ALBERT

Description: RNA sequencing (RNAseq) has become the method of choice for transcriptome analysis, yet no consensus exists as to the most appropriate pipeline for its analysis, with current benchmarks suffering important limitations. Here, we address these challenges through a rich benchmarking resource harnessing (i) two RNAseq datasets including ERCC ExFold spike-ins; (ii) Nanostring measurements of a panel of 150 genes on the same samples; (iii) a set of internal, genetically-determined controls; (iv) a reanalysis of the SEQC dataset; and (v) a focus on relative quantification (i.e. across-samples). We use this resource to compare different approaches to each step of RNAseq analysis, from alignment to differential expression testing. We show that methods providing the best absolute quantification do not necessarily provide good relative quantification across samples, that count-based methods are superior for gene-level relative quantification, and that the new generation of pseudo-alignment-based software performs as well as established methods, at a fraction of the computing time. We also assess the impact of library type and size on quantification and differential expression analysis. Finally, we have created a R package and a web platform to enable the simple and streamlined application of this resource to the benchmarking of future methods.

Description: Lying below Vatnajökull ice cap in Iceland, Bárðarbunga stratovolcano began experiencing wholesale caldera collapse in 2014 August 16, one of the largest such events recorded in the modern instrumental era. Simultaneous with this collapse is the initiation of a plate boundary rifting episode north of the caldera. Observations using the international constellation of radar satellites indicate rapid 50 cm d –1 subsidence of the glacier surface overlying the collapsing caldera and metre-scale crustal deformation in the active rift zone. Anomalous earthquakes around the rim of the caldera with highly nondouble-couple focal mechanisms provide a mechanical link to the dynamics of the collapsing magma chamber. A model of the collapse consistent with available geodetic and seismic observations suggests that the majority of the observed subsidence occurs aseismically via a deflating sill-like magma chamber.

Description: MicroRNAs (miRNAs), small non-coding RNA molecules, are important biomarkers for research and medical purposes. Here, we describe the development of a fast and simple method using highly fluorescent oligonucleotide-silver nanocluster probes (DNA/AgNCs) to efficiently detect specific miRNAs. Due to the great sequence diversity of miRNAs in humans and other organisms, a uniform strategy for miRNA detection is attractive. The concept presented is an oligonucleotide-based locking-to-unlocking system that can be endowed with miRNA complementarity while maintaining the same secondary structure. The locking-to-unlocking system is based on fold-back anchored DNA templates that consist of a cytosine-rich loop for AgNCs stabilization, an miRNA recognition site and an overlap region for hairpin stabilization. When an miRNA is recognized, fluorescence in the visible region is specifically extinguished in a concentration-dependent manner. Here, the exact composition of the fold-back anchor for the locking-to-unlocking system has been systematically optimized, balancing propensity for loop-structure formation, encapsulation of emissive AgNCs and target sensitivity. It is demonstrated that the applied strategy successfully can detect a number of cancer related miRNAs in RNA extracts from human cancer cell lines.

Description: Studies of discrete volcanic explosions, that usually last less than 2 or 3 min, have suggested that the partitioning of seismic-acoustic energy is likely related to a range of physical mechanisms that depend on magma properties and other physical constraints such as the location of the fragmentation surface. In this paper, we explore the energy partition of a paroxysmal eruptive phase of Tungurahua volcano that lasted for over 4 hr, on 2006 July 14–15, using seismic-acoustic information recorded by stations on its flanks (near field). We find evidence of a linear scaling between seismic and acoustic energies, with time-dependent intensities, during the sustained explosive phase of the eruption. Furthermore, we argue that this scaling can be explained by two different processes: (1) the fragmentation region ultimately acts as the common source of energy producing both direct seismic waves, that travel through the volcanic edifice, and direct acoustic waves coming from a disturbed atmosphere above the summit; (2) the coupling of acoustic waves with the ground to cause seismic waves. Both processes are concurrent, however we have found that the first one is dominant for seismic records below 4 Hz. Here we use the linear scaling of intensities to construct seismic and acoustic indices, which, we argue, could be used to track an ongoing eruption. Thus, especially in strong paroxysms that can produce pyroclastic flows, the index correlation and their levels can be used as quantitative monitoring parameters to assess the volcanic hazard in real time. Additionally, we suggest from the linear scaling that the source type for both cases, seismic and acoustic, is dipolar and dominant in the near field.

