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  • 1
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    Arkadia enhances BMP signalling through ubiquitylation and degradation of Smad6 (2015)
    Oxford University Press
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    Publication Date: 2015-07-01
    Description: Arkadia, a positive regulator of Smad-dependent signalling via the transforming growth factor-β (TGF-β) family, is an E3 ubiquitin ligase that induces ubiquitylation and proteasome-dependent degradation of TGF-β suppressors such as Smad7, c-Ski and SnoN. In this study, we examined the effects of Arkadia on bone morphogenetic protein (BMP)-induced osteoblast differentiation. Knockdown of Arkadia reduced mineralization and expression of osteoblast differentiation markers. Furthermore, we showed that Smad6, a BMP-specific inhibitory Smad, is a target of Arkadia: wild-type (WT) Arkadia, but not the C937A (CA) mutant lacking E3 ubiquitin-ligase activity, induced ubiquitylation and proteasome-dependent degradation of Smad6. Accordingly, protein levels of Smad6, Smad7 and c-Ski were elevated in MEFs from Arkadia KO mice. Finally, expression of Arkadia attenuated blockade of BMP signalling by Smad6 in a transcriptional reporter assay. These results demonstrate that Smad6 is a novel target of Arkadia, and that Arkadia positively regulates BMP signalling via degradation of Smad6, Smad7 and c-Ski/SnoN.
    Print ISSN: 0021-924X
    Electronic ISSN: 1756-2651
    Topics: Biology , Chemistry and Pharmacology
    Published by Oxford University Press on behalf of The Japanese Biochemical Society (JBS).
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  • 2
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    Arkadia enhances BMP signalling through ubiquitylation and degradation of Smad6 (2015)
    Oxford University Press
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    Publication Date: 2015-03-11
    Print ISSN: 0021-924X
    Electronic ISSN: 1756-2651
    Topics: Biology , Chemistry and Pharmacology
    Published by Oxford University Press on behalf of Japanese Biochemical Society.
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  • 3
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    Characterisation of novel RUNX2 mutation with alanine tract expansion from Japanese cleidocranial dysplasia patient (2015)
    Oxford University Press
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    Publication Date: 2015-12-31
    Description: Cleidocranial dysplasia (CCD; MIM 119600) is an autosomal dominant skeletal dysplasia characterised by hypopalstic and/or aplastic clavicles, midface hypoplasia, absent or delayed closure of cranial sutures, moderately short stature, delayed eruption of permanent dentition and supernumerary teeth. The molecular pathogenesis can be explained in about two-thirds of CCD patients by haploinsufficiency of the RUNX2 gene. In our current study, we identified a novel and rare variant of the RUNX2 gene (c.181_189dupGCGGCGGCT) in a Japanese patient with phenotypic features of CCD. The insertion led an alanine tripeptide expansion (+3Ala) in the polyalanine tract. To date, a RUNX2 variant with alanine decapeptide expansion (+10Ala) is the only example of a causative variant of RUNX2 with polyalanine tract expansion to be reported, whilst RUNX2 (+1Ala) has been isolated from the healthy population. Thus, precise analyses of the RUNX2 (+3Ala) variant were needed to clarify whether the tripeptide expanded RUNX2 is a second disease-causing mutant with alanine tract expansion. We therefore investigated the biochemical properties of the mutant RUNX2 (+3Ala), which contains 20 alanine residues in the polyalanine tract. When transfected in COS7 cells, RUNX2 (+3Ala) formed intracellular ubiquitinated aggregates after 24h, and exerted a dominant negative effect in vitro . At 24h after gene transfection, whereas slight reduction was observed in RUNX2 (+10Ala), all of these mutants significantly activated osteoblast-specific element-2, a cis-acting sequence in the promoter of the RUNX2 target gene osteocalcin. The aggregation growth of RUNX2 (+3Ala) was clearly lower and slower than that of RUNX2 (+10Ala). Furthermore, we investigated several other RUNX2 variants with various alanine tract lengths, and found that the threshold for aggregation may be RUNX2 (+3Ala). We conclude that RUNX2 (+3Ala) is the cause of CCD in our current case, and that the accumulation of intracellular aggregates in vitro is related to the length of the alanine tract.
    Print ISSN: 0267-8357
    Electronic ISSN: 1464-3804
    Topics: Biology , Medicine
    Published by Oxford University Press
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  • 4
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    Acrylamide induces specific DNA adduct formation and gene mutations in a carcinogenic target site, the mouse lung (2015)
    Oxford University Press
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    Publication Date: 2015-02-17
    Description: Acrylamide (AA) is a contaminant in heated foods and is carcinogenic in multiple organs of rodents. There have been many reports regarding AA-induced DNA modification and genotoxicity. However, the data are insufficient to understand fully the relationship between the two events. A recent report demonstrated carcinogenicity in the mouse lung. The lung is advantageous for investigation of AA-induced genotoxicity because DNA adduct levels are relatively high in this organ. In the present study, reporter gene mutation assays and quantitative analyses of specific DNA adducts were performed in the lungs of mature gpt delta mice treated with AA at doses of 100, 200 and 400 p.p.m. in drinking water for 4 weeks. N7-GA-Gua was detected in all AA-treated mice in a dose-dependent manner. gpt mutant frequencies (MFs) were significantly increased in the middle- and high-dose groups. In the analysis of mutation spectra, significant increases in GC-TA transversions and single base deletion mutations were observed in the high-dose group. Spi – MFs were significantly increased in the high-dose group. Analysis of Spi – mutants revealed significant increases in the frequencies of single base deletion mutation in runs of G/C and A/T. Analyses of immature mice under the same experimental conditions showed that there were no differences of susceptibility to AA-induced genotoxicity in the two age classes. The overall data clearly show the causal relationship between AA-induced DNA adducts and the gene mutations at carcinogenic target sites.
