ALBERT

Description: Secreted phosphoprotein 24 kDa (Spp24) is an apatite- and BMP/TGF-β cytokine-binding phosphoprotein found in serum and many tissues, including bone. N-terminally intact degradation products ranging in size from 14 kDa to 23 kDa have been found in bone. The cleavage sites in Spp24 that produce these short forms have not been definitively identified, and the biological activities and mechanisms of action of Spp24 and its degradation products have not been fully elucidated. We found that the C-terminus of Spp24 is labile to proteolysis by furin, kallikrein, lactoferrin, and trypsin, indicating that both extracellular and intracellular proteolytic events could account for the generation of biologically-active Spp18, Spp16, and Spp14. We determined the effects of these truncation products on kinase-mediated signal transduction, gene expression, and osteoblastic differentiation in W-20-17 bone marrow stromal cells cultured in basal or pro-osteogenic media. After culturing for five days, all forms inhibited BMP-2-stimulated osteoblastic differentiation, assessed as induction of alkaline phosphatase activity, in basal, but not pro-osteogenic media. After 10 days, they also inhibited BMP-2-stimulated mineral deposition in pro-osteogenic media. Spp24 had no effect on Erk1/2 phosphorylation, but Spp18 stimulated short-term Erk1/2, MEK 1/2, and p38 phosphorylation. Pertussis toxin and a MEK1/2 inhibitor ablated Spp18-stimulated Erk 1/2 phosphorylation, indicating a role for G i proteins and MEK1/2 in the Spp18-stimulated Erk1/2 phosphorylation cascade. Truncation products, but not full-length Spp24, stimulated RUNX2, ATF4, and CSF1 transcription. This suggests that Spp24 truncation products have effects on osteoblastic differentiation mediated by kinase pathways that are independent of exogenous BMP/TGF-β cytokines. This article is protected by copyright. All rights reserved

Description: Similar to phosphorylation, transient conjugation of ubiquitin to target proteins (ubiquitination) mediated by the concerted action of ubiquitin ligases and de-ubiquitinating enzymes (DUBs) can affect substrate function. As obligate intracellular parasites, viruses rely on different cellular pathways for their own replication and the well conserved ubiquitin conjugating/de-conjugating system is not an exception. Viruses not only usurp the host proteins involved in the ubiquitination/de-ubiquitination process, but they also encode their own ubiquitin ligases and DUBs. Here we report that an N-terminal variant of the herpes simplex virus (HSV) type-1 large tegument protein VP1/2 (VP1/2 1–767 ), encompassing an active DUB domain (herpesvirus tegument ubiquitin specific protease, htUSP), and TSG101, a component of the endosomal sorting complex required for transport (ESCRT)-I, functionally interact. In particular, VP1/2 1–767 modulates TSG101 ubiquitination and influences its intracellular distribution. Given the role played by the ESCRT machinery in crucial steps of both cellular pathways and viral life cycle, the identification of TSG101 as a cellular target for the HSV-1 specific de-ubiquitinating enzyme contributes to the clarification of the still under debate function of viral encoded DUBs highly conserved throughout the Herpesviridae family. This article is protected by copyright. All rights reserved

Description: Sarcopenia and osteoporosis have recently been noted for their relationship with locomotive syndrome and increased number of older people. Sarcopenia is defined by decreased muscle mass and impaired muscle function, which may be associated with frailty. Several clinical data have indicated that increased muscle mass is related to increased bone mass and reduced fracture risk. Genetic, endocrine and mechanical factors as well as inflammatory and nutritional states concurrently affect muscle tissues and bone metabolism. Several genes, including myostatin and α-actinin 3, have been shown in a genome-wide association study (GWAS) to be associated with both sarcopenia and osteoporosis. Vitamin D, growth hormone and testosterone as well as pathological disorders, such as an excess in glucocorticoid and diabetes, affect both muscle and bone. Basic and clinical research of bone metabolism and muscle biology suggests that bone interacts with skeletal muscle via signaling from local and humoral factors in addition to their musculoskeletal function. However, the physiological and pathological mechanisms related to muscle and bone interactions remain unclear. We found that Tmem119 may play a critical role in the commitment of myoprogenitor cells to the osteoblast lineage. We also reported that osteoglycin and FAM5C might be muscle-derived humoral osteogenic factors. Other factors, including myostatin, osteonectin, insulin-like growth factor I, irisin and osteocalcin, may be associated with the interactions between muscle tissues and bone metabolism. This article is protected by copyright. All rights reserved

Description: The transition to a pathological phenotype such as Barrett's esophagus occurs via induction of resistance upon repeated contact with gastric refluxate in esophagus. This study examined the molecular changes within normal esophageal epithelial cells (EECs) under short-term acid loading and the role of these changes in defensive resistance against acidic cytotoxicity. After primary cultured EECs were exposed to pH 4-acidified medium (AM4), cell viability was determined by the MTT assay. Reactive oxygen species (ROS) and NAD(P)H oxidase (NOX) activity were measured. Activation of the mitogen-activated protein kinases (MAPKs) MEK/ERK1/2, p38 and JNK; phosphoinositol-3-kinase (PI3K)/Akt, and nuclear factor-kappa B (NF-κB) were detected by Western blot analysis or immunofluorescence staining. AM4 incubation induced intracellular ROS generation accompanied by increase in NOX activity, which was further increased by Na + /H + exchange-1 (NHE1)-dependent inhibition but was prevented by inhibition of NOX or mitochondria complex I. AM4 also induced phosphorylation of MEK/ERK1/2, p38 MAPK, PI3K/Akt, and nuclear translocation of NF-κB, and all these effects, except for p38 MAPK phosphorylation, were abolished by inhibition of ROS. ROS-dependent PI3K/Akt activation, which mediates NF-κB nuclear translocation, was inhibited by protein tyrosine kinase (PTK) inhibitors and NHE1-specific inhibitor. All inhibitors of NHE, ROS, PTK, PI3K or NF-κB further decreased AM4-induced cell viability. Acid loading in the presence of NHE1-dependent protection induced ROS generation by activating NOX and mitochondria complex I, which stimulated PTK/PI3K/Akt/NF-κB-dependent survival in EEC. Our data indicate that normal EEC initially respond to acid loading through intrinsic survival activation. This article is protected by copyright. All rights reserved

Description: Key Points Net COS production in wheat field soil and roots scaled linearly by temperature Sterilized wheat field soil demonstrated abiotic COS production mechanism Lab observations of COS fluxes from soil confirmed previous field measurements

Description: Biological invasions of aquatic systems disrupt ecological communities, and cause major changes in diversity and ecosystem function. The Laurentian Great Lakes of North America have been dramatically altered by such invasions, especially zebra (Dreissena polymorpha) and quagga (D. rostriformis bugensis) mussels. Responses to mussel invasions have included increased water clarity, and decreased chlorophyll and phytoplankton abundance. Although not all systems have responded similarly, in general, mussels have changed nutrient dynamics and physical habitat conditions. Therefore examination of different impacts can help us further understand mechanisms that underlie ecosystem responses to biological invasions. To aid our understanding of ecosystem impacts, we sampled established locations along a well-studied trophic gradient in Green Bay, Lake Michigan, after the 1993 zebra mussel invasion. A strong trophic gradient remained during the period sampled after the mussel invasion (2000–2012). However, mean summer chlorophyll increased and other measures of phytoplankton biomass (microscope and electronic cell counting) did not change significantly. Multivariate analyses of phytoplankton community structure demonstrate a significant community shift after the invasion. Cyanobacteria increased in dominance, with Microcystis becoming the major summer taxon in lower Green Bay. Diatom diversity and abundance also increased and Chlorophyta became rare. Phytoplankton responses along the trophic gradient of Green Bay to zebra mussel invasion highlight the importance of mussel effects on nutrient dynamics and phytoplankton diversity and function.

Description: The CRIPSR-Cas9 system consists of a site-specific, targetable DNA nuclease that holds great potential in gene editing and genome-wide screening applications. In order to apply the CRISPR-Cas9 system to these assays successfully, the rate at which Cas9 induces DNA breaks at undesired loci must be understood. We characterised the rate of Cas9 off-target activity in typical Cas9 experiments in two human and one mouse cell lines. We analysed the Cas9 cutting activity of 12 gRNAs in both their targeted sites and ∼90 predicted off-target sites per gRNA. In a Cas9-based knock-out experiment, gRNAs induced detectable Cas9 cutting activity in all on-target sites and in only a few off-target sites genome-wide in human 293FT, human iPS cells and mouse ES cells. Both the cutting rates and DNA repair patterns were highly correlated between the two human cell lines in both on-target and off-target sites. In clonal Cas9 cutting analysis in mouse ES cells, bi-allelic Cas9 cutting was observed with low off-target activity. Our results show that off-target activity of Cas9 is low and predictable by the degree of sequence identity between the gRNA and a potential off-target site. Off-target Cas9 activity can be minimized by selecting gRNAs with few off-target sites of near-complementarity. genesis 00:00–00, 2014. © 2014 Wiley Periodicals, Inc.

Description: In this study we first present updated riverine total alkalinity (TA) loads to the various Baltic Sea sub-basins, based on monthly measurements in 82 of the major rivers that represent 85% of the total runoff. Simulations in the coupled physical-biogeochemical BALTSEM model show that these river loads together with North Sea water inflows are not sufficient to reproduce observed TA concentrations in the system, demonstrating the large influence from internal sources. Budget calculations indicate that the required internal TA generation must be similar to river loads in magnitude. The non-riverine source in the system amounts to about 2.4 mmol m -2 d -1 on average. We argue here that the majority of this source is related to denitrification together with unresolved sediment processes such as burial of reduced sulfur and/or silicate weathering. This hypothesis is supported by studies on sediment processes on a global scale, and also by data from sediment cores in the Baltic Sea. In a model simulation with all internal TA sources and sinks switched on, the net absorption of atmospheric CO 2 increased by 0.78 mol C m -2 y -1 compared to a simulation where TA was treated as a passive tracer. Our results clearly illustrate how pelagic TA sources together with anaerobic mineralization in coastal sediments generate a significant carbon sink along the aquatic continuum, mitigating CO 2 evasions from coastal and estuarine systems.

Description: ABSTRACT The recent advances on ascidian pigment sensory organ development and function represent a fascinating platform to get insight on the basic programs of chordate eye formation. This review aims to summarize current knowledge, at the structural and molecular levels, on the two main building blocks of ascidian light sensory organs, i.e . pigment cells and photoreceptor cells. The unique features of these structures (e.g. simplicity and well characterized cell lineage) are indeed making it possible to dissect the developmental programs at single cell resolution and will soon provide a panel of molecular tools to be exploited for a deep developmental and comparative-evolutionary analysis. © 2014 Wiley Periodicals, Inc.

Description: Dynamic interactions with DNA allow replication protein A to direct single-stranded DNA-intermediates into different pathways for synthesis or repair. On pages 1156–1161 , Chen and Wold review recent discoveries that show that replication protein A (RPA), the major eukaryotic single-stranded DNA-binding protein, binds DNA dynamically and that this is important for correct processing of DNA intermediates. The cover shows a model of human RPA interacting with ssDNA based on the known structures of the domains of human RPA and the structure of Ustalago RPA bound to DNA. The three subunits of RPA are shown in blue (RPA1), red (RPA2), and green (RPA3) with ssDNA shown in black.

Description: The aim of this study was to investigate the involvement of exchange proteins directly activated by cyclic adenosine (ADO) monophosphate (EPAC) in 4-mer hyaluronan (HA) oligosaccharide-induced inflammatory response in mouse normal synovial fibroblasts (NSF). Treatment of NSF with 4-mer HA increased Toll-like receptor-4, TNF-alpha and IL-1beta mRNA expression and of the related proteins, as well as nuclear factor kappaB (NF-kB) activation. Addition to NSF, previously stimulated with 4-mer HA oligosaccharides, of ADO significantly reduced NF-kB activation, TNF-alpha and IL-1beta expression. The pre-treatment of NSF with cyclic ADO monophosphate and/or PKA and/or EPAC-specific inhibitors significantly inhibited the anti-inflammatory effect exerted by ADO. In particular, the EPAC inhibitor reduced the ADO effect to a major extent than the PKA inhibitor. These results mean that both PKA and EPAC pathways are involved in ADO-induced NF-kB inhibition although EPAC seems to be more involved than PKA. Copyright © 2014 John Wiley & Sons, Ltd.

Description: Temporally controlled induction of gene expression is a useful technique for analyzing gene function. To make such a technique possible in Ciona intestinalis embryos, we employed the cis -regulatory region of the heat-shock protein 70 (HSP70) gene Ci-HSPA1/6/7-like for heat-inducible gene expression in C. intestinalis embryos. We showed that Ci-HSPA1/6/7-like becomes heat shock-inducible by the 32-cell stage during embryogenesis. The 5'-upstream region of Ci-HSPA1/6/7-like , which contains heat-shock elements indispensable for heat-inducible gene expression, induces the heat shock-dependent expression of a reporter gene in the whole embryo from the 32-cell to the middle gastrula stages and in progressively restricted areas of embryos in subsequent stages. We assessed the effects of heat-shock treatments in different conditions on the normality of embryos and induction of transgene expression. We evaluated the usefulness of this technique through overexpression experiments on the well-characterized, developmentally relevant gene, Ci-Bra , and showed that this technique is applicable for inferring the gene function in C. intestinalis . © 2014 Wiley Periodicals, Inc.

Description: Osteolytic bone diseases are characterized by excessive osteoclast formation and activation. Protein kinase C (PKC)-dependent pathways regulate cell growth, differentiation and apoptosis in many cellular systems, and have been implicated in cancer development and osteoclast formation. A number of PKC inhibitors with anti-cancer properties have been developed, but whether they might also influence osteolysis (a common complication of bone invading cancers) is unclear. We studied the effects of the PKC inhibitor compound, GF109203X on osteoclast formation and activity, processes driven by receptor activator of NFκB ligand (RANKL). We found that GF109203X strongly and dose dependently suppresses osteoclastogenesis and osteoclast activity in RANKL-treated primary mouse bone marrow cells. Consistent with this GF109203X reduced expression of key osteoclastic genes, including cathepsin K, calcitonin receptor, tartrate resistant acid phosphatase (TRAP) and the proton pump subunit V-ATPase-d2 in RANKL-treated primary mouse bone marrow cells. Expression of these proteins is dependent upon RANKL-induced NF-κB and NFAT transcription factor actions; both were reduced in osteoclast progenitor populations by GF109203X treatment, notably NFATc1 levels. Furthermore, we showed that GF109203X inhibits RANKL-induced calcium oscillation. Together, this study shows GF109203X may block osteoclast functions, suggesting that pharmacological blockade of PKC-dependent pathways has therapeutic potential in osteolytic diseases. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.

Description: The sesquiterpene lactone Parthenolide (PN) exerted a cytotoxic effect on MDA-MB231 cells, a triple-negative breast cancer (TNBC) cell line, but its effectiveness was scarce when employed at low doses. This represents an obstacle for a therapeutic utilization of PN. In order to overcome this difficulty we associated to PN the suberoylanilide hydroxamic acid (SAHA), an histone deacetylase inhibitor. Our results show that SAHA synergistically sensitized MDA-MB231 cells to the cytotoxic effect of PN. It is noteworthy that treatment with PN alone stimulated the survival pathway Akt/mTOR and the consequent nuclear translocation of Nrf2, while treatment with SAHA alone induced autophagic activity. However when the cells were treated with SAHA/PN combination, SAHA suppressed PN effect on Akt/mTOR/Nrf2 pathway, while PN reduced the prosurvival autophagic activity of SAHA. In addition SAHA/PN combination induced GSH depletion, fall in Δψm, release of cytochrome c, activation of caspase 3 and apoptosis. Finally we demonstrated that combined treatment maintained both hyperacetylation of histones H3 and H4 induced by SAHA and down-regulation of DNMT1 expression induced by PN. Inhibition of the DNA-binding activity of NF-kB, which is determined by PN, was also observed after combined treatment. In conclusion, combination of PN to SAHA inhibits the cytoprotective responses induced by the single compounds, but does not alter the mechanisms leading to the cytotoxic effects. Taken together our results suggest that this combination could be a candidate for TNBC therapy. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.

Description: There have been few assessments of the performance of alternative resistance surfaces, and little is known about how connectivity modeling approaches differ in their ability to predict organism movements. In this paper, we evaluate the performance of four connectivity modeling approaches applied to two resistance surfaces in predicting the locations of highway crossings by American black bears in the northern Rocky Mountains, USA. We found that a resistance surface derived directly from movement data greatly outperformed a resistance surface produced from analysis of genetic differentiation, despite their heuristic similarities. Our analysis also suggested differences in the performance of different connectivity modeling approaches. Factorial least cost paths appeared to slightly outperform other methods on the movement-derived resistance surface, but had very poor performance on the resistance surface obtained from multi-model landscape genetic analysis. Cumulative resistant kernels appeared to offer the best combination of high predictive performance and sensitivity to differences in resistance surface parameterization. Our analysis highlights that even when two resistance surfaces include the same variables and have a high spatial correlation of resistance values, they may perform very differently in predicting animal movement and population connectivity.