Description: Cap analysis of gene expression (CAGE) is a high-throughput method for transcriptome analysis that provides a single base-pair resolution map of transcription start sites (TSS) and their relative usage. Despite their high resolution and functional significance, published CAGE data are still underused in promoter analysis due to the absence of tools that enable its efficient manipulation and integration with other genome data types. Here we present CAGEr , an R implementation of novel methods for the analysis of differential TSS usage and promoter dynamics, integrated with CAGE data processing and promoterome mining into a first comprehensive CAGE toolbox on a common analysis platform. Crucially, we provide collections of TSSs derived from most published CAGE datasets, as well as direct access to FANTOM5 resource of TSSs for numerous human and mouse cell/tissue types from within R, greatly increasing the accessibility of precise context-specific TSS data for integrative analyses. The CAGEr package is freely available from Bioconductor at http://www.bioconductor.org/packages/release/bioc/html/CAGEr.html .

Description: The variability and complexity of the transcription initiation process was examined by adapting RNA ligase-mediated rapid amplification of 5' cDNA ends (5'-RACE) to Next-Generation Sequencing (NGS). We oligo-labelled 5'-m 7 G-capped mRNA from two genes, the simple mono-exonic Beta-2-Adrenoceptor (ADRB2R) and the complex multi-exonic Glucocorticoid Receptor (GR, NR3C1) , and detected a variability in TSS location that has received little attention up to now. Transcription was not initiated at a fixed TSS, but from loci of 4 to 10 adjacent nucleotides. Individual TSSs had frequencies from 〈0.001% to 38.5% of the total gene-specific 5' m 7 G-capped transcripts. ADRB2R used a single locus consisting of 4 adjacent TSSs. Unstimulated, the GR used a total of 358 TSSs distributed throughout 38 loci, that were principally in the 5' UTRs and were spliced using established donor and acceptor sites. Complete demethylation of the epigenetically sensitive GR promoter with 5-azacytidine induced one new locus and 127 TSSs, 12 of which were unique. We induced GR transcription with dexamethasone and Interferon-, adding one new locus and 185 additional TSSs distributed throughout the promoter region. In-vitro the TSS microvariability regulated mRNA translation efficiency and the relative abundance of the different GR N-terminal protein isoform levels.

Description: One of the unresolved questions concerning fault deformation is the degree and cause of localization of shear at depth beneath a fault. Geologic observations of exhumed shear zones indicate that while the motion is no longer planar, it can still be localized near the down-dip extension of the fault; however, the degree of localization is uncertain. We employ simple analytic and numerical models to investigate the structural form of distributed shear beneath a strike-slip fault, and the relative importance of the physical mechanisms that have the potential to localize a shear zone. For a purely depth dependent viscosity, = 0 exp (– z / z 0 ), we find that a shear zone develops with a half-width $\delta _w\sim \sqrt{z_0}$ for small z 0 at the base of the layer, where lengths are non-dimensionalized by the layer thickness ( d km). Including a non-linear stress–strain-rate relation ( $\dot{\epsilon }\propto \sigma ^n$ ) scales w by $1/\sqrt{n}$ , comparable to deformation length scales in thin viscous sheet calculations. We find that the primary control on the shear-zone width is the depth dependence of viscosity that arises from the temperature dependence of viscosity and the increase in temperature with depth. As this relationship is exponential, scaling relations give a dimensional half-width that scales approximately as $$ \tilde{\delta }_w\approx T_{\frac{1}{2}}\sqrt{\frac{Rd}{nQ\beta }}\text{ km}, $$ where $T_{\frac{1}{2}}$ (K) is the temperature at the midpoint of the layer, R (J mol –1 K –1 ) the gas constant, Q (J mol –1 ) the activation energy and β (K km –1 ) the geothermal gradient. This relation predicts the numerical results for the parameter range consistent with continental rheologies to within 2–5 per cent and shear-zone half-widths from 2 to 6 km. The inclusion of shear-stress heating reduces w by only an additional 5–25 per cent, depending on the initial width of the shear zone. While the width of the shear zone may not decrease significantly, local temperature increases from shear-stress heating range from 50 to 300 °C resulting in a reduction in viscosities beneath the fault of several orders of magnitude and a concomitant reduction in the stresses needed to drive the motion.