    Print ISSN: 0267-8357
    Electronic ISSN: 1464-3804
    Topics: Biology , Medicine
    Published by Oxford University Press
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  • 5
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    Central mass profiles of the nearby cool-core galaxy clusters Hydra A and A478 (2016)
    Oxford University Press
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    Publication Date: 2016-01-21
    Description: We perform a weak-lensing study of the nearby cool-core galaxy clusters, Hydra A ( z  = 0.0538) and A478 ( z  = 0.0881), of which the brightest cluster galaxies (BCGs) host the powerful activities of active galactic nuclei (AGNs). For each cluster, the observed tangential shear profile is described well by either a single Navarro–Frenk–White model or a two-component model including the BCG as an unresolved point mass. For A478, we determine the BCG and its host-halo masses from a joint fit to weak-lensing and stellar photometry measurements. We find that the choice of initial mass functions (IMFs) can introduce a factor of 2 uncertainty in the BCG mass, whereas the BCG host-halo mass is constrained well by data. We perform a joint analysis of the weak-lensing and stellar kinematics data available for the Hydra A cluster, which allows us to constrain the central mass profile without assuming specific IMFs. We find that the central mass profile ( r  〈 300 kpc) determined from the joint analysis is in excellent agreement with those from independent measurements, including dynamical masses estimated from the cold gas disc component, X-ray hydrostatic total mass estimates, and the central stellar mass estimated with the Salpeter IMF. The observed dark matter fraction around the BCG for Hydra A is found to be smaller than those predicted by adiabatic contraction models, suggesting the importance of other physical processes, such as AGN feedback and/or dissipationless mergers.
    Print ISSN: 0035-8711
    Electronic ISSN: 1365-2966
    Topics: Physics
    Published by Oxford University Press
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  • 6
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    ALMA imprint of intergalactic dark structures in the gravitational lens SDP.81 (2016)
    Oxford University Press
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    Publication Date: 2016-02-20
    Description: We present an analysis of the Atacama Large Millimeter/submillimeter Array long baseline science verification data of the gravitational lens system SDP.81. We fit the positions of the brightest clumps at redshift z = 3.042 and a possible active galactic nucleus component of the lensing galaxy at redshift z = 0.2999 in the band 7 continuum image using a canonical lens model, a singular isothermal ellipsoid plus an external shear. Then, we measure the ratio of fluxes in some apertures at the source plane where the lensed images are inversely mapped. We find that the aperture flux ratios of band 7 continuum image are perturbed by 10–20 per cent with a significance at 2 ~ 3 level. Moreover, we measure the astrometric shifts of multiply lensed images near the caustic using the CO(8–7) line. Using a lens model best fitted to the band 7 continuum image, we reconstruct the source image of the CO(8–7) line by taking linear combination of inverted quadruply lensed images. At the 50th channel (rest-frame velocity 28.6 km s –1 ) of the CO(8–7) line, we find an imprint of astrometric shifts of the order of 0.01 arcsec in the source image. Based on a semi-analytic calculation, we find that the observed anomalous flux ratios and the astrometric shifts can be explained by intergalactic dark structures in the line of sight. A compensated homogeneous spherical clump with a mean surface mass density of the order of 10 8 M h –1 arcsec –2 can explain the observed anomaly and astrometric shifts simultaneously.
    Print ISSN: 0035-8711
    Electronic ISSN: 1365-2966
    Topics: Physics
    Published by Oxford University Press
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  • 7
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    Expression and structural characterization of anti-T-antigen single-chain antibodies (scFvs) and analysis of their binding to T-antigen by surface plasmon resonance and NMR spectroscopy (2013)
    Oxford University Press
    Details
    Publication Date: 2013-12-03
    Description: T-antigen (Galβ1-3GalNAcα-1-Ser/Thr), also known as Thomsen–Friedenreich antigen (TF antigen), is an oncofetal antigen commonly found in cancerous tissues. Availability of anti-T-antigen human antibodies could lead to the development of cancer diagnostics and therapeutics. Four groups of single-chain variable fragment (scFv) genes were previously isolated from a phage library (Matsumoto-Takasaki et al. (2009) Isolation and characterization of anti-T-antigen single chain antibodies from a phage library. BioSci Trends 3:87–95.). Here, four anti-T-antigen scFv genes belonging to Group 1–4 were expressed and produced in a Drosophila S2 cell expression system. ELISA and surface plasmon resonance (SPR) analyses confirmed the binding activity of 1E8 scFv protein to various T-antigen presenting conjugates. NMR experiments provided evidence of the folded nature of the 1E8 scFv protein. ScFv-ligand contact was identified by STD NMR, indicating that the galactose unit of T-antigen at the non-reducing end was primarily recognized by 1E8 scFv. This thus provides direct evidence of T-antigen specificity.