Description: The tenet that ecological opportunity drives adaptive diversification has been central to theories of speciation since Darwin, yet no widely accepted definition or mechanistic framework for the concept currently exists. We propose a definition for ecological opportunity that provides an explicit mechanism for its action. In our formulation, ecological opportunity refers to environmental conditions that both permit the persistence of a lineage within a community, as well as generate divergent natural selection within that lineage. Thus, ecological opportunity arises from two fundamental elements: (1) niche availability, the ability of a population with a phenotype previously absent from a community to persist within that community and (2) niche discordance, the diversifying selection generated by the adaptive mismatch between a population's niche-related traits and the newly encountered ecological conditions. Evolutionary response to ecological opportunity is primarily governed by (1) spatiotemporal structure of ecological opportunity, which influences dynamics of selection and development of reproductive isolation and (2) diversification potential, the biological properties of a lineage that determine its capacity to diversify. Diversification under ecological opportunity proceeds as an increase in niche breadth, development of intraspecific ecotypes, speciation, and additional cycles of diversification that may themselves be triggered by speciation. Extensive ecological opportunity may exist in depauperate communities, but it is unclear whether ecological opportunity abates in species-rich communities. Because ecological opportunity should generally increase during times of rapid and multifarious environmental change, human activities may currently be generating elevated ecological opportunity – but so far little work has directly addressed this topic. Our framework highlights the need for greater synthesis of community ecology and evolutionary biology, unifying the four major components of the concept of ecological opportunity. The tenet that ecological opportunity drives adaptive diversification has been central to theories of speciation since Darwin, yet there is currently no widely accepted definition or mechanistic framework for the concept. We propose an explicit definition and mechanism for ecological opportunity that includes its two fundamental elements: (1) niche availability, the opportunity for initial persistence of a new phenotype within a community, and (2) niche discordance, the rearrangement of ecological constraints that generates diversifying selection.

Description: Santín et al. (2014) report the conversion of different boreal forest biomass pools to pyrogenic organic matter (PyOM) during a forest fire, and suggest that ~100 Tg C y −1 may be converted to PyOM in boreal forests globally. They further suggest that PyOM formation represents a missing C sink. The phrase ‘missing C sink’ derives from a lack of closure in the atmospheric C budget. Approximately ⅓ of the CO 2 emitted to the atmosphere via burning of fossil fuels and land use change cannot be accounted for after oceanic uptake and atmospheric accumulations are tallied (Schlesinger and Bernhardt 2013). This article is protected by copyright. All rights reserved.

Description: A significant part of the soil organic carbon that is eroded in uplands is deposited and buried in colluvial settings. Understanding the fate of this deposited soil organic carbon (SOC) is of key importance for the understanding of the role of (accelerated) erosion in the global C cycle: the residence time of the deposited carbon will determine if, and for how long, accelerated erosion due to human disturbance will induce sequestration of SOC from the atmosphere to the soil. Experimental studies may provide useful information, but, given the time scale under consideration, the response of the colluvial SOC can only be simulated using numerical models which need careful calibration using field data. In this study, we present a depth explicit SOC model (ICBM-DE) including soil profile evolution due to sedimentation to simulate the long-term C dynamics in colluvial soils. The SOC profile predicted by our model is in good agreement with field observations. The C burial efficiency (the ratio of current C content of the buried sediments to the original C content at the time of sedimentation) of deposited sediments exponentially decreases with time and gradually reached an equilibrium value. This equilibrium C burial efficiency is positively correlated with the sedimentation rate. The sedimentation rate is crucial for the long-term dynamics of the deposited SOC as it controls the time that buried sediments spend at a given soil depth, thereby determining its temporal evolution of C input and decomposition rate during the burial process: C input and decomposition rate vary with depth due to the vertical variation of root distribution and soil environmental factors such as (but not limited to) humidity, temperature and aeration. The model demonstrates that, for the profiles studied, it takes ca. 300 yr for the buried SOC to lose half of its C load. It would also take centuries for the SOC accumulated in colluvial soils over the past decades due to soil redistribution under mechanized agriculture to be released to the atmosphere after the application of soil conservation measures such as conservation tillage.

Description: Massive DNA sequencing has significantly increased the amount of data available for population genetics and molecular ecology studies. However, the parallel computation of simple statistics within and between populations from large panels of polymorphic sites is not yet available, making the exploratory analyses of a set or subset of data a very laborious task. Here, we present 4P (parallel processing of polymorphism panels), a stand-alone software program for the rapid computation of genetic variation statistics (including the joint frequency spectrum) from millions of DNA variants in multiple individuals and multiple populations. It handles a standard input file format commonly used to store DNA variation from empirical or simulation experiments. The computational performance of 4P was evaluated using large SNP (single nucleotide polymorphism) datasets from human genomes or obtained by simulations. 4P was faster or much faster than other comparable programs, and the impact of parallel computing using multicore computers or servers was evident. 4P is a useful tool for biologists who need a simple and rapid computer program to run exploratory population genetics analyses in large panels of genomic data. It is also particularly suitable to analyze multiple data sets produced in simulation studies. Unix, Windows, and MacOs versions are provided, as well as the source code for easier pipeline implementations. 4P is a new computer program for parallel computation of basic population genetics statistics (including the joint allele frequency spectrum) from large panel of SNPs. 4P is fast and simple to use, read files with genomic data saved in classical formats, do not require ad hoc scripts, and it very fast with servers but also when multi-core stand-alone computers are used. It is particularly suitable to analyse multiple data sets produced in simulation studies.

Description: Publication date: Available online 13 December 2014 Source: FEBS Open Bio Author(s): Tadaho Nakamura , Takeo Yoshikawa , Fumito Naganuma , Attayeb Mohsen , Tomomitsu Iida , Yamato Miura , Akira Sugawara , Kazuhiko Yanai Pancreatic α-cells secrete glucagon to maintain energy homeostasis. Although histamine has an important role in energy homeostasis, the expression and function of histamine receptors in pancreatic α-cells remains unknown. We found that the histamine H 3 receptor (H 3 R) was expressed in mouse pancreatic α-cells and αTC1.6 cells, a mouse pancreatic α-cell line. H 3 R inhibited glucagon secretion from αTC1.6 cells by inhibiting an increase in intracellular Ca 2+ concentration. We also found that immepip, a selective H 3 R agonist, decreased serum glucagon concentration in rats. These results suggest that H 3 R modulates glucagon secretion from pancreatic α-cells.

Description: Tumor cells display different bioenergetic profiles when compared to normal cells. In the present work we showed metabolic reprogramming by means of inhibitors of histone deacetylase (HDACis), sodium butyrate and trichostatin A in breast cancer cells representing different stages of aggressiveness and metabolic profile. When testing the effect of NaB and TSA on viability of cells, it was shown that non-tumorigenic MCF-10A cells were less affected by increasing doses of the drugs than the tumorigenic, hormone dependent, tightly cohesive MCF-7, T-47D and the highly metastatic triple-negative MDA-MB 231 cells. T-47D cells were the most sensitive to treatment with both, NaB and TSA. Experiments measuring anchorage- independent growth of tumor cells showed that MCF-7, T-47D, and MDA-MB-231 cells were equally sensitive to the treatment with NaB. The NaB induced an attenuation of glycolysis, reflected by a decrease in lactate release in MCF-7 and T47D lines. Pyruvate kinase activity was significantly enhanced by NaB in MDA-MB-231 cells only. In contrast, the inhibitor enhanced lactate dehydrogenase activity specifically in T-47 D cells. Glucose-6-phosphate dehydrogenase activity was shown to be differentially modulated by NaB in the cell lines investigated: the enzyme was inhibited in MCF-7 cells, whereas in T-47D and MDA-MB-231 cells, G6PDH was activated. NaB and TSA were able to significantly increase the oxygen consumption by MDA-MB-231 and T-47D cells. Collectively the results show that epigenetic changes associated to acetylation of proteins in general affect the energy metabolism in all cancer cell lines and that mitochondria may occupy a central role in metastasis. This article is protected by copyright. All rights reserved

Description: Genomic and trascriptomic profiling has recently contributed details to the characterization of luminal B breast cancer. We explored the contribution of anthropometric, metabolic and molecular determinants to the multifaceted heterogeneity of this breast cancer subtype, with a specific focus on the association between body mass index (BMI), pre-treatment fasting glucose, hormone receptors and expression of human epidermal growth factor receptor 2 (HER2). Extensively annotated specimens were obtained from 154 women with luminal B breast cancer diagnosed at two Italian comprehensive cancer centres. Participants' characteristics were descriptively analyzed overall and by HER2 status (positive vs negative). BMI (〈25 vs ≥25), pre-treatment fasting glucose (〈median value of 94 mg/dl vs ≥94) and percentage of hormone receptors were tested for association with HER2 expression in regression models. In univariate models, BMI, fasting glucose and, at a lesser extent, percentage of estrogen receptors (ER) were significantly and inversely associated with HER2 expression (OR: 0.32, 95%CI: 0.16-0.66; 0.43, 0.23-0.0.82; 0.96, 0.94-0.97, respectively). The multivariate models confirmed the protective role of BMI and ER on HER2 expression, with luminal B HER2 positive patients being significantly less frequent among women within the highest category of BMI and percentage expression of ER compared with their counterparts (OR: 0.22, 95%CI: 0.09-0.53; 0.95, 0.93-0.97). In conclusions, BMI and percentage of ER representation are inversely associated with HER2 expression in luminal B breast cancers. Upon confirmatory findings, this might help identify patient subgroups who may best benefit from the use of interventions targeting insulin resistance in well depicted breast cancer scenarios. This article is protected by copyright. All rights reserved

Description: Citations published in online supplementary material (OSM) are invisible to search engines used to calculate citation counts, potentially negatively impacting popular performance indices and journal rankings that rely on citation counts for quantification. To quantify the number of citations that are “lost” in OSM, we conducted a systematic survey of supplementary citation practices in four high-ranking, society-run journals from two geographical locations (Europe and North America). In 2012, 6% of all citations were only included in the OSM and were therefore not included in citation counts. We found a significant increase in the number of references invisible to citation counting services over the last two decades. A solution to this problem is urgently required and could include journal indexing of citations in OSM or the inclusion of all references in the main text. Citations published in online supplementary material (OSM) are invisible to search engines used to calculate citation counts, potentially negatively impacting popular performance indices and journal rankings that rely on citation counts for quantification. We conducted a systematic survey of historical supplementary citation practices in four journals from two geographical locations (Europe and North America) and found a significant increase in the number of references invisible to citation counting services over the last two decades. A solution to this problem is urgently required and could include journal indexing of citations in OSM or the inclusion of all references in the main text.

Description: Histone deacetylases (HDACs) play a crucial role in the regulation of gene expression through remodeling of chromatin structures. However, the molecular mechanisms involved in this event remain unknown. In this study, we sought to examine whether HDAC inhibition-mediated protective effects involved HDAC4 sumoylation, degradation, and the proteasome pathway. Isolated neonatal mouse ventricular myocytes (NMVM) and H9c2 cardiomyoblasts were subjected to 48 hours of hypoxia (H) (1% O 2 ) and 2 hours of reoxygenation (R). Treatment of cardiomyocytes with trichostatin A (TSA) attenuated H/R-elicited injury, as indicated by a reduction of lactate dehydrogenase (LDH) leakage, an increase in cell viability, and decrease in apoptotic positive cardiomyocytes. MG132, a potent proteasome pathway inhibitor, abrogated TSA-induced protective effects, which was associated with the accumulation of ubiquitinated HDAC4. NMVM transduced with adenoviral HDAC4 led to an exaggeration of H/R-induced injury. TSA treatment resulted in a decrease in HDAC4 in cardiomyocytes infected with adenoviral HDAC4, and HDAC4-induced injury was attenuated by TSA. HDAC inhibition resulted in a significant reduction in reactive oxygen species (ROS) in cardiomyoblasts exposed to H/R, which was attenuated by blockade of the proteasome pathway. Cardiomyoblasts carrying wild type and sumoylation mutation (K559R) were established to examine effects of HDAC4 sumoylation and ubiquitination on H/R injury. Disruption of HDAC4 sumoylation brought about HDAC4 accumulation and impairment of HDAC4 ubiquitination in association with enhanced susceptibility of cardiomyoblasts to H/R. Taken together, these results demonstrated that HDAC inhibition stimulates proteasome dependent degradation of HDAC4, which is associated with HDAC4 sumoylation to induce these protective effects. This article is protected by copyright. All rights reserved

Description: Tree-ring analysis is often used to assess long-term trends in tree growth. A variety of growth-trend detection methods (GDMs) exist to disentangle age/size trends in growth from long-term growth changes. However, these detrending methods strongly differ in approach, with possible implications for their output. Here we critically evaluate the consistency, sensitivity, reliability and accuracy of four most widely used GDMs: Conservative Detrending applies mathematical functions to correct for decreasing ring-widths with age; Basal Area Correction transforms diameter into basal-area growth; Regional Curve Standardization detrends individual tree-ring series using average age/size trends; and Size Class Isolation calculates growth trends within separate size classes. First, we evaluated whether these GDMs produce consistent results applied to an empirical tree-ring dataset of Melia azedarach , a tropical tree species from Thailand. Three GDMs yielded similar results – a growth decline over time – but the widely used Conservative Detrending method did not detect any change. Second, we assessed the sensitivity (probability of correct growth trend detection), reliability (1- probability of detecting false trends), and accuracy (whether the strength of imposed trends is correctly detected) of these GDMs, by applying them to simulated growth trajectories with different imposed trends: no trend, strong trends (-6% and +6% change per decade), and weak trends (-2%, +2%). All methods except Conservative Detrending, showed high sensitivity, reliability and accuracy to detect strong imposed trends. However, these were considerably lower in the weak or no-trend scenarios. Basal Area Correction showed good sensitivity and accuracy, but low reliability, indicating uncertainty of trend-detection using this method. Our study reveals that the choice of GDM influences results of growth-trend studies. We recommend applying multiple methods when analysing trends and encourage performing sensitivity and reliability analysis. Finally, we recommend Size Class Isolation and Regional Curve Standardization, as these methods showed highest reliability to detect long-term growth trends. This article is protected by copyright. All rights reserved.

Description: Surface-to-atmosphere emissions of dimethyl sulfide (DMS) may impact global climate through the formation of gaseous sulfuric acid, which can yield secondary sulfate aerosols and contribute to new particle formation. While oceans are generally considered the dominant source of DMS, a shortage of ecosystem observations prevents an accurate analysis of terrestrial DMS sources. Using mass spectrometry, we quantified ambient DMS mixing ratios within and above a primary rainforest ecosystem in the central Amazon Basin in real-time (2010–2011) and at high vertical resolution (2013–2014). Elevated but highly variable DMS mixing ratios were observed within the canopy, showing clear evidence of a net ecosystem source to the atmosphere during both day and night in both the dry and wet seasons. Periods of high DMS mixing ratios lasting up to 8 hours (up to 160 ppt) often occurred within the canopy and near the surface during many evenings and nights. Daytime gradients showed mixing ratios (up to 80 ppt) peaking near the top of the canopy as well as near the ground following a rain event. The spatial and temporal distribution of DMS suggests that ambient levels and their potential climatic impacts are dominated by local soil and plant emissions. A soil source was confirmed by measurements of DMS emission fluxes from Amazon soils as a function of temperature and soil moisture. Furthermore, light and temperature dependent DMS emissions were measured from seven tropical tree species. Our study has important implications for understanding terrestrial DMS sources and their role in coupled land-atmosphere climate feedbacks.