Description: As the use of RNA-seq has popularized, there is an increasing consciousness of the importance of experimental design, bias removal, accurate quantification and control of false positives for proper data analysis. We introduce the NOISeq R-package for quality control and analysis of count data. We show how the available diagnostic tools can be used to monitor quality issues, make pre-processing decisions and improve analysis. We demonstrate that the non-parametric NOISeqBIO efficiently controls false discoveries in experiments with biological replication and outperforms state-of-the-art methods. NOISeq is a comprehensive resource that meets current needs for robust data-aware analysis of RNA-seq differential expression.

Description: Methods that permit controlled changes in the expression of genes are important tools for biological and medical research, and for biotechnological applications. Conventional methods are directed at individually changing each gene, its regulatory elements or its mRNA's translation rate. We demonstrate that the CRISPR-associated DNA-binding Cascade complex can be used for efficient, long-lasting and programmable gene silencing. When Cascade is targeted to a promoter sequence the transcription of the downstream gene is inhibited, resulting in dramatically reduced expression. The specificity of Cascade binding is provided by the integral crRNA component, which is easily designed to target virtually any stretch of DNA. Cascade targeted to the ORF sequence of the gene can also silence expression, albeit at lower efficiency. The system can be used to silence plasmid and chromosome targets, simultaneously target several genes and is active in different bacterial species and strains. The findings described here are an addition to the expanding range of CRISPR-based technologies and may be adapted to additional organisms and cell systems.

Description: We present a study of the seismic velocity variations that occurred in the structure before the large 2010 eruption of Merapi volcano. For the first time to our knowledge, the technique of coda wave interferometry is applied to both families of similar events (multiplets) and to correlation functions of seismic noise. About half of the seismic events recorded at the summit stations belong to one of the ten multiplets identified, including 120 similar events that occurred in the last 20 hr preceding the eruption onset. Daily noise cross-correlation functions (NCF) were calculated for the six pairs of short-period stations available. Using the stretching method, we estimate time-series of apparent velocity variation (AVV) for each multiplet and each pair of stations. No significant velocity change is detected until September 2010. From 10 October to the beginning of the eruption on 26 October, a complex pattern of AVV is observed with amplitude of up to ±1.5 per cent. Velocity decrease is first observed from families of deep events and then from shallow earthquakes. In the same period, AVV with different signs and chronologies are estimated from NCF calculated for various station pairs. The location in the horizontal plane of the velocity perturbations related with the AVV obtained from NCF is estimated by using an approach based on the radiative transfer approximation. Although their spatial resolution is limited, the resulting maps display velocity decrease in the upper part of the edifice in the period 12–25 October. After the eruption onset, the pattern of velocity perturbations is significantly modified with respect to the previous one. We interpret these velocity variations in the framework of a scenario of magmatic intrusion that integrates most observations. The perturbation of the stress field associated with the magma migration can induce both decrease and increase of the seismic velocity of rocks. Thus the detected AVVs can be considered as precursors of volcanic eruptions in andesitic volcanoes, without taking their sign into account.