    Print ISSN: 0021-924X
    Electronic ISSN: 1756-2651
    Topics: Biology , Chemistry and Pharmacology
    Published by Oxford University Press on behalf of The Japanese Biochemical Society (JBS).
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  • 8
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    Cyanide-insensitive quinol oxidase (CIO) from Gluconobacter oxydans is a unique terminal oxidase subfamily of cytochrome bd (2013)
    Oxford University Press
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    Publication Date: 2013-05-30
    Description: Cyanide-insensitive terminal quinol oxidase (CIO) is a subfamily of cytochrome bd present in bacterial respiratory chain. We purified CIO from the Gluconobacter oxydans membranes and characterized its properties. The air-oxidized CIO showed some or weak peaks of reduced haemes b and of oxygenated and ferric haeme d , differing from cytochrome bd . CO- and NO-binding difference spectra suggested that haeme d serves as the ligand-binding site of CIO. Notably, the purified CIO showed an extraordinary high ubiquinol-1 oxidase activity with the pH optimum of pH 5–6. The apparent V max value of CIO was 17-fold higher than that of G. oxydans cytochrome bo 3 . In addition, compared with Escherichia coli cytochrome bd , the quinol oxidase activity of CIO was much more resistant to cyanide, but sensitive to azide. The K m value for O 2 of CIO was 7- to 10-fold larger than that of G. oxydans cytochrome bo 3 or E. coli cytochrome bd . Our results suggest that CIO has unique features attributable to the structure and properties of the O 2 -binding site, and thus forms a new sub-group distinct from cytochrome bd . Furthermore, CIO of acetic acid bacteria may play some specific role for rapid oxidation of substrates under acidic growth conditions.
    Print ISSN: 0021-924X
    Electronic ISSN: 1756-2651
    Topics: Biology , Chemistry and Pharmacology
    Published by Oxford University Press on behalf of The Japanese Biochemical Society (JBS).
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  • 9
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    Ochratoxin A induces DNA double-strand breaks and large deletion mutations in the carcinogenic target site of gpt delta rats (2013)
    Oxford University Press
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    Publication Date: 2013-12-20
    Description: Ochratoxin A (OTA) is a carcinogen targeting proximal tubules at the renal outer medulla (ROM) in rodents. We previously reported that OTA increased mutant frequencies of the red / gam gene (Spi – ), primarily deletion mutations. In the present study, Spi – assays and mutation spectrum analyses in the Spi – mutants were performed using additional samples collected in our previous study. Spi – assay results were similar to those in our previous study, revealing large (〉1kb) deletion mutations in the red/gam gene. To clarify the molecular progression from DNA damage to gene mutations, in vivo comet assays and analysis of DNA damage/repair-related mRNA and/or protein expression was performed using the ROM of gpt delta rats treated with OTA at 70, 210 or 630 µg/kg/day by gavage for 4 weeks. Western blotting and immunohistochemical staining demonstrated that OTA increased -H2AX expression specifically at the carcinogenic target site. In view of the results of comet assays, we suspected that OTA was capable of inducing double-strand breaks (DSBs) at the target sites. mRNA and/or protein expression levels of homologous recombination (HR) repair-related genes ( Rad51 , Rad18 and Brip1 ), but not nonhomologous end joining-related genes, were increased in response to OTA in a dose-dependent manner. Moreover, dramatic increases in the expression of genes involved in G 2 /M arrest ( Chek1 and Wee1 ) and S/G 2 phase ( Ccna2 and Cdk1 ) were observed, suggesting that DSBs induced by OTA were repaired predominantly by HR repair, possibly due to OTA-specific cell cycle regulation, consequently producing large deletion mutations at the carcinogenic target site.
    Print ISSN: 0267-8357
    Electronic ISSN: 1464-3804
    Topics: Biology , Medicine
    Published by Oxford University Press
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  • 10
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    PINBPA: Cytoscape app for network analysis of GWAS data (2015)
    Oxford University Press
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    Publication Date: 2015-01-10
    Description: : Protein interaction network-based pathway analysis (PINBPA) for genome-wide association studies (GWAS) has been developed as a Cytoscape app, to enable analysis of GWAS data in a network fashion. Users can easily import GWAS summary-level data, draw Manhattan plots, define blocks, prioritize genes with random walk with restart, detect enriched subnetworks and test the significance of subnetworks via a user-friendly interface. Availability and implementation: PINBPA app is freely available in Cytoscape app store. Contact: pmousavi@cs.queensu.ca and sebaran@cgl.ucsf.edu Supplementary information: Supplementary data are available at Bioinformatics online.
    Print ISSN: 1367-4803
    Electronic ISSN: 1460-2059
    Topics: Biology , Computer Science , Medicine
    Published by Oxford University Press
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