Description: Question Hemiparasitic plants can profoundly affect the structure of the community in which they occur, mainly due to parasitic suppression of hosts. As a consequence, non-host species have the opportunity to colonize resulting gaps. In contrast to most grassland species, hemiparasites are generally short-lived and can reach high densities; as a consequence, vegetation gaps are left after their death. These gaps form microsites more suitable for seed germination and therefore might increase recruitment of other species. Which species suffer or profit from parasitism and is there a positive effect on seedling establishment? Location Semi-natural grasslands in northern Belgium. Methods We selected two hemiparasitic plant species from contrasting vegetation types: Rhinanthus angustifolius growing in mesotrophic grassland and Pedicularis sylvatica growing in oligotrophic heath-grassland. A weeding experiment was set up at six sites in which the hemiparasite was repeatedly removed in half of the plots during three growing seasons. The abundance change of individual species was compared between weeded and control plots. After the second growing season, seeds of up to ten species were added. The number of seedlings in the third year was then compared between weeded and control plots. Results Rhinanthus removal significantly affected the abundance of species relative to control plots, both positively and negatively, and decreased the species evenness. Pedicularis removal only increased the abundance of some species. Only Juncaceae (but no other graminoid families) increased after Rhinanthus and Pedicularis weeding, and there was considerable variation within functional groups. Moreover, based on our observations, we propose as a new hypothesis that species with persistent clonal spread are more vulnerable to parasitism. Once attached, resources are potentially drawn from a whole network of interconnected ramets. Finally, only half of the sown species successfully established seedlings; hemiparasite removal had a significantly negative effect on seedling number for two of these species. Conclusions Effects of hemiparasites on species differ considerably, also within functional groups. Persistent clonal spread emerges as an important plant trait determining vulnerability to hemiparasites. Finally, our results suggest that hemiparasitic plants might have a limited positive effect on seedling establishment in these semi-natural grasslands where chances for successful establishment were shown to be low. We studied the effect of 2 grassland hemiparasites on individual species in the established phase and in the establishment phase. Based on our observations, we propose an exciting new hypothesis: that species with persistent clonal spread are more vulnerable to parasitism because once attached, resources are potentially drawn from a whole network of interconnected ramets.

Description: (http://onlinelibrary.wiley.com/doi/10.1111/gcb.12697/full) The above article, published online on 18 August 2014 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the authors, Dr Melanie Harsch and Associate Professor Janneke Hille Ris Lambers, journal Editor‐in‐Chief, Professor Stephen Long, and John Wiley & Sons Ltd. The retraction has been agreed for the following reasons: a coding error affected the results and therefore invalidated the broad‐scale conclusions presented in the article. The article presented broad‐scale patterns of species distribution shifts in response to recent climate change. Unfortunately, it has since been found that one approach used to account for sampling bias, the null model approach, was affected by the coding error. Following the identification of the coding error, we are therefore retracting the article. We thank Drs Adam Wolf and William Anderegg for bringing this issue to our attention. Reference Harsch MA, Hille Ris Lambers J (2014) Species distributions shift downward across western North America. Global Change Biology. doi: 10.1111/gcb.12697.

Description: Serine protease inhibitors (PIs) have been described in many plant species and are universal throughout the plant kingdom, where trypsin inhibitors is the most common type. In the present study, trypsin and chymotrypsin inhibitory activity was detected in the seed flour extracts of 13 selected cultivars/accessions of cowpea. Two cowpea cultivars, Cream7 and Buff, were found to have higher trypsin and chymotrypsin inhibitory potential compared to other tested cultivars for which they have been selected for further purification studies using ammonium sulfate fractionation and DEAE-Sephadex A-25 column. Cream7-purified proteins showed two bands on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) corresponding to molecular mass of 17.10 and 14.90 kDa, while the purified protein from Buff cultivar showed a single band corresponding mass of 16.50 kDa. The purified inhibitors were stable at temperature below 60°C and were active at wide range of pH from 2 to 12. The kinetic analysis revealed noncompetitive type of inhibition for both inhibitors against both enzymes. The inhibitor constant ( K i ) values suggested high affinity between inhibitors and enzymes. Purified inhibitors were found to have deep and negative effects on the mean larval weight, larval mortality, pupation, and mean pupal weight of Spodoptera littoralis , where Buff PI was more effective than Cream7 PI. It may be concluded that cowpea PI gene(s) could be potential insect control protein for future studies in developing insect-resistant transgenic plants.

Description: The vitamin A-redox hypothesis provides an explanation for honest signaling of phenotypic quality by carotenoid-dependent traits. A key aspect of the vitamin A-redox hypothesis, applicable to both yellow and red coloration, is the hypothesized negative feedback of tightly regulated Vitamin A plasma levels on the enzyme responsible for sequestering both Vitamin A and carotenoids from the gut. We performed a meta-analysis and find that vitamin A levels are positively related to carotenoid plasma levels ( r  =   0.50, P  = 0.0002). On the basis of this finding and further theoretical considerations, we propose that the vitamin A-redox hypothesis is unlikely to explain carotenoid-dependent honest signaling. The vitamin A-redox hypothesis provides an explanation for honest signaling of phenotypic quality by carotenoid-dependent traits. We performed a meta-analysis and find that vitamin A levels are positively related to carotenoid plasma levels ( r  = 0.50, P  = 0.0002). On the basis of this finding and further theoretical considerations we propose that the vitamin A-redox hypothesis is unlikely to explain carotenoid-dependent honest signaling.

Description: Seaweed beds play a key role in providing essential habitats and energy to coastal areas, with enhancements in productivity and biodiversity and benefits to human societies. However, the spatial extent of seaweed beds around Japan has decreased due to coastal reclamation, water quality changes, rising water temperatures, and heavy grazing by herbivores. Using monthly mean sea surface temperature (SST) data from 1960 to 2099 and SST-based indices, we quantitatively evaluated the effects of warming seawater on the spatial extent of suitable versus unsuitable habitats for temperate seaweed Ecklonia cava, which is predominantly found in southern Japanese waters. SST data were generated using the most recent multiple climate projection models and emission scenarios (the Representative Concentration Pathways or RCPs) used in the Coupled Model Intercomparison Project phase 5 (CMIP5). In addition, grazing by Siganus fuscescens, an herbivorous fish, was evaluated under the four RCP simulations. Our results suggest that continued warming may drive a poleward shift in the distribution of E. cava, with large differences depending on the climate scenario. For the lowest emission scenario (RCP2.6), most existing E. cava populations would not be impacted by seawater warming directly but would be adversely affected by intensified year-round grazing. For the highest emission scenario (RCP8.5), previously suitable habitats throughout coastal Japan would become untenable for E. cava by the 2090s, due to both high-temperature stress and intensified grazing. Our projections highlight the importance of not only mitigating regional warming due to climate change, but also protecting E. cava from herbivores to conserve suitable habitats on the Japanese coast. In this study, we quantitatively evaluated the effects of warming seawater on the spatial extent of suitable versus unsuitable habitats for temperate seaweed using the most recent multiple climate projection models and emission scenarios. Our results suggest that continued warming may drive a poleward shift of temperate seaweeds, with large differences depending on the severity of warming, and most of seaweeds' populations would be exposed to intensified grazing by herbivores, even if it were possible to mitigate regional warming.

Description: Endothelial cells play a major role in the initiation and perpetuation of the inflammatory process in health and disease, including their pivotal role in leukocyte recruitment. The role of pro-inflammatory transcription factors in this process has been well-described, including NF-κB. However, much less is known regarding transcription factors that play an anti-inflammatory role in endothelial cells. Myocyte enhancer factor 2 C (MEF2C) is a transcription factor known to regulate angiogenesis in endothelial cells. Here, we report that MEF2C plays a critical function as an inhibitor of endothelial cell inflammation. Tumor necrosis factor (TNF)-α inhibited MEF2C expression in endothelial cells. Knockdown of MEF2C in endothelial cells resulted in the upregulation of pro-inflammatory molecules and stimulated leukocyte adhesion to endothelial cells. MEF2C knockdown also resulted in NF-κB activation in endothelial cells. Conversely, MEF2C overexpression by adenovirus significantly repressed TNF-α induction of pro-inflammatory molecules, activation of NF-κB, and leukocyte adhesion to endothelial cells. This inhibition of leukocyte adhesion by MEF2C was partially mediated by induction of KLF2. In mice, lipopolysaccharide (LPS)-induced leukocyte adhesion to the retinal vasculature was significantly increased by endothelial cell-specific ablation of MEF2C. Taken together, these results demonstrate that MEF2C is a novel negative regulator of inflammation in endothelial cells and may represent a therapeutic target for vascular inflammation. This article is protected by copyright. All rights reserved

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Changmeng Cai , Housheng Hansen He , Shuai Gao , Sen Chen , Ziyang Yu , Yanfei Gao , Shaoyong Chen , Mei Wei Chen , Jesse Zhang , Musaddeque Ahmed , Yang Wang , Eric Metzger , Roland Schüle , X. Shirley Liu , Myles Brown , Steven P. Balk Lysine-Specific Demethylase 1 (LSD1, KDM1A) functions as a transcriptional corepressor through demethylation of histone 3 lysine 4 (H3K4) but has a coactivator function on some genes through mechanisms that are unclear. We show that LSD1, interacting with CoREST, associates with and coactivates androgen receptor (AR) on a large fraction of androgen-stimulated genes. A subset of these AR/LSD1-associated enhancer sites have histone 3 threonine 6 phosphorylation (H3T6ph), and these sites are further enriched for androgen-stimulated genes. Significantly, despite its coactivator activity, LSD1 still mediates H3K4me2 demethylation at these androgen-stimulated enhancers. FOXA1 is also associated with LSD1 at AR-regulated enhancer sites, and a FOXA1 interaction with LSD1 enhances binding of both proteins at these sites. These findings show that LSD1 functions broadly as a regulator of AR function, that it maintains a transcriptional repression function at AR-regulated enhancers through H3K4 demethylation, and that it has a distinct AR-linked coactivator function mediated by demethylation of other substrates. Graphical abstract Teaser Cai et al. show that lysine-specific demethylase 1 (LSD1), although generally a transcriptional corepressor through H3K4 demethylation, functions broadly as a coactivator for androgen receptor and interacts with FOXA1 on androgen-stimulated genes. LSD1-mediated H3K4 demethylation persists at these sites, indicating a distinct coactivator function mediated by demethylation of other substrates.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Raajit Rampal , Altuna Alkalin , Jozef Madzo , Aparna Vasanthakumar , Elodie Pronier , Jay Patel , Yushan Li , Jihae Ahn , Omar Abdel-Wahab , Alan Shih , Chao Lu , Patrick S. Ward , Jennifer J. Tsai , Todd Hricik , Valeria Tosello , Jacob E. Tallman , Xinyang Zhao , Danette Daniels , Qing Dai , Luisa Ciminio , Iannis Aifantis , Chuan He , Francois Fuks , Martin S. Tallman , Adolfo Ferrando , Stephen Nimer , Elisabeth Paietta , Craig B. Thompson , Jonathan D. Licht , Christopher E. Mason , Lucy A. Godley , Ari Melnick , Maria E. Figueroa , Ross L. Levine Somatic mutations in IDH1/IDH2 and TET2 result in impaired TET2-mediated conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC). The observation that WT1 inactivating mutations anticorrelate with TET2/IDH1/IDH2 mutations in acute myeloid leukemia (AML) led us to hypothesize that WT1 mutations may impact TET2 function. WT1 mutant AML patients have reduced 5hmC levels similar to TET2/IDH1/IDH2 mutant AML. These mutations are characterized by convergent, site-specific alterations in DNA hydroxymethylation, which drive differential gene expression more than alterations in DNA promoter methylation. WT1 overexpression increases global levels of 5hmC, and WT1 silencing reduced 5hmC levels. WT1 physically interacts with TET2 and TET3, and WT1 loss of function results in a similar hematopoietic differentiation phenotype as observed with TET2 deficiency. These data provide a role for WT1 in regulating DNA hydroxymethylation and suggest that TET2 IDH1/IDH2 and WT1 mutations define an AML subtype defined by dysregulated DNA hydroxymethylation. Graphical abstract Teaser Mutational studies in patients with acute myeloid leukemia (AML) have identified recurrent mutations in TET2 and IDH1/IDH2 , and these mutations result in a reduction in 5-hydroxymethylcytosine (5hmC) levels. Rampal et al. demonstrate that WT1 mutations anticorrelate with TET2 and IDH1/IDH2 mutations, and WT1 mutant AMLs have decreased 5hmC levels, consistent with reduced TET2 function.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Shong Lau , Daniella Rylander Ottosson , Johan Jakobsson , Malin Parmar Recent findings show that human fibroblasts can be directly programmed into functional neurons without passing via a proliferative stem cell intermediate. These findings open up the possibility of generating subtype-specific neurons of human origin for therapeutic use from fetal cell, from patients themselves, or from matched donors. In this study, we present an improved system for direct neural conversion of human fibroblasts. The neural reprogramming genes are regulated by the neuron-specific microRNA, miR-124, such that each cell turns off expression of the reprogramming genes once the cell has reached a stable neuronal fate. The regulated system can be combined with integrase-deficient vectors, providing a nonintegrative and self-regulated conversion system that rids problems associated with the integration of viral transgenes into the host genome. These modifications make the system suitable for clinical use and therefore represent a major step forward in the development of induced neurons for cell therapy. Graphical abstract Teaser Lau et al. now use miRNA targeting to build a self-regulating neural conversion system. Combined with nonintegrating vectors, this system can efficiently drive conversion of human fibroblasts into functional induced neurons (iNs) suitable for clinical applications.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Lucas Leclère , Fabian Rentzsch Patterning of the metazoan dorsoventral axis is mediated by a complex interplay of BMP signaling regulators. Repulsive guidance molecule (RGM) is a conserved BMP coreceptor that has not been implicated in axis specification. We show that NvRGM is a key positive regulator of BMP signaling during secondary axis establishment in the cnidarian Nematostella vectensis . NvRGM regulates first the generation and later the shape of a BMP-dependent Smad1/5/8 gradient with peak activity on the side opposite the  NvBMP/NvRGM/NvChordin expression domain. Full knockdown of Smad1/5/8 signaling blocks the formation of endodermal structures, the mesenteries, and the establishment of bilateral symmetry, while altering the gradient through partial NvRGM or NvBMP knockdown shifts the boundaries of asymmetric gene expression and the positioning of the mesenteries along the secondary axis. These findings provide insight into the diversification of axis specification mechanisms and identify a previously unrecognized role for RGM in BMP-mediated axial patterning. Graphical abstract Teaser Leclère and Rentzsch identify repulsive guidance molecule (RGM) as an essential regulator of the BMP morphogen gradient that controls the formation and patterning of the secondary body axis in the sea anemone Nematostella . The evolutionary conservation of RGM-BMP interactions indicates that this function might also be important for bilaterian embryogenesis.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Grace E. Kim , Jack Kronengold , Giulia Barcia , Imran H. Quraishi , Hilary C. Martin , Edward Blair , Jenny C. Taylor , Olivier Dulac , Laurence Colleaux , Rima Nabbout , Leonard K. Kaczmarek Disease-causing mutations in ion channels generally alter intrinsic gating properties such as activation, inactivation, and voltage dependence. We examined nine different mutations of the KCNT1 (Slack) Na + -activated K + channel that give rise to three distinct forms of epilepsy. All produced many-fold increases in current amplitude compared to the wild-type channel. This could not be accounted for by increases in the intrinsic open probability of individual channels. Rather, greatly increased opening was a consequence of cooperative interactions between multiple channels in a patch. The degree of cooperative gating was much greater for all of the mutant channels than for the wild-type channel, and could explain increases in current even in a mutant with reduced unitary conductance. We also found that the same mutation gave rise to different forms of epilepsy in different individuals. Our findings indicate that a major consequence of these mutations is to alter channel-channel interactions. Graphical abstract Teaser Slack KCNT1 channels regulate how neurons respond to sustained stimulation. Kim et al. characterized nine KCNT1 mutations found in epilepsy patients with severe intellectual disabilities and showed that, in isolation, channel behavior is unaltered. However, in groups, mutant channels interact with each other abnormally, increasing current that flows through the channels.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Juanjuan Ou , Hongming Miao , Yinyan Ma , Feng Guo , Jia Deng , Xing Wei , Jie Zhou , Ganfeng Xie , Hang Shi , Bingzhong Xue , Houjie Liang , Liqing Yu How cancer cells shift metabolism to aerobic glycolysis is largely unknown. Here, we show that deficiency of α/β-hydrolase domain-containing 5 (Abhd5), an intracellular lipolytic activator that is also known as comparative gene identification 58 (CGI-58), promotes this metabolic shift and enhances malignancies of colorectal carcinomas (CRCs). Silencing of Abhd5 in normal fibroblasts induces malignant transformation. Intestine-specific knockout of Abhd5 in Apc Min/+ mice robustly increases tumorigenesis and malignant transformation of adenomatous polyps. In colon cancer cells, Abhd5 deficiency induces epithelial-mesenchymal transition by suppressing the AMPKα-p53 pathway, which is attributable to increased aerobic glycolysis. In human CRCs, Abhd5 expression falls substantially and correlates negatively with malignant features. Our findings link Abhd5 to CRC pathogenesis and suggest that cancer cells develop aerobic glycolysis by suppressing Abhd5-mediated intracellular lipolysis. Graphical abstract Teaser Cancer cells shift their metabolism to aerobic glycolysis (i.e., fermentation of glucose as energy in the presence of ample oxygen), but the underlying mechanisms remain elusive. Ou et al. identify Abhd5, an activator of intracellular fat breakdown, as a suppressor of this metabolic shift and associated malignancies in colon cancer.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Emmanuel Eggermann , Yves Kremer , Sylvain Crochet , Carl C.H. Petersen Internal brain states affect sensory perception, cognition, and learning. Many neocortical areas exhibit changes in the pattern and synchrony of neuronal activity during quiet versus active behaviors. Active behaviors are typically associated with desynchronized cortical dynamics. Increased thalamic firing contributes importantly to desynchronize mouse barrel cortex during active whisker sensing. However, a whisking-related cortical state change persists after thalamic inactivation, which is mediated at least in part by acetylcholine, as we show here by using whole-cell recordings, local pharmacology, axonal calcium imaging, and optogenetic stimulation. During whisking, we find prominent cholinergic signals in the barrel cortex, which suppress spontaneous cortical activity. The desynchronized state of barrel cortex during whisking is therefore driven by at least two distinct signals with opposing functions: increased thalamic activity driving glutamatergic excitation of the cortex and increased cholinergic input suppressing spontaneous cortical activity. Graphical abstract Teaser Eggermann et al. now find that the desynchronized state of the barrel cortex during active whisker sensing is accompanied by increased cholinergic input, which suppresses slow spontaneous cortical activity in excitatory layer 2/3 barrel cortex neurons.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Nancy A. Chow , Luke D. Jasenosky , Anne E. Goldfeld Interferon γ (IFN-γ) priming sensitizes monocytes and macrophages to lipopolysaccharide (LPS) stimulation, resulting in augmented expression of a set of genes including TNF . Here, we demonstrate that IFN-γ priming of LPS-stimulated TNF transcription requires a distal TNF/LT locus element 8 kb upstream of the TNF transcription start site (hHS-8). IFN-γ stimulation leads to increased DNase I accessibility of hHS-8 and its recruitment of interferon regulatory factor 1 (IRF1), and subsequent LPS stimulation enhances H3K27 acetylation and induces enhancer RNA synthesis at hHS-8. Ablation of IRF1 or targeting the hHS-8 IRF1 binding site in vivo with Cas9 linked to the KRAB repressive domain abolishes IFN-γ priming, but does not affect LPS induction of the gene. Thus, IFN-γ poises a distal enhancer in the TNF/LT locus by chromatin remodeling and IRF1 recruitment, which then drives enhanced TNF gene expression in response to a secondary toll-like receptor (TLR) stimulus. Graphical abstract Teaser Interferon γ (IFN-γ) priming is a critical immune event that enhances the monocyte and macrophage response, particularly expression of the TNF gene, to toll-like receptor (TLR) signaling. Chow et al. demonstrate that IFN-γ priming requires a distal enhancer element within the TNF/LT locus, thereby expanding the role of distal regulatory elements in the innate immune response.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Daniel W. Hagey , Jonas Muhr Organ formation and maintenance depends on slowly self-renewing stem cells that supply an intermediate population of rapidly dividing progenitors, but how this proliferative hierarchy is regulated is unknown. By performing genome-wide single-cell and functional analyses in the cortex, we demonstrate that reduced Sox2 expression is a key regulatory signature of the transition between stem cells and rapidly dividing progenitors. In stem cells, Sox2 is expressed at high levels, which enables its repression of proproliferative genes, of which Cyclin D1 is the most potent target. Sox2 confers this function through binding to low-affinity motifs, which facilitate the recruitment of Gro/Tle corepressors in synergy with Tcf/Lef proteins. Upon differentiation, proneural factors reduce Sox2 expression, which derepresses Cyclin D1 and promotes proliferation. Our results show how concentration-dependent Sox2 occupancy of DNA motifs of varying affinities translates into recruitment of repressive complexes, which regulate the proliferative dynamics of neural stem and progenitor cells. Graphical abstract Teaser Hagey and Muhr show that high levels of Sox2 maintain stem cells of the developing cortex in a slowly self-renewing state by directly repressing cell-cycle genes. They further demonstrate that proneural protein-induced commitment to differentiation induces a rapidly dividing state via the reduction of Sox2 expression levels.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Jakub O. Westholm , Pedro Miura , Sara Olson , Sol Shenker , Brian Joseph , Piero Sanfilippo , Susan E. Celniker , Brenton R. Graveley , Eric C. Lai Circularization was recently recognized to broadly expand transcriptome complexity. Here, we exploit massive Drosophila total RNA-sequencing data, >5 billion paired-end reads from >100 libraries covering diverse developmental stages, tissues, and cultured cells, to rigorously annotate >2,500 fruit fly circular RNAs. These mostly derive from back-splicing of protein-coding genes and lack poly(A) tails, and the circularization of hundreds of genes is conserved across multiple Drosophila species. We elucidate structural and sequence properties of Drosophila circular RNAs, which exhibit commonalities and distinctions from mammalian circles. Notably, Drosophila circular RNAs harbor >1,000 well-conserved canonical miRNA seed matches, especially within coding regions, and coding conserved miRNA sites reside preferentially within circularized exons. Finally, we analyze the developmental and tissue specificity of circular RNAs and note their preferred derivation from neural genes and enhanced accumulation in neural tissues. Interestingly, circular isoforms increase substantially relative to linear isoforms during CNS aging and constitute an aging biomarker. Graphical abstract Teaser Westholm et al. annotate Drosophila circular RNAs from a massive collection of total RNA-seq data, providing insights into their biogenesis and function. In particular, circularizing exons are predominantly associated with long flanking introns, are preferred locations of conserved coding miRNA sites, and accumulate to highest levels in the aging CNS.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Bo-Kuan Wu , Charles Brenner Hypermethylation-mediated tumor suppressor gene (TSG) silencing is a central epigenetic alteration in RAS-dependent tumorigenesis. Ten-eleven translocation (TET) enzymes can depress DNA methylation by hydroxylation of 5-methylcytosine (5mC) bases to 5-hydroxymethylcytosine (5hmC). Here, we report that suppression of TET1 is required for KRAS-induced DNA hypermethylation and cellular transformation. In distinct nonmalignant cell lines, oncogenic KRAS promotes transformation by inhibiting TET1 expression via the ERK-signaling pathway. This reduces chromatin occupancy of TET1 at TSG promoters, lowers levels of 5hmC, and increases levels of 5mC and 5mC-dependent transcriptional silencing. Restoration of TET1 expression by ERK pathway inhibition or ectopic TET1 reintroduction in KRAS-transformed cells reactivates TSGs and inhibits colony formation. KRAS knockdown increases TET1 expression and diminishes colony-forming ability, whereas KRAS/TET1 double knockdown bypasses the KRAS dependence of KRAS-addicted cancer cells. Thus, suppression of TET1-dependent DNA demethylation is critical for KRAS-mediated transformation. Graphical abstract Teaser Wu and Brenner now show that KRAS drives ERK-dependent mRNA suppression of TET1 and that this is necessary to achieve tumor suppressor gene hypermethylation and malignant transformation.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Zekiye Buket Yılmaz , Bente Kofahl , Patrick Beaudette , Katharina Baum , Inbal Ipenberg , Falk Weih , Jana Wolf , Gunnar Dittmar , Claus Scheidereit The mechanisms that govern proteolytic maturation or complete destruction of the precursor proteins p100 and p105 are fundamental to homeostasis and activation of NF-κB; however, they remain poorly understood. Using mass-spectrometry-based quantitative analysis of noncanonical LTβR-induced signaling, we demonstrate that stimulation induces simultaneous processing of both p100 and p105. The precursors not only form hetero-oligomers but also interact with the ATPase VCP/p97, and their induced proteolysis strictly depends on the signal response domain (SRD) of p100, suggesting that the SRD-targeting proteolytic machinery acts in cis and in trans . Separation of cellular pools by isotope labeling revealed synchronous dynamics of p105 and p100 proteolysis. The generation of p50 and p52 from their precursors depends on functional VCP/p97. We have developed quantitative mathematical models that describe the dynamics of the system and predict that p100-p105 complexes are signal responsive. Graphical abstract Teaser Proteolytic precursor processing is a hallmark of the NF-κB system. Yilmaz et al. show that in lymphotoxin-stimulated cells p100 acts upstream of p105, resulting in concurrent production of p52 and p50. Both precursors form complexes and bind to segregase (p97/VCP), which promotes proteasomal processing. The findings are supported by mass spectrometry and incorporated in quantitative mathematical models.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Lisa L. Korn , Harper G. Hubbeling , Paige M. Porrett , Qi Yang , Lisa G. Barnett , Terri M. Laufer Regulatory T cells (Tregs) are CD4 + T cells that maintain immune homeostasis and prevent autoimmunity. Like all CD4 + T cells, Tregs require antigen-specific signals via T cell receptor-major histocompatibility complex class II (TCR-MHCII) interactions for their development. However, the requirement for MHCII in Treg homeostasis in tissues such as intestinal lamina propria (LP) is unknown. We examined LP Treg homeostasis in a transgenic mouse model that lacks peripheral TCR-MHCII interactions and generation of extrathymic Tregs (iTregs). Thymically generated Tregs entered the LP of weanlings and proliferated independently of MHCII to fill the compartment. The adult LP was a closed niche; new thymic Tregs were excluded, and Tregs in parabiotic pairs were LP resident. The isolated LP niche was interleukin-2 (IL-2) independent but dependent on commensal bacteria. Thus, an LP Treg niche can be filled, isolated, and maintained independently of antigen signals and iTregs. This niche may represent a tissue-specific mechanism for maintaining immune tolerance. Graphical abstract Teaser Regulatory T cells (Tregs) maintain immune homeostasis and prevent autoimmunity. Korn et al. describe a unique Treg niche in the intestinal lamina propria that does not require T cell receptor signals for development or maintenance and is physiologically isolated from the Tregs that circulate through lymphoid organs. Maintenance of this niche is dependent upon local commensal bacteria. The authors propose that this isolated niche may represent a tissue-specific mechanism for maintaining immune tolerance.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Romain Christiano , Nagarjuna Nagaraj , Florian Fröhlich , Tobias C. Walther How cells maintain specific levels of each protein and whether that control is evolutionarily conserved are key questions. Here, we report proteome-wide steady-state protein turnover rate measurements for the evolutionarily distant but ecologically similar yeasts, Saccharomyces cerevisiae and Schizosaccharomyces pombe . We find that the half-life of most proteins is much longer than currently thought and determined to a large degree by protein synthesis and dilution due to cell division. However, we detect a significant subset of proteins (∼15%) in both yeasts that are turned over rapidly. In addition, the relative abundances of orthologous proteins between the two yeasts are highly conserved across the 400 million years of evolution. In contrast, their respective turnover rates differ considerably. Our data provide a high-confidence resource for studying protein degradation in common yeast model systems. Graphical abstract Teaser Christiano et al. report protein turnover rates for S. cerevisiae and S. pombe . Overall, protein half-life is far less conserved than protein abundance. Protein turnover profiling of hrd1 Δ, an endoplasmic-reticulum-associated degradation (ERAD)-defective mutant, identifies candidate substrates for this pathway in budding yeast.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Brian C. Shonesy , Rebecca J. Bluett , Teniel S. Ramikie , Rita Báldi , Daniel J. Hermanson , Philip J. Kingsley , Lawrence J. Marnett , Danny G. Winder , Roger J. Colbran , Sachin Patel Endocannabinoid (eCB) signaling has been heavily implicated in the modulation of anxiety and depressive behaviors and emotional learning. However, the role of the most-abundant endocannabinoid 2-arachidonoylglycerol (2-AG) in the physiological regulation of affective behaviors is not well understood. Here, we show that genetic deletion of the 2-AG synthetic enzyme diacylglycerol lipase α (DAGLα) in mice reduces brain, but not circulating, 2-AG levels. DAGLα deletion also results in anxiety-like and sex-specific anhedonic phenotypes associated with impaired activity-dependent eCB retrograde signaling at amygdala glutamatergic synapses. Importantly, acute pharmacological normalization of 2-AG levels reverses both phenotypes of DAGLα-deficient mice. These data suggest 2-AG deficiency could contribute to the pathogenesis of affective disorders and that pharmacological normalization of 2-AG signaling could represent an approach for the treatment of mood and anxiety disorders. Graphical abstract Teaser The role of the primary endogenous cannabinoid 2-AG in mood and anxiety regulation is not well understood. Shonesy et al. show that deletion of a primary 2-AG synthetic enzyme, DAGLα, results in anxiety and sex-specific depressive phenotypes, which can be rapidly reversed by pharmacological normalization of endocannabinoid levels.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Kiran Batta , Magdalena Florkowska , Valerie Kouskoff , Georges Lacaud Recent reports have shown that somatic cells, under appropriate culture conditions, could be directly reprogrammed to cardiac, hepatic, or neuronal phenotype by lineage-specific transcription factors. In this study, we demonstrate that both embryonic and adult somatic fibroblasts can be efficiently reprogrammed to clonal multilineage hematopoietic progenitors by the ectopic expression of the transcription factors ERG, GATA2, LMO2, RUNX1c, and SCL. These reprogrammed cells were stably expanded on stromal cells and possessed short-term reconstitution ability in vivo. Loss of p53 function facilitated reprogramming to blood, and p53 −/− reprogrammed cells efficiently generated erythroid, megakaryocytic, myeloid, and lymphoid lineages. Genome-wide analyses revealed that generation of hematopoietic progenitors was preceded by the appearance of hemogenic endothelial cells expressing endothelial and hematopoietic genes. Altogether, our findings suggest that direct reprogramming could represent a valid alternative approach to the differentiation of embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) for disease modeling and autologous blood cell therapies. Graphical abstract Teaser Batta et al. demonstrate that murine fibroblasts are reprogrammed to hematopoietic progenitors, with erythroid, megakaryocyte, and myeloid potential, by ectopic expression of hematopoietic transcription factors. Reprogramming efficiency is increased by loss of p53 function, and generation of blood cells is preceded by the appearance of hemogenic endothelium.

Description: Publication date: Available online 4 December 2014 Source: Cell Reports Author(s): Nancy F. Ramia , Michael Spilman , Li Tang , Yaming Shao , Joshua Elmore , Caryn Hale , Alexis Cocozaki , Nilakshee Bhattacharya , Rebecca M. Terns , Michael P. Terns , Hong Li , Scott M. Stagg The Cmr complex is the multisubunit effector complex of the type III-B clustered regularly interspaced short palindromic repeats (CRISPR)-Cas immune system. The Cmr complex recognizes a target RNA through base pairing with the integral CRISPR RNA (crRNA) and cleaves the target at multiple regularly spaced locations within the complementary region. To understand the molecular basis of the function of this complex, we have assembled information from electron microscopic and X-ray crystallographic structural studies and mutagenesis of a complete Pyrococcus furiosus Cmr complex. Our findings reveal that four helically packed Cmr4 subunits, which make up the backbone of the Cmr complex, act as a platform to support crRNA binding and target RNA cleavage. Interestingly, we found a hook-like structural feature associated with Cmr4 that is likely the site of target RNA binding and cleavage. Our results also elucidate analogies in the mechanisms of crRNA and target molecule binding by the distinct Cmr type III-A and Cascade type I-E complexes. Graphical abstract Teaser Ramia et al. show that the helical core of the type III-B Cmr CRISPR-Cas effector complex, made up of multiple Cmr4 subunits, forms the platform for a corresponding number of cleavages of the target RNA. Comparison with the type I-E Cascade structure reveals strikingly similar mechanisms of crRNA and target binding.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Christos G. Gkogkas , Arkady Khoutorsky , Ruifeng Cao , Seyed Mehdi Jafarnejad , Masha Prager-Khoutorsky , Nikolaos Giannakas , Archontia Kaminari , Apostolia Fragkouli , Karim Nader , Theodore J. Price , Bruce W. Konicek , Jeremy R. Graff , Athina K. Tzinia , Jean-Claude Lacaille , Nahum Sonenberg Fragile X syndrome (FXS) is the leading genetic cause of autism. Mutations in Fmr1 (fragile X mental retardation 1 gene) engender exaggerated translation resulting in dendritic spine dysmorphogenesis, synaptic plasticity alterations, and behavioral deficits in mice, which are reminiscent of FXS phenotypes. Using postmortem brains from FXS patients and Fmr1 knockout mice (Fmr1 −/y ), we show that phosphorylation of the mRNA 5′ cap binding protein, eukaryotic initiation factor 4E (eIF4E), is elevated concomitant with increased expression of matrix metalloproteinase 9 (MMP-9) protein. Genetic or pharmacological reduction of eIF4E phosphorylation rescued core behavioral deficits, synaptic plasticity alterations, and dendritic spine morphology defects via reducing exaggerated translation of Mmp9 mRNA in Fmr1 −/y mice, whereas MMP-9 overexpression produced several FXS-like phenotypes. These results uncover a mechanism of regulation of synaptic function by translational control of Mmp-9 in FXS, which opens the possibility of new treatment avenues for the diverse neurological and psychiatric aspects of FXS. Graphical abstract Teaser Fragile X syndrome (FXS) is caused by dysregulation of translation in the brain. Gkogkas et al. show that phosphorylation of eukaryotic translation initiation factor 4E (eIF4E) is increased in FXS postmortem brains and Fmr1 −/y mice. Downregulation of eIF4E phosphorylation in Fmr1 −/y mice rescues defects in dendritic spine morphology, synaptic plasticity, and social interaction via normalization of MMP-9 expression.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Wei-Qun Fang , Wei-Wei Chen , Liwen Jiang , Kai Liu , Wing-Ho Yung , Amy K.Y. Fu , Nancy Y. Ip The functional integrity of the neocortex depends upon proper numbers of excitatory and inhibitory neurons; however, the consequences of dysregulated neuronal production during the development of the neocortex are unclear. As excess cortical neurons are linked to the neurodevelopmental disorder autism, we investigated whether the overproduction of neurons leads to neocortical malformation and malfunction in mice. We experimentally increased the number of pyramidal neurons in the upper neocortical layers by using the small molecule XAV939 to expand the intermediate progenitor population. The resultant overpopulation of neurons perturbs development of dendrites and spines of excitatory neurons and alters the laminar distribution of interneurons. Furthermore, these phenotypic changes are accompanied by dysregulated excitatory and inhibitory synaptic connection and balance. Importantly, these mice exhibit behavioral abnormalities resembling those of human autism. Thus, our findings collectively suggest a causal relationship between neuronal overproduction and autism-like features, providing developmental insights into the etiology of autism. Graphical abstract Teaser Fang et al. generated a mouse model with excessive excitatory neurons in the neocortex by manipulating embryonic neurogenesis. Overproduction of neurons results in autism-like anatomical and behavioral features. These findings suggest a causal relationship between overproduction of neurons and cortical malfunction and provide developmental insights into the etiology of autism.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Lauren J. Manderfield , Kurt A. Engleka , Haig Aghajanian , Mudit Gupta , Steven Yang , Li Li , Julie E. Baggs , John B. Hogenesch , Eric N. Olson , Jonathan A. Epstein Loss of Pax3, a developmentally regulated transcription factor expressed in premigratory neural crest, results in severe developmental defects and embryonic lethality. Although Pax3 mutations produce profound phenotypes, the intrinsic transcriptional activation exhibited by Pax3 is surprisingly modest. We postulated the existence of transcriptional coactivators that function with Pax3 to mediate developmental functions. A high-throughput screen identified the Hippo effector proteins Taz and Yap65 as Pax3 coactivators. Synergistic coactivation of target genes by Pax3-Taz/Yap65 requires DNA binding by Pax3, is Tead independent, and is regulated by Hippo kinases Mst1 and Lats2. In vivo, Pax3 and Yap65 colocalize in the nucleus of neural crest progenitors in the dorsal neural tube. Neural crest deletion of Taz and Yap65 results in embryo-lethal neural crest defects and decreased expression of the Pax3 target gene, Mitf . These results suggest that Pax3 activity is regulated by the Hippo pathway and that Pax factors are Hippo effectors. Graphical abstract Teaser Hippo signaling is a conserved kinase cascade that mediates transcription through a Yap65/Taz-Tead complex and governs organ size. Manderfield et al. have identified Yap65 and Taz as coactivators of Pax factors and established neural crest as a novel site of Hippo signaling. The Pax3-Yap65/Taz complex is regulated by upstream Hippo kinases and is Tead independent. Deletion of Yap65 and Taz in neural crest generated differentiation defects in specific neural crest derivatives.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Chris M. Woodard , Brian A. Campos , Sheng-Han Kuo , Melissa J. Nirenberg , Michael W. Nestor , Matthew Zimmer , Eugene V. Mosharov , David Sulzer , Hongyan Zhou , Daniel Paull , Lorraine Clark , Eric E. Schadt , Sergio Pablo Sardi , Lee Rubin , Kevin Eggan , Mathew Brock , Scott Lipnick , Mahendra Rao , Stephen Chang , Aiqun Li , Scott A. Noggle Parkinson’s disease (PD) has been attributed to a combination of genetic and nongenetic factors. We studied a set of monozygotic twins harboring the heterozygous glucocerebrosidase mutation ( GBA N370S) but clinically discordant for PD. We applied induced pluripotent stem cell (iPSC) technology for PD disease modeling using the twins’ fibroblasts to evaluate and dissect the genetic and nongenetic contributions. Utilizing fluorescence-activated cell sorting, we obtained a homogenous population of “footprint-free” iPSC-derived midbrain dopaminergic (mDA) neurons. The mDA neurons from both twins had ∼50% GBA enzymatic activity, ∼3-fold elevated α-synuclein protein levels, and a reduced capacity to synthesize and release dopamine. Interestingly, the affected twin’s neurons showed an even lower dopamine level, increased monoamine oxidase B (MAO-B) expression, and impaired intrinsic network activity. Overexpression of wild-type GBA and treatment with MAO-B inhibitors normalized α-synuclein and dopamine levels, suggesting a combination therapy for the affected twin. Graphical abstract Teaser Woodard et al. studied a set of identical twins harboring GBA N370S mutation but clinically discordant for Parkinson's disease (PD). Using iPSC technology, they found that enzymes GBA and MAO-B could be therapeutic targets in PD. These findings shed light into future studies using iPSCs to model idiopathic PD cases.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Hyokjoon Kwon , Sarnia Laurent , Yan Tang , Haihong Zong , Pratibha Vemulapalli , Jeffrey E. Pessin Adipose tissue inflammation is one pathway shown to mediate insulin resistance in obese humans and rodents. Obesity induces dynamic cellular changes in adipose tissue to increase proinflammatory cytokines and diminish anti-inflammatory cytokines. However, we have found that anti-inflammatory interleukin-13 (IL-13) is unexpectedly induced in adipose tissue of obese humans and high-fat diet (HFD)-fed mice, and the source of IL-13 is primarily the adipocyte. Moreover, HFD-induced proinflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and IL-1β mediate IL-13 production in adipocytes in an IKKβ-dependent manner. In contrast, adipocyte-specific IKKβ-deficient mice show diminished IL-13 expression and enhanced inflammation after HFD feeding, resulting in a worsening of the insulin-resistant state. Together these data demonstrate that although IKKβ activates the expression of proinflammatory mediators, in adipocytes, IKKβ signaling also induces the expression of the anti-inflammatory cytokine IL-13, which plays a unique protective role by limiting adipose tissue inflammation and insulin resistance. Graphical abstract Teaser IKKβ is known to be a proinflammatory mediator. However, IKKβ in adipocytes also mediates IL-13 expression to suppress high-fat-diet-induced inflammation in adipose tissue. This feedback mechanism may be the molecular basis of diet-induced chronic low-grade inflammation, resulting in systemic insulin resistance.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Morgane Belle , David Godefroy , Chloé Dominici , Céline Heitz-Marchaland , Pavol Zelina , Farida Hellal , Frank Bradke , Alain Chédotal Clearing techniques have been developed to transparentize mouse brains, thereby preserving 3D structure, but their complexity has limited their use. Here, we show that immunolabeling of axonal tracts followed by optical clearing with solvents (3DISCO) and light-sheet microscopy reveals brain connectivity in mouse embryos and postnatal brains. We show that the Robo3 receptor is selectively expressed by medial habenula axons forming the fasciculus retroflexus (FR) and analyzed the development of this commissural tract in mutants of the Slit/Robo and DCC/Netrin pathways. Netrin-1 and DCC are required to attract FR axons to the midline, but the two mutants exhibit specific and heterogeneous axon guidance defects. Moreover, floor-plate-specific deletion of Slit ligands with a conditional Slit2 allele perturbs not only midline crossing by FR axons but also their anteroposterior distribution. In conclusion, this method represents a unique and powerful imaging tool to study axonal connectivity in mutant mice. Graphical abstract Teaser Clearing techniques have recently been developed to look at mouse brains, but they are complex and expensive. Belle et al. now describe a simple procedure that combines immunolabeling, solvent-based clearing, and light-sheet fluorescence microscopy. This technique allows large-scale screening of axon guidance defects and other developmental disorders in mutant mice.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Emmanuelle Godefroy , Anne Gallois , Juliana Idoyaga , Miriam Merad , Navpreet Tung , Ngozi Monu , Yvonne Saenger , Yichun Fu , Rajesh Ravindran , Bali Pulendran , Francine Jotereau , Sergio Trombetta , Nina Bhardwaj Matrix metalloproteinase-2 (MMP-2) is involved in several physiological mechanisms, including wound healing and tumor progression. We show that MMP-2 directly stimulates dendritic cells (DCs) to both upregulate OX40L on the cell surface and secrete inflammatory cytokines. The mechanism underlying DC activation includes physical association with Toll-like receptor-2 (TLR2), leading to NF-κB activation, OX40L upregulation on DCs, and ensuing T H 2 differentiation. Significantly, MMP-2 polarizes T cells toward type 2 responses in vivo, in a TLR2-dependent manner. MMP-2-dependent type 2 polarization may represent a key immune regulatory mechanism for protection against a broad array of disorders, such as inflammatory, infectious, and autoimmune diseases, which can be hijacked by tumors to evade immunity. Graphical abstract Teaser Godefroy et al. now demonstrate that matrix metalloproteinase-2 (MMP-2) directly interacts with and activates dendritic cells (DCs) via Toll-like receptor-2. MMP-2-exposed DCs upregulate OX40L, promoting type 2 polarization both in vitro and in vivo. This may represent a key immune regulatory mechanism involved in a variety of inflammatory disorders.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Catalin Chimerel , Edward Emery , David K. Summers , Ulrich Keyser , Fiona M. Gribble , Frank Reimann It has long been speculated that metabolites, produced by gut microbiota, influence host metabolism in health and diseases. Here, we reveal that indole, a metabolite produced from the dissimilation of tryptophan, is able to modulate the secretion of glucagon-like peptide-1 (GLP-1) from immortalized and primary mouse colonic L cells. Indole increased GLP-1 release during short exposures, but it reduced secretion over longer periods. These effects were attributed to the ability of indole to affect two key molecular mechanisms in L cells. On the one hand, indole inhibited voltage-gated K + channels, increased the temporal width of action potentials fired by L cells, and led to enhanced Ca 2+ entry, thereby acutely stimulating GLP-1 secretion. On the other hand, indole slowed ATP production by blocking NADH dehydrogenase, thus leading to a prolonged reduction of GLP-1 secretion. Our results identify indole as a signaling molecule by which gut microbiota communicate with L cells and influence host metabolism. Graphical abstract Teaser Indole is the main metabolite produced by gut bacteria from tryptophan. Chimerel et al. demonstrate that indole modulates the hormone secretion of enteroendocrine L cells and reveal the molecular mechanism behind this modulation. These findings suggest that the production of indole by bacteria could have a major impact on host metabolism.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Qing-ming Gao , Keshun Yu , Ye Xia , M.B. Shine , Caixia Wang , DuRoy Navarre , Aardra Kachroo , Pradeep Kachroo The plant galactolipids monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) have been linked to the anti-inflammatory and cancer benefits of a green leafy vegetable diet in humans due to their ability to regulate the levels of free radicals like nitric oxide (NO). Here, we show that DGDG contributes to plant NO as well as salicylic acid biosynthesis and is required for the induction of systemic acquired resistance (SAR). In contrast, MGDG regulates the biosynthesis of the SAR signals azelaic acid (AzA) and glycerol-3-phosphate (G3P) that function downstream of NO. Interestingly, DGDG is also required for AzA-induced SAR, but MGDG is not. Notably, transgenic expression of a bacterial glucosyltransferase is unable to restore SAR in dgd1 plants even though it does rescue their morphological and fatty acid phenotypes. These results suggest that MGDG and DGDG are required at distinct steps and function exclusively in their individual roles during the induction of SAR. Graphical abstract Teaser The galactolipids monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) constitute ∼80% of total membrane lipids in plants. Gao et al. now show that these galactolipids function nonredundantly to regulate systemic acquired resistance (SAR). Furthermore, they show that the terminal galactose on the α-galactose-β-galactose head group of DGDG is critical for SAR.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Rafael B. Blasco , Elif Karaca , Chiara Ambrogio , Taek-Chin Cheong , Emre Karayol , Valerio G. Minero , Claudia Voena , Roberto Chiarle Generation of genetically engineered mouse models (GEMMs) for chromosomal translocations in the endogenous loci by a knockin strategy is lengthy and costly. The CRISPR/Cas9 system provides an innovative and flexible approach for genome engineering of genomic loci in vitro and in vivo. Here, we report the use of the CRISPR/Cas9 system for engineering a specific chromosomal translocation in adult mice in vivo. We designed CRISPR/Cas9 lentiviral vectors to induce cleavage of the murine endogenous Eml4 and Alk loci in order to generate the Eml4-Alk gene rearrangement recurrently found in non-small-cell lung cancers (NSCLCs). Intratracheal or intrapulmonary inoculation of lentiviruses induced Eml4-Alk gene rearrangement in lung cells in vivo. Genomic and mRNA sequencing confirmed the genome editing and the production of the Eml4-Alk fusion transcript. All mice developed Eml4-Alk -rearranged lung tumors 2 months after the inoculation, demonstrating that the CRISPR/Cas9 system is a feasible and simple method for the generation of chromosomal rearrangements in vivo. Graphical abstract Teaser Blasco et al. demonstrate that CRISPR/Cas9 technology can be exploited to generate targeted chromosomal rearrangements in vivo in mice in a time- and cost-effective approach.

Description: Publication date: Available online 26 November 2014 Source: Cell Reports Author(s): Andrea Galmozzi , Si B. Sonne , Svetlana Altshuler-Keylin , Yutaka Hasegawa , Kosaku Shinoda , Ineke H.N. Luijten , Jae Won Chang , Louis Z. Sharp , Benjamin F. Cravatt , Enrique Saez , Shingo Kajimura Obesity develops when energy intake chronically exceeds energy expenditure. Because brown adipose tissue (BAT) dissipates energy in the form of heat, increasing energy expenditure by augmenting BAT-mediated thermogenesis may represent an approach to counter obesity and its complications. The ability of BAT to dissipate energy is dependent on expression of mitochondrial uncoupling protein 1 (UCP1). To facilitate the identification of pharmacological modulators of BAT UCP1 levels, which may have potential as antiobesity medications, we developed a transgenic model in which luciferase activity faithfully mimics endogenous UCP1 expression and its response to physiologic stimuli. Phenotypic screening of a library using cells derived from this model yielded a small molecule that increases UCP1 expression in brown fat cells and mice. Upon adrenergic stimulation, compound-treated mice showed increased energy expenditure. These tools offer an opportunity to identify pharmacologic modulators of UCP1 expression and uncover regulatory pathways that impact BAT-mediated thermogenesis. Graphical abstract Teaser Pharmacological activation of brown adipose tissue (BAT) thermogenesis and energy dissipation, a process mediated by UCP1, may be useful to counter the energy imbalance that engenders obesity. Galmozzi et al. have developed an in vivo model to monitor UCP1 expression in real time and identified a small molecule that increases UCP1 levels. Mice treated with this molecule show greater energy expenditure upon adrenergic stimulation. Discovery of compounds with this ability is an important stride toward enhancing BAT function in obese individuals.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Yanmeng Guo , Yuping Wang , Qingxiu Wang , Zuoren Wang In Drosophila larvae, the class IV dendritic arborization (da) neurons are polymodal nociceptors. Here, we show that ppk26 (CG8546) plays an important role in mechanical nociception in class IV da neurons. Our immunohistochemical and functional results demonstrate that ppk26 is specifically expressed in class IV da neurons. Larvae with mutant ppk26 showed severe behavioral defects in a mechanical nociception behavioral test but responded to noxious heat stimuli comparably to wild-type larvae. In addition, functional studies suggest that ppk26 and ppk (also called ppk1 ) function in the same pathway, whereas piezo functions in a parallel pathway. Consistent with these functional results, we found that PPK and PPK26 are interdependent on each other for their cell surface localization. Our work indicates that PPK26 and PPK might form heteromeric DEG/ENaC channels that are essential for mechanotransduction in class IV da neurons. Graphical abstract Teaser Sensing painful stimuli is of vital importance for animal survival. Guo et al. now find that PPK26 is selectively expressed in class IV dendritic arborization neurons and contributes to mechanical nociception but not thermal nociception in Drosophila larvae.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Amol Ugale , Gudmundur L. Norddahl , Martin Wahlestedt , Petter Säwén , Pekka Jaako , Cornelis Jan Pronk , Shamit Soneji , Jörg Cammenga , David Bryder Studies of developmental pathways of hematopoietic stem cells (HSCs) have defined lineage relationships throughout the blood system. This is relevant to acute myeloid leukemia (AML), where aggressiveness and therapeutic responsiveness can be influenced by the initial stage of transformation. To address this, we generated a mouse model in which the mixed-lineage leukemia/eleven-nineteen-leukemia (MLL-ENL) transcription factor can be conditionally activated in any cell type. We show that AML can originate from multiple hematopoietic progenitor subsets with granulocytic and monocytic potential, and that the normal developmental position of leukemia-initiating cells influences leukemic development. However, disease failed to arise from HSCs. Although it maintained or upregulated the expression of target genes associated with leukemic development, MLL-ENL dysregulated the proliferative and repopulating capacity of HSCs. Therefore, the permissiveness for development of AML may be associated with a narrower window of differentiation than was previously appreciated, and hijacking the self-renewal capacity of HSCs by a potent oncogene is insufficient for leukemic development. Graphical abstract Teaser The cellular origin of leukemia driven by MLL fusions has been suggested to underlie heterogeneity in aggressiveness and prognosis. Ugale et al. now use an inducible MLL-ENL mouse model to study leukemia initiation and competence throughout the hematopoietic hierarchy. Although AML development could originate from multiple progenitor subsets, the most primitive stem cells were unexpectedly unable to initiate disease.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Nila Roy Choudhury , Jakub S. Nowak , Juan Zuo , Juri Rappsilber , Steven H. Spoel , Gracjan Michlewski RNA binding proteins have thousands of cellular RNA targets and often exhibit opposite or passive molecular functions. Lin28a is a conserved RNA binding protein involved in pluripotency and tumorigenesis that was previously shown to trigger TuT4-mediated pre-let-7 uridylation, inhibiting its processing and targeting it for degradation. Surprisingly, despite binding to other pre-microRNAs (pre-miRNAs), only pre-let-7 is efficiently uridylated by TuT4. Thus, we hypothesized the existence of substrate-specific cofactors that stimulate Lin28a-mediated pre-let-7 uridylation or restrict its functionality on non-let-7 pre-miRNAs. Through RNA pull-downs coupled with quantitative mass spectrometry, we identified the E3 ligase Trim25 as an RNA-specific cofactor for Lin28a/TuT4-mediated uridylation. We show that Trim25 binds to the conserved terminal loop (CTL) of pre-let-7 and activates TuT4, allowing for more efficient Lin28a-mediated uridylation. These findings reveal that protein-modifying enzymes, only recently shown to bind RNA, can guide the function of canonical ribonucleoprotein (RNP) complexes in cis , thereby providing an additional level of specificity. Graphical abstract Teaser Lin28a triggers TuT4-mediated pre-let-7 uridylation. Despite binding to other pre-microRNAs, only pre-let-7 is efficiently uridylated by TuT4. Choudhury et al. show that Trim25 is an RNA-specific cofactor for Lin28a/TuT4-mediated uridylation. These findings reveal that Trim25 can guide the function of canonical RNP complexes in cis , thereby providing an additional level of specificity.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Anita Parmigiani , Aida Nourbakhsh , Boxiao Ding , Wei Wang , Young Chul Kim , Konstantin Akopiants , Kun-Liang Guan , Michael Karin , Andrei V. Budanov The mechanistic target of rapamycin complex 1 (mTORC1) kinase is a sensor of different environmental conditions and regulator of cell growth, metabolism, and autophagy. mTORC1 is activated by Rag GTPases, working as RagA:RagB and RagC:RagD heterodimers. Rags control mTORC1 activity by tethering mTORC1 to the lysosomes where it is activated by Rheb GTPase. RagA:RagB, active in its GTP-bound form, is inhibited by GATOR1 complex, a GTPase-activating protein, and GATOR1 is in turn negatively regulated by GATOR2 complex. Sestrins are stress-responsive proteins that inhibit mTORC1 via activation of AMP-activated protein kinase (AMPK) and tuberous sclerosis complex. Here we report an AMPK-independent mechanism of mTORC1 inhibition by Sestrins mediated by their interaction with GATOR2. As a result of this interaction, the Sestrins suppress mTOR lysosomal localization in a Rag-dependent manner. This mechanism is potentially involved in mTORC1 regulation by amino acids, rotenone, and tunicamycin, connecting stress response with mTORC1 inhibition. Graphical abstract Teaser The mTORC1 kinase integrates various environmental signals to regulate cell growth and metabolism. Parmigiani et al. identified a mechanism of mTORC1 regulation by Sestrins via interaction with GATOR2 and suppression of mTOR lysosomal localization.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Nicholas J. Bessman , Atrish Bagchi , Kathryn M. Ferguson , Mark A. Lemmon The epidermal growth factor receptor (EGFR) plays pivotal roles in development and is mutated or overexpressed in several cancers. Despite recent advances, the complex allosteric regulation of EGFR remains incompletely understood. Through efforts to understand why the negative cooperativity observed for intact EGFR is lost in studies of its isolated extracellular region (ECR), we uncovered unexpected relationships between ligand binding and receptor dimerization. The two processes appear to compete. Surprisingly, dimerization does not enhance ligand binding (although ligand binding promotes dimerization). We further show that simply forcing EGFR ECRs into preformed dimers without ligand yields ill-defined, heterogeneous structures. Finally, we demonstrate that extracellular EGFR-activating mutations in glioblastoma enhance ligand-binding affinity without directly promoting EGFR dimerization, suggesting that these oncogenic mutations alter the allosteric linkage between dimerization and ligand binding. Our findings have important implications for understanding how EGFR and its relatives are activated by specific ligands and pathological mutations. Graphical abstract Teaser Although the epidermal growth factor receptor (EGFR) was the first growth factor receptor for which ligand-induced dimerization was established, the relationship between growth factor binding and EGFR dimerization remains unclear. Bessman et al. use a range of biophysical methods to study growth factor binding to EGFR variants, including those found in glioblastoma, to shed light on ligand-specific allosteric control of EGFR.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Kristen L. Karlin , Gourish Mondal , Jessica K. Hartman , Siddhartha Tyagi , Sarah J. Kurley , Chris S. Bland , Tiffany Y.T. Hsu , Alexander Renwick , Justin E. Fang , Ilenia Migliaccio , Celetta Callaway , Amritha Nair , Rocio Dominguez-Vidana , Don X. Nguyen , C. Kent Osborne , Rachel Schiff , Li-Yuan Yu-Lee , Sung Y. Jung , Dean P. Edwards , Susan G. Hilsenbeck , Jeffrey M. Rosen , Xiang H.-F. Zhang , Chad A. Shaw , Fergus J. Couch , Thomas F. Westbrook Defining the molecular networks that drive breast cancer has led to therapeutic interventions and improved patient survival. However, the aggressive triple-negative breast cancer subtype (TNBC) remains recalcitrant to targeted therapies because its molecular etiology is poorly defined. In this study, we used a forward genetic screen to discover an oncogenic network driving human TNBC. S CYL1, T EX14, and P LK1 (“STP axis”) cooperatively trigger degradation of the REST tumor suppressor protein, a frequent event in human TNBC. The STP axis induces REST degradation by phosphorylating a conserved REST phospho-degron and bridging REST interaction with the ubiquitin-ligase βTRCP. Inhibition of the STP axis leads to increased REST protein levels and impairs TNBC transformation, tumor progression, and metastasis. Expression of the STP axis correlates with low REST protein levels in human TNBCs and poor clinical outcome for TNBC patients. Our findings demonstrate that the STP-REST axis is a molecular driver of human TNBC. Graphical abstract Teaser Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype for which the molecular drivers are poorly understood. Karlin et al. now demonstrate that S CYL1, T EX14, and P LK1 (“STP axis”) cooperatively trigger degradation of the REST tumor suppressor protein, a frequent event that may provide a therapeutic entry point for human TNBC.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Christina L. Zheng , Nicholas J. Wang , Jongsuk Chung , Homayoun Moslehi , J. Zachary Sanborn , Joseph S. Hur , Eric A. Collisson , Swapna S. Vemula , Agne Naujokas , Kami E. Chiotti , Jeffrey B. Cheng , Hiva Fassihi , Andrew J. Blumberg , Celeste V. Bailey , Gary M. Fudem , Frederick G. Mihm , Bari B. Cunningham , Isaac M. Neuhaus , Wilson Liao , Dennis H. Oh , James E. Cleaver , Philip E. LeBoit , Joseph F. Costello , Alan R. Lehmann , Joe W. Gray , Paul T. Spellman , Sarah T. Arron , Nam Huh , Elizabeth Purdom , Raymond J. Cho Somatic mutations in cancer are more frequent in heterochromatic and late-replicating regions of the genome. We report that regional disparities in mutation density are virtually abolished within transcriptionally silent genomic regions of cutaneous squamous cell carcinomas (cSCCs) arising in an XPC −/− background. XPC −/− cells lack global genome nucleotide excision repair (GG-NER), thus establishing differential access of DNA repair machinery within chromatin-rich regions of the genome as the primary cause for the regional disparity. Strikingly, we find that increasing levels of transcription reduce mutation prevalence on both strands of gene bodies embedded within H3K9me3-dense regions, and only to those levels observed in H3K9me3-sparse regions, also in an XPC-dependent manner. Therefore, transcription appears to reduce mutation prevalence specifically by relieving the constraints imposed by chromatin structure on DNA repair. We model this relationship among transcription, chromatin state, and DNA repair, revealing a new, personalized determinant of cancer risk. Graphical abstract Teaser Zheng et al. report that variable mutation densities within cancer genomes result from differential access of DNA repair machinery, imposed by chromatin state. By showing that transcription restores DNA repair to tightly packaged DNA, their study reveals natural differences in expression level as a potentially important modulator of oncogene mutation rate.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Ana Freije , Rut Molinuevo , Laura Ceballos , Marta Cagigas , Pilar Alonso-Lecue , René Rodriguez , Pablo Menendez , Daniel Aberdam , Ernesto De Diego , Alberto Gandarillas Tumor suppressor p53 is a major cellular guardian of genome integrity, and its inactivation is the most frequent genetic alteration in cancer, rising up to 80% in squamous cell carcinoma (SCC). By adapting the small hairpin RNA (shRNA) technology, we inactivated endogenous p53 in primary epithelial cells from the epidermis of human skin. We show that either loss of endogenous p53 or overexpression of a temperature-sensitive dominant-negative conformation triggers a self-protective differentiation response, resulting in cell stratification and expulsion. These effects follow DNA damage and exit from mitosis without cell division. p53 preserves the proliferative potential of the stem cell compartment and limits the power of proto-oncogene MYC to drive cell cycle stress and differentiation. The results provide insight into the role of p53 in self-renewal homeostasis and help explain why p53 mutations do not initiate skin cancer but increase the likelihood that cancer cells will appear. Graphical abstract Teaser The p53 tumor suppressor is frequently inactivated in squamous cell carcinoma, yet loss of p53 does not initiate nonmelanoma skin cancer, suggesting that epithelial skin cells have self-protective mechanisms. Freije et al. show that p53 enhances proliferation and inhibits differentiation in keratinocytes by preventing endoreplication. p53 loss leads to squamous differentiation and expulsion of mutant cells, which may confer the epidermis with a molecular protective response.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Andreas Gewies , Oliver Gorka , Hanna Bergmann , Konstanze Pechloff , Franziska Petermann , Katharina M. Jeltsch , Martina Rudelius , Mark Kriegsmann , Wilko Weichert , Marion Horsch , Johannes Beckers , Wolfgang Wurst , Mathias Heikenwalder , Thomas Korn , Vigo Heissmeyer , Jürgen Ruland The paracaspase Malt1 is a central regulator of antigen receptor signaling that is frequently mutated in human lymphoma. As a scaffold, it assembles protein complexes for NF-κB activation, and its proteolytic domain cleaves negative NF-κB regulators for signal enforcement. Still, the physiological functions of Malt1-protease are unknown. We demonstrate that targeted Malt1-paracaspase inactivation induces a lethal inflammatory syndrome with lymphocyte-dependent neurodegeneration in vivo. Paracaspase activity is essential for regulatory T cell (Treg) and innate-like B cell development, but it is largely dispensable for overcoming Malt1-dependent thresholds for lymphocyte activation. In addition to NF-κB inhibitors, Malt1 cleaves an entire set of mRNA stability regulators, including Roquin-1, Roquin-2, and Regnase-1, and paracaspase inactivation results in excessive interferon gamma (IFNγ) production by effector lymphocytes that drive pathology. Together, our results reveal distinct threshold and modulatory functions of Malt1 that differentially control lymphocyte differentiation and activation pathways and demonstrate that selective paracaspase blockage skews systemic immunity toward destructive autoinflammation. Graphical abstract Teaser The paracaspase Malt1 is a key regulator of antigen receptor signaling and frequently mutated in human lymphoma. Gewies et al. demonstrate that the proteolytic function of Malt1 is largely dispensable for lymphocyte activation but critical for regulatory T cell (Treg) and innate-like B cell development and protection from IFNγ-mediated autoinflammation in vivo.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Anne Biton , Isabelle Bernard-Pierrot , Yinjun Lou , Clémentine Krucker , Elodie Chapeaublanc , Carlota Rubio-Pérez , Nuria López-Bigas , Aurélie Kamoun , Yann Neuzillet , Pierre Gestraud , Luca Grieco , Sandra Rebouissou , Aurélien de Reyniès , Simone Benhamou , Thierry Lebret , Jennifer Southgate , Emmanuel Barillot , Yves Allory , Andrei Zinovyev , François Radvanyi Extracting relevant information from large-scale data offers unprecedented opportunities in cancerology. We applied independent component analysis (ICA) to bladder cancer transcriptome data sets and interpreted the components using gene enrichment analysis and tumor-associated molecular, clinicopathological, and processing information. We identified components associated with biological processes of tumor cells or the tumor microenvironment, and other components revealed technical biases. Applying ICA to nine cancer types identified cancer-shared and bladder-cancer-specific components. We characterized the luminal and basal-like subtypes of muscle-invasive bladder cancers according to the components identified. The study of the urothelial differentiation component, specific to the luminal subtypes, showed that a molecular urothelial differentiation program was maintained even in those luminal tumors that had lost morphological differentiation. Study of the genomic alterations associated with this component coupled with functional studies revealed a protumorigenic role for PPARG in luminal tumors. Our results support the inclusion of ICA in the exploitation of multiscale data sets. Graphical abstract Teaser Extracting biological insights from large-scale data is both a challenge and an opportunity. Biton et al. now analyze bladder tumor transcriptomes. An enrichment analysis of contributing genes combined with molecular and clinical annotations of tumor samples identifies biologically relevant components, some of which are shared with other carcinoma types.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Malgorzata Gozdecka , Stephen Lyons , Saki Kondo , Janet Taylor , Yaoyong Li , Jacek Walczynski , Gerald Thiel , Wolfgang Breitwieser , Nic Jones JNK and p38 phosphorylate a diverse set of substrates and, consequently, can act in a context-dependent manner to either promote or inhibit tumor growth. Elucidating the functions of specific substrates of JNK and p38 is therefore critical for our understanding of these kinases in cancer. ATF2 is a phosphorylation-dependent transcription factor and substrate of both JNK and p38. Here, we show ATF2 suppresses tumor formation in an orthotopic model of liver cancer and cellular transformation in vitro. Furthermore, we find that suppression of tumorigenesis by JNK requires ATF2. We identify a transcriptional program activated by JNK via ATF2 and provide examples of JNK- and ATF2-dependent genes that block cellular transformation. Significantly, we also show that ATF2-dependent gene expression is frequently downregulated in human cancers, indicating that amelioration of JNK-ATF2-mediated suppression may be a common event during tumor development. Graphical abstract Teaser Given the large number of substrates phosphorylated by stress-activated kinases, identifying key effectors of their antitumorigenic function is a major challenge. Gozdecka et al. provide evidence that ATF2 mediates the tumor-suppressive effect of JNK through a transcriptional program that is frequently downregulated in human tumors.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Shaheen Kabir , Dirk Hockemeyer , Titia de Lange The conserved protein Rap1 functions at telomeres in fungi, protozoa, and vertebrates. Like yeast Rap1, human Rap1 has been implicated in telomere length regulation and repression of nonhomologous end-joining (NHEJ) at telomeres. However, mouse telomeres lacking Rap1 do not succumb to NHEJ. To determine the functions of human Rap1, we generated several transcription activator-like effector nuclease (TALEN)-mediated human cell lines lacking Rap1. Loss of Rap1 did not affect the other components of shelterin, the modification of telomeric histones, the subnuclear position of telomeres, or the 3′ telomeric overhang. Telomeres lacking Rap1 did not show a DNA damage response, NHEJ, or consistent changes in their length, indicating that Rap1 does not have an important function in protection or length regulation of human telomeres. As human Rap1, like its mouse and unicellular orthologs, affects gene expression, we propose that the conservation of Rap1 reflects its role in transcriptional regulation rather than a function at telomeres. Graphical abstract Teaser Kabir et al. now employ a TALEN-mediated genome editing strategy to make Rap1 knockouts in a diverse array of human cell lines and test the function of this conserved telomere protein. They find that, unlike its yeast counterparts, mammalian Rap1 is not essential for telomere protection or length regulation, but its role in transcriptional regulation is maintained, indicating why Rap1 remains so conserved.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Carrie L. Simms , Benjamin H. Hudson , John W. Mosior , Ali S. Rangwala , Hani S. Zaher Chemical damage to RNA affects its functional properties and thus may pose a significant hurdle to the translational apparatus; however, the effects of damaged mRNA on the speed and accuracy of the decoding process and their interplay with quality-control processes are not known. Here, we systematically explore the effects of oxidative damage on the decoding process using a well-defined bacterial in vitro translation system. We find that the oxidative lesion 8-oxoguanosine (8-oxoG) reduces the rate of peptide-bond formation by more than three orders of magnitude independent of its position within the codon. Interestingly, 8-oxoG had little effect on the fidelity of the selection process, suggesting that the modification stalls the translational machinery. Consistent with these findings, 8-oxoG mRNAs were observed to accumulate and associate with polyribosomes in yeast strains in which no-go decay is compromised. Our data provide compelling evidence that mRNA-surveillance mechanisms have evolved to cope with damaged mRNA. Graphical abstract Teaser Oxidative damage to RNA has received relatively little attention despite evidence that it can accumulate in cells and is associated with numerous disease states. Simms et al. demonstrate that a single modified residue in an mRNA can lead to ribosomal stalling. Cells in which no-go decay is compromised show increased levels of 8-oxoG mRNA, suggesting that mRNA surveillance mechanisms may have evolved to cope with damaged mRNA.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Lawrence D. Gaspers , Paula J. Bartlett , Antonio Politi , Paul Burnett , Walson Metzger , Jane Johnston , Suresh K. Joseph , Thomas Höfer , Andrew P. Thomas Receptor-mediated oscillations in cytosolic Ca 2+ concentration ([Ca 2+ ] i ) could originate either directly from an autonomous Ca 2+ feedback oscillator at the inositol 1,4,5-trisphosphate (IP 3 ) receptor or as a secondary consequence of IP 3 oscillations driven by Ca 2+ feedback on IP 3 metabolism. It is challenging to discriminate these alternatives, because IP 3 fluctuations could drive Ca 2+ oscillations or could just be a secondary response to the [Ca 2+ ] i spikes. To investigate this problem, we constructed a recombinant IP 3 buffer using type-I IP 3 receptor ligand-binding domain fused to GFP (GFP-LBD), which buffers IP 3 in the physiological range. This IP 3 buffer slows hormone-induced [IP 3 ] dynamics without changing steady-state [IP 3 ]. GFP-LBD perturbed [Ca 2+ ] i oscillations in a dose-dependent manner: it decreased both the rate of [Ca 2+ ] i rise and the speed of Ca 2+ wave propagation and, at high levels, abolished [Ca 2+ ] i oscillations completely. These data, together with computational modeling, demonstrate that IP 3 dynamics play a fundamental role in generating [Ca 2+ ] i oscillations and waves. Graphical abstract Teaser Gaspers et al. use a genetically encoded IP 3 buffer to suppress IP 3 dynamics during hormonal stimulation. Using this approach, they find that positive feedback of Ca 2+ on IP 3 formation is an essential component, generating long-period, baseline-separated Ca 2+ oscillations and intracellular Ca 2+ waves.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Panos Roussos , Amanda C. Mitchell , Georgios Voloudakis , John F. Fullard , Venu M. Pothula , Jonathan Tsang , Eli A. Stahl , Anastasios Georgakopoulos , Douglas M. Ruderfer , Alexander Charney , Yukinori Okada , Katherine A. Siminovitch , Jane Worthington , Leonid Padyukov , Lars Klareskog , Peter K. Gregersen , Robert M. Plenge , Soumya Raychaudhuri , Menachem Fromer , Shaun M. Purcell , Kristen J. Brennand , Nikolaos K. Robakis , Eric E. Schadt , Schahram Akbarian , Pamela Sklar A large portion of common variant loci associated with genetic risk for schizophrenia reside within noncoding sequence of unknown function. Here, we demonstrate promoter and enhancer enrichment in schizophrenia variants associated with expression quantitative trait loci (eQTL). The enrichment is greater when functional annotations derived from the human brain are used relative to peripheral tissues. Regulatory trait concordance analysis ranked genes within schizophrenia genome-wide significant loci for a potential functional role, based on colocalization of a risk SNP, eQTL, and regulatory element sequence. We identified potential physical interactions of noncontiguous proximal and distal regulatory elements. This was verified in prefrontal cortex and -induced pluripotent stem cell–derived neurons for the L-type calcium channel ( CACNA1C ) risk locus. Our findings point to a functional link between schizophrenia-associated noncoding SNPs and 3D genome architecture associated with chromosomal loopings and transcriptional regulation in the brain. Graphical abstract Teaser Roussos et al. find that schizophrenia risk variants are enriched for alleles that affect gene expression and lie within promoters or enhancers. For the L-type calcium channel ( CACNA1C ), the risk variant is associated with transcriptional regulation in the brain and is positioned within an enhancer sequence that physically interacts though chromosome loops with the promoter region of the gene.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Denise Cook , Erin Nuro , Emma V. Jones , Haider F. Altimimi , W. Todd Farmer , Valentina Gandin , Edith Hanna , Ruiting Zong , Alessandro Barbon , David L. Nelson , Ivan Topisirovic , Joseph Rochford , David Stellwagen , Jean-Claude Béïque , Keith K. Murai Translational control of mRNAs allows for rapid and selective changes in synaptic protein expression that are required for long-lasting plasticity and memory formation in the brain. Fragile X Related Protein 1 (FXR1P) is an RNA-binding protein that controls mRNA translation in nonneuronal cells and colocalizes with translational machinery in neurons. However, its neuronal mRNA targets and role in the brain are unknown. Here, we demonstrate that removal of FXR1P from the forebrain of postnatal mice selectively enhances long-term storage of spatial memories, hippocampal late-phase long-term potentiation (L-LTP), and de novo GluA2 synthesis. Furthermore, FXR1P binds specifically to the 5′ UTR of GluA2 mRNA to repress translation and limit the amount of GluA2 that is incorporated at potentiated synapses. This study uncovers a mechanism for regulating long-lasting synaptic plasticity and spatial memory formation and reveals an unexpected divergent role of FXR1P among Fragile X proteins in brain plasticity. Graphical abstract Teaser Control over protein synthesis is important for long-lasting plasticity and memory storage in the brain. Cook, Nuro, et al. now reveal that the RNA-binding protein FXR1P acts as a molecular brake that limits synthesis and synaptic incorporation of the AMPAR subunit GluA2, ultimately constraining long-term plasticity and memory formation.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Fengtao Su , Shibani Mukherjee , Yanyong Yang , Eiichiro Mori , Souparno Bhattacharya , Junya Kobayashi , Steven M. Yannone , David J. Chen , Aroumougame Asaithamby WRN, the protein defective in Werner syndrome (WS), is a multifunctional nuclease involved in DNA damage repair, replication, and genome stability maintenance. It was assumed that the nuclease activities of WRN were critical for these functions. Here, we report a nonenzymatic role for WRN in preserving nascent DNA strands following replication stress. We found that lack of WRN led to shortening of nascent DNA strands after replication stress. Furthermore, we discovered that the exonuclease activity of MRE11 was responsible for the shortening of newly replicated DNA in the absence of WRN. Mechanistically, the N-terminal FHA domain of NBS1 recruits WRN to replication-associated DNA double-stranded breaks to stabilize Rad51 and to limit the nuclease activity of its C-terminal binding partner MRE11. Thus, this previously unrecognized nonenzymatic function of WRN in the stabilization of nascent DNA strands sheds light on the molecular reason for the origin of genome instability in WS individuals. Graphical abstract Teaser Su et al. uncover a nonenzymatic function for WRN in DNA replication, giving insight into the molecular origin of genome instability in Werner syndrome individuals. The authors find that WRN recruitment to replication-associated DNA double-strand breaks prevents excessive MRE11-mediated degradation of nascent DNA strands.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): David A. Gorczyca , Susan Younger , Shan Meltzer , Sung Eun Kim , Li Cheng , Wei Song , Hye Young Lee , Lily Yeh Jan , Yuh Nung Jan A major gap in our understanding of sensation is how a single sensory neuron can differentially respond to a multitude of different stimuli (polymodality), such as propio- or nocisensation. The prevailing hypothesis is that different stimuli are transduced through ion channels with diverse properties and subunit composition. In a screen for ion channel genes expressed in polymodal nociceptive neurons, we identified Ppk26, a member of the trimeric degenerin/epithelial sodium channel (DEG/ENaC) family, as being necessary for proper locomotion behavior in Drosophila larvae in a mutually dependent fashion with coexpressed Ppk1, another member of the same family. Mutants lacking Ppk1 and Ppk26 were defective in mechanical, but not thermal, nociception behavior. Mutants of Piezo, a channel involved in mechanical nociception in the same neurons, did not show a defect in locomotion, suggesting distinct molecular machinery for mediating locomotor feedback and mechanical nociception. Graphical abstract Teaser Using a screen for ion channels in polymodal nociceptive neurons, Gorczyca et al. identify Ppk26, a member of the trimeric DEG/ENaC channel family, as necessary for both proper locomotive behavior and mechanical, but not thermal, nociception.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Anindya Chatterjee , Joydeep Ghosh , Baskar Ramdas , Raghuveer Singh Mali , Holly Martin , Michihiro Kobayashi , Sasidhar Vemula , Victor H. Canela , Emily R. Waskow , Valeria Visconte , Ramon V. Tiu , Catherine C. Smith , Neil Shah , Kevin D. Bunting , H. Scott Boswell , Yan Liu , Rebecca J. Chan , Reuben Kapur Oncogenic mutations of FLT3 and KIT receptors are associated with poor survival in patients with acute myeloid leukemia (AML) and myeloproliferative neoplasms (MPNs), and currently available drugs are largely ineffective. Although Stat5 has been implicated in regulating several myeloid and lymphoid malignancies, how precisely Stat5 regulates leukemogenesis, including its nuclear translocation to induce gene transcription, is poorly understood. In leukemic cells, we show constitutive activation of focal adhesion kinase (FAK) whose inhibition represses leukemogenesis. Downstream of FAK, activation of Rac1 is regulated by RacGEF Tiam1, whose inhibition prolongs the survival of leukemic mice. Inhibition of the Rac1 effector PAK1 prolongs the survival of leukemic mice in part by inhibiting the nuclear translocation of Stat5. These results reveal a leukemic pathway involving FAK/Tiam1/Rac1/PAK1 and demonstrate an essential role for these signaling molecules in regulating the nuclear translocation of Stat5 in leukemogenesis. Graphical abstract Teaser A significant impediment in treatment of leukemia, induced by oncogenic FLT3 and KIT receptors, is inadequate understanding of critical signaling pathways that lead to the development of this disease. In this study, Chatterjee et al. show an essential role of FAK/Tiam1/Rac1/PAK1 pathway in regulating nuclear translocation of Stat5 leading to leukemogenesis, in the context of oncogenic mutations of FLT3 and KIT, and provide multiple potential therapeutic targets to treat leukemia.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Shogo Tanabe , Toshihide Yamashita Multiple sclerosis (MS) is a chronic autoimmune disease characterized by inflammation, demyelination, and neurodegeneration in the CNS. Although it is important to prevent neurodegeneration for alleviating neurological disability, the molecular mechanism of neurodegeneration remains largely unknown. Here, we report that repulsive guidance molecule-a (RGMa), known to regulate axonal growth, is associated with neurodegeneration in experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. RGMa is highly expressed in interleukin-17-producing CD4 + T cells (Th17 cells). We induced EAE by adoptive transfer of myelin oligodendrocyte glycoprotein (MOG)-specific Th17 cells and then inhibited RGMa with a neutralizing antibody. Inhibition of RGMa improves EAE scores and reduces neuronal degeneration without altering immune or glial responses. Th17 cells induce cultured cortical neuron death through RGMa-neogenin and Akt dephosphorylation. Our results demonstrate that RGMa is involved in Th17-cell-mediated neurodegeneration and that RGMa-specific antibody may have a therapeutic effect in MS. Graphical abstract Teaser The mechanism of neurodegeneration under inflammation in the CNS remains largely unknown. Tanabe and Yamashita demonstrate that RGMa is highly expressed in Th17 cells and induces dephosphorylation of Akt, leading to death of neurons. A neutralizing antibody to RGMa attenuates axonal degeneration and severity of Th17-cell-mediated autoimmune encephalomyelitis.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Patricia A. Possik , Judith Müller , Carmen Gerlach , Juliana C.N. Kenski , Xinyao Huang , Aida Shahrabi , Oscar Krijgsman , Ji-Ying Song , Marjon A. Smit , Bram Gerritsen , Cor Lieftink , Kristel Kemper , Magali Michaut , Roderick L. Beijersbergen , Lodewyk Wessels , Ton N. Schumacher , Daniel S. Peeper To identify factors preferentially necessary for driving tumor expansion, we performed parallel in vitro and in vivo negative-selection short hairpin RNA (shRNA) screens. Melanoma cells harboring shRNAs targeting several DNA damage response (DDR) kinases had a greater selective disadvantage in vivo than in vitro, indicating an essential contribution of these factors during tumor expansion. In growing tumors, DDR kinases were activated following hypoxia. Correspondingly, depletion or pharmacologic inhibition of DDR kinases was toxic to melanoma cells, including those that were resistant to BRAF inhibitor, and this could be enhanced by angiogenesis blockade. These results reveal that hypoxia sensitizes melanomas to targeted inhibition of the DDR and illustrate the utility of in vivo shRNA dropout screens for the identification of pharmacologically tractable targets. Graphical abstract Teaser Specific parameters lacking in vitro, but present in vivo, influence tumor behavior and therapeutic dependencies. Possik et al. now perform parallel in vitro and in vivo negative-selection screens and uncover a critical requirement for DNA damage response kinases in expanding tumors, a liability that could be exploited pharmacologically.

Description: Publication date: 20 November 2014 Source: Cell Reports, Volume 9, Issue 4 Author(s): Debabrata Panja , Justin W. Kenney , Laura D’Andrea , Francesca Zalfa , Anni Vedeler , Karin Wibrand , Rikiro Fukunaga , Claudia Bagni , Christopher G. Proud , Clive R. Bramham BDNF signaling contributes to protein-synthesis-dependent synaptic plasticity, but the dynamics of TrkB signaling and mechanisms of translation have not been defined. Here, we show that long-term potentiation (LTP) consolidation in the dentate gyrus of live rodents requires sustained (hours) BDNF-TrkB signaling. Surprisingly, this sustained activation maintains an otherwise labile signaling pathway from TrkB to MAP-kinase-interacting kinase (MNK). MNK activity promotes eIF4F translation initiation complex formation and protein synthesis in mechanistically distinct early and late stages. In early-stage translation, MNK triggers release of the CYFIP1/FMRP repressor complex from the 5′-mRNA cap. In late-stage translation, MNK regulates the canonical translational repressor 4E-BP2 in a synapse-compartment-specific manner. This late stage is coupled to MNK-dependent enhanced dendritic mRNA translation. We conclude that LTP consolidation in the dentate gyrus is mediated by sustained BDNF signaling to MNK and MNK-dependent regulation of translation in two functionally and mechanistically distinct stages. Graphical abstract Teaser The logic of translational control in synaptic plasticity is not well understood. Panja et al. show that long-term potentiation in the dentate gyrus of live rodents is a two-stage process driven by brain-derived neurotrophic factor signaling to MAP-kinase-interacting kinase and activation of functionally and mechanistically distinct forms of translation.

Description: Capture–mark–recapture (CMR) approaches are the backbone of many studies in population ecology to gain insight on the life cycle, migration, habitat use, and demography of target species. The reliable and repeatable recognition of an individual throughout its lifetime is the basic requirement of a CMR study. Although invasive techniques are available to mark individuals permanently, noninvasive methods for individual recognition mainly rest on photographic identification of external body markings, which are unique at the individual level. The re-identification of an individual based on comparing shape patterns of photographs by eye is commonly used. Automated processes for photographic re-identification have been recently established, but their performance in large datasets (i.e., 〉 1000 individuals) has rarely been tested thoroughly. Here, we evaluated the performance of the program AMPHIDENT, an automatic algorithm to identify individuals on the basis of ventral spot patterns in the great crested newt ( Triturus cristatus ) versus the genotypic fingerprint of individuals based on highly polymorphic microsatellite loci using GENECAP. Between 2008 and 2010, we captured, sampled and photographed adult newts and calculated for 1648 samples/photographs recapture rates for both approaches. Recapture rates differed slightly with 8.34% for GENECAP and 9.83% for AMPHIDENT. With an estimated rate of 2% false rejections (FRR) and 0.00% false acceptances (FAR), AMPHIDENT proved to be a highly reliable algorithm for CMR studies of large datasets. We conclude that the application of automatic recognition software of individual photographs can be a rather powerful and reliable tool in noninvasive CMR studies for a large number of individuals. Because the cross-correlation of standardized shape patterns is generally applicable to any pattern that provides enough information, this algorithm is capable of becoming a single application with broad use in CMR studies for many species. Capture-mark-recapture (CMR) of individuals is the backbone of many studies in population ecology. Automated processes for photographic re-identification of individuals have been recently established but their performance in large datasets have rarely been tested thoroughly. By applying genetic fingerprinting we show that cross-correlated automatic photo-identification of individuals is highly efficient in large capture-mark–recapture studies.

Description: Predator community composition can alter habitat quality for prey by changing the strength and direction of consumptive effects. Whether predator community composition also alters prey density via nonconsumptive effects during habitat selection is not well known, but is important for understanding how changes to predator communities will alter prey populations. We tested the hypothesis that predator community composition (presence of caged trout, caged dragonflies, or caged trout + dragonflies) alters colonization of aquatic mesocosms by ovipositing aquatic insects. In a previous experiment in this system, we found a spatial contagion effect, in which insects avoided pools with predators, but only when predator-free pools were isolated (~5 m away from predator pools). Here, we removed the isolated predator-free pools, allowing us to test whether insects would make fine-scale (~1 m) oviposition decisions in the absence of preferred isolated pools. We also estimated consumptive effects by allowing predators to feed on colonists for 5 days following colonization. All insects collected after 21 days were dipterans, dominated by Chironomidae. Total colonization, measured as the number of developing larvae after 21 days, was not affected by either predator presence or composition. Consumption was significant in the trout only treatment, reducing larval insect density by 46 ± 37% (mean ± SE). No other predator treatment significantly reduced prey density, although the proportion of chironomid larvae in protective cases increased in response to direct predation from dragonflies, indicating an antipredatory behavioral response. Taken together, these results reveal that predator community composition altered larval survival and behavior, but colonizing females either did not or could not assess these risks across small scales during oviposition. We found no evidence that predatory trout or dragonflies altered colonization of adjacent mesocosms by aquatic dipterans (i.e., larvae of the adult Chironomid pictured above). However, both trout and dragonflies (but not trout + dragonflies) had consumptive effects on the colonized larval community. We interpret our results in light of previous studies as a function of (1) a failure of colonizing insects to differentiate predator risk at small scales; and (2) possible spatial contagion effects that override colonization decisions at small scales.

Description: Identification of units within species worthy of separate management consideration is an important area within conservation. Mitochondrial DNA (mtDNA) surveys can potentially contribute to this by identifying phylogenetic and population structure below the species level. The American crocodile ( Crocodylus acutus ) is broadly distributed throughout the Neotropics. Its numbers have been reduced severely with the species threatened throughout much of its distribution. In Colombia, the release of individuals from commercial captive populations has emerged as a possible conservation strategy that could contribute to species recovery. However, no studies have addressed levels of genetic differentiation or diversity within C. acutus in Colombia, thus complicating conservation and management decisions. Here, sequence variation was studied in mtDNA cytochrome b and cytochrome oxidase I gene sequences in three Colombian captive populations of C. acutus . Two distinct lineages were identified: C. acutus -I, corresponding to haplotypes from Colombia and closely related Central American haplotypes; and C. acutus -II, corresponding to all remaining haplotypes from Colombia. Comparison with findings from other studies indicates the presence of a single “northern” lineage (corresponding to C. acutus -I) distributed from North America (southern Florida), through Central America and into northern South America. The absence of C. acutus -II haplotypes from North and Central America indicates that the C. acutus -II lineage probably represents a separate South American lineage. There appears to be sufficient divergence between lineages to suggest that they could represent two distinct evolutionary units. We suggest that this differentiation needs to be recognized for conservation purposes because it clearly contributes to the overall genetic diversity of the species. All Colombian captive populations included in this study contained a mixture of representatives of both lineages. As such, we recommend against the use of captive-bred individuals for conservation strategies until further genetic information is available.

Description: Marine organisms are simultaneously exposed to anthropogenic stressors with likely interactive effects, including synergisms in which the combined effects of multiple stressors are greater than the sum of individual effects. Early life stages of marine organisms are potentially vulnerable to the stressors associated with global change, but identifying general patterns across studies, species and response variables is challenging. This review represents the first meta-analysis of multi-stressor studies to target early marine life stages (embryo to larvae), particularly between temperature, salinity and pH as these are the best studied. Knowledge gaps in research on multiple abiotic stressors and early life stages are also identified. The meta-analysis yielded several key results: 1) Synergistic interactions (65% of individual tests) are more common than additive (17%) or antagonistic (17%) interactions. 2) Larvae are generally more vulnerable than embryos to thermal and pH stress. 3) Survival is more likely than sub-lethal responses to be affected by thermal, salinity, and pH stress. 4) Interaction types vary among stressors, ontogenetic stages, and biological responses, but they are more consistent among phyla. 5) Ocean acidification is a greater stressor for calcifying than non-calcifying larvae. Although more ecologically realistic than single-factor studies, multifactorial studies may still oversimplify complex systems, and so meta-analyses of the data from them must be cautiously interpreted with regard to extrapolation to field conditions. Nonetheless our results identify taxa with early life stages that may be particularly vulnerable (e.g. molluscs, echinoderms) or robust (e.g. arthropods, cnidarians) to abiotic stress. We provide a list of recommendations for future multiple stressor studies, particularly those focussed on early marine life stages. This article is protected by copyright. All rights reserved.

Description: Questions What are the driving factors for the species richness of understorey herbs of isolated forest patches? How can the relative direct and indirect effects of patch area, site quality, habitat isolation and continuity be disentangled? What are the effects on different species groups? Location Eutrophic hardwood forest patches in northwestern Germany. The isolation of these patches results from extensive agricultural land use and prevailing poor soil conditions in the surrounding landscape. Thus, the patches are embedded within a matrix of both open habitats and species-poor acidic forests. Methods We surveyed 76 eutrophic forest patches and recorded all understorey herb species. Several measured site variables were divided into soil and structural components, and into variables describing mean site conditions and site heterogeneity. Partial least squares structural equation modelling (PLS-SEM) was employed to determine the direct and indirect effects of patch area, site heterogeneity, mean site conditions, isolation and habitat continuity on species richness of several species groups. Results The most important factor determining species richness was patch area, which showed the strongest direct effect on eutrophic forest specialist species. Richness of forest matrix species, which also occur in the surrounding forest areas, was not directly influenced by area. Soil heterogeneity as well as structural heterogeneity increased with area, but only soil heterogeneity was relevant for species richness. Soil heterogeneity was more important for forest species in general and stress tolerators than for generalist species, open habitat species, competitors and ruderals. Mean site conditions influenced species richness of generalists, open habitat species, competitors and nutrient-demanding species. Isolation had a (negative) effect only on habitat specialists. Conclusions The relative effects of driving factors varied considerably between species groups. From our results for eutrophic forest specialists, we conclude that large and well-connected habitat patches are especially important for the successful conservation of the unique and partly endangered flora of eutrophic hardwood forests. We determined the relative effects of driving factors on species richness of eutrophic forest patches and found considerable differences between different species groups. Richness of specialists was driven by soil heterogeneity, direct area effects and isolation whereas these factors hardly affected generalists and open habitat species. We conclude that large, well-connected habitat patches play an important role with respect to conservation.

Description: Ramathal et al. have employed an elegant xenotransplantation technique to study the fate of human induced pluripotent stem cells (hiPSCs) from fertile males and from males carrying Y chromosome deletions of the azoospermia factor ( AZF ) region. When placed in a mouse testis niche, hiPSCs from fertile males differentiate into germ cell-like cells (GCLCs). Highlighting the crucial role of cell autonomous factors in male sterility, hiPSCs derived from azoospermic males prove to be less successful under similar circumstances. Their studies argue that the agametic “Sertoli cell only” phenotype of two of the AZF deletions likely arises from a defect in the maintenance of germline stem cells (GSCs) rather than from a defect in their specification. These observations underscore the importance of the dialogue between the somatic niche and its inhabitant stem cells, and open up interesting questions concerning the functioning of the somatic niche and how it communicates to the GSCs. In Ramathal et al., human induced pluripotent stem cells (hiPSCs) from wild type and azoospermic men adopt germ cell fate in vitro. Upon xenotransplantation into mouse seminiferous tubules, these hiPSCs acquired germ cell-like (GCL) fate. Remarkably, samples from azoospermic males were less efficient in both contexts.

Description: Human adult stem cells, which are capable of self-renewal and differentiation into other cell types, can be isolated from various tissues. There are no ethical and rejection problems as in the case of embryonic stem cells, so they are a promising source for cell therapy. The human body contains a great amount of adipose tissue that contains high numbers of mesenchymal stem cells. Human adipose-derived stem cells (hADSCs) could be easily induced to form neuron-like cells, and because of its availability and abundance, we can use it for clinical cell therapy. On the other hand, T 3 hormone as a known neurotropic factor has important impressions on the nervous system. The aim of this study was to explore the effects of T 3 treatment on neural differentiation of hADSCs. ADSCs were harvested from human adipose tissue, after neurosphere formation, and during final differentiation, treatment with T 3 was performed. Immunocytochemistry, real-time RT-PCR, Western blotting techniques were used for detection of nestin, MAP2, and GFAP markers in order to confirm the effects of T 3 on neural differentiation of hADSCs. Our results showed an increase in the number of glial cells but reduction in neuronal cells number fallowing T 3 treatment. Copyright © 2014 John Wiley & Sons, Ltd.

Description: Palmitate induces insulin resistance and apoptosis in insulin target tissues. Rosiglitazone (RSG), a peroxisome proliferator-activated receptor γ (PPAR γ ) agonist, can activate both pro-apoptotic and anti-apoptotic pathways in different cells; however, its effect on palmitate-induced apoptosis in skeletal muscle cells remains to be elucidated. After differentiation of C2C12 cells, myotubes were treated with palmitate, RSG and GW9662 (PPAR γ antagonist). MTT and terminal deoxynucleotide transferase dUTP nick end labelling (TUNEL) assays and caspase-3 activity were used to investigate the apoptosis. To study the underlying mechanism, glucose uptake, gene expression and protein levels were evaluated. A total of 0.75 mM palmitate reduced cell viability by 43% and increased TUNEL-positive cells and caspase-3 activity by 15-fold and 6.6-fold, respectively. RSG (10 μM) could markedly decrease the level of TUNEL-positive cells and caspase-3 activity in palmitate-treated cells. The protective effect of RSG on apoptosis was abrogated by GW9662. To investigate the molecular mechanism of this effect, gene expression and protein level of protein tyrosine phosphatase 1B (PTP1B) were evaluated. Palmitate and RSG individually increased the expression and protein level of PTP1B, whereas combined treatment (palmitate and RSG) were able to further increase the expression of PTP1B in C2C12 cells. We also evaluated the effect of RSG on palmitate-induced insulin resistance in muscle cells. RSG could significantly improve glucose uptake by 0.4-fold in myotubes treated with palmitate. Moreover, RSG could restore the phosphorylation of Akt in palmitate-treated cells. These data suggest that RSG protects skeletal muscle cells against palmitate-induced apoptosis and this effect appears to be mediated via the PPAR γ -dependent and PTP1B-independent mechanisms. Copyright © 2014 John Wiley & Sons, Ltd. SIGNIFICANCE OF THE STUDY Saturated free fatty acids (FFAs), such as palmitate, have been shown to induce cellular apoptosis. Strategies for preventing the cytotoxic effect of palmitate are useful in reduction of diabetes complications. In this study, we introduced RSG as an agent that protects skeletal muscle cells against palmitate-induced apoptosis and insulin resistance. It appears that RSG protects skeletal muscle cells against palmitate-induced apoptosis via the PPARγ-dependent and PTP1B-independent mechanisms. Given the role of FFAs in skeletal muscle apoptosis, these findings support the idea that RSG can ameliorate diabetes complications such as skeletal muscle loss.

Description: The expression of Hes1 is increased following myocardial infarct and other ischemic cardiomyopathies, but the role of Hes1 in cardioprotection provided by ischemic postconditioning (IPost) remains unclear. In this study, we used gain and loss of function approaches to investigate the role of Hes1 in cardioprotection during IPost. Primary cardiac myocytes exposed to ischemia reperfusion injury (IRI) and IPost were used as the experimental model. The results showed that Hes1 expression was increased during myocardial IPost, and Hes1 promoted the viability while inhibited the apoptosis of cardiomyocytes. Moreover, Hes1 inhibited the opening of mitochondrial permeability transition pore (mPTP) and the generation of reactive oxygen species in primary cardiac myocytes exposed to IRI. Mechanistically, we found that Hes1-mediated cardioprotection was related to the downregulation of phosphatase and tensin homolog and the activation of phosphatidylinositol 3-kinase/Akt and signal transducer and activator of transcription 3 signalling. These data demonstrate that Hes1 is upregulated and mediates cardioprotection provided by IPost and suggest that Hes1 is a potential new target for the treatment of ischemic cardiomyopathy. Copyright © 2014 John Wiley & Sons, Ltd.

Description: The aim of this study was to study RAS-siRNA blocking RAS pathway and suppressing cell growth in human oesophageal squamous cell carcinoma in nude mice. The methods in this study was to construct RAS-siRNA expression vector, establish 40 oesophageal squamous cell carcinoma xenograft animal models and divided them into five groups: control group, siRNA control group, RAS-siRNA group, paclitaxel group and RAS-siRNA and paclitaxel group. We observed tumour growth in nude mice, studied histology by HE staining, tumour growth inhibition by TUNEL assay and detected the RAS, MAPK and cyclin D1 protein expression by immunohistochemistry and western blot. We have obtained the following results: (i) successfully established animal models; (ii) nude mice in each group after treatment inhibited tumour volume was significantly reduced compared with the control group ( p  〈 0.05); (iii) compared with the control group, the number of apoptotic cells were significantly increased in the siRNA control group and the RAS-siRNA group, and the number of apoptosis cells in the paclitaxel and RAS-siRNA group is significantly most than the paclitaxel group and RAS-siRNA group ( p  〈 0.05); and (iv) after treatment, RAS, MAPK and cyclin D1 expression in five groups was decreasing gradually. After adding paclitaxel, the protein expression in the paclitaxel and RAS-siRNA group was significantly lower than that of paclitaxel group, negative control and paclitaxel group ( p  〈 0.05). We therefore conclude that RAS-siRNA can block the RAS signal transduction pathway, reduce the activity of tumour cells, arrest tumour cell cycle, promote apoptosis, inhibit cell proliferation and increase tumour cell sensitivity to chemotherapeutic drugs. Copyright © 2014 John Wiley & Sons, Ltd.

Description: There is increasing evidence suggesting that oxidative stress plays an important role in the development of many chronic and degenerative conditions such as diabetic encephalopathy and depression. Considering that diabetic rats and mice present higher depressive-like behaviour when submitted to the forced swimming test and that treatment with insulin and/or clonazepam is able to reverse the behavioural changes of the diabetic rats, the present work investigated the antioxidant status, specifically total antioxidant reactivity and antioxidant potential of insulin and clonazepam, as well as the effect of this drugs upon protein oxidative damage and reactive species formation in cortex, hippocampus and striatum from diabetic rats submitted to forced swimming test. It was verified that longer immobility time in diabetic rats and insulin plus clonazepam treatment reversed this depressive-like behaviour. Moreover, data obtained in this study allowed to demonstrate through different parameters such as protein carbonyl content, 2′7′-dichlorofluorescein oxidation, catalase, superoxide dismutase, glutathione peroxidase assay, total radical-trapping antioxidant potential and total antioxidant reactivity that there is oxidative stress in cortex, hippocampus and striatum from diabetic rats under depressive-like behaviour and highlight the insulin and/or clonazepam effect in these different brain areas, restoring antioxidant status and protein damage. Copyright © 2014 John Wiley & Sons, Ltd.