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Bryl, R.

Publications and source records attributed to Bryl, R..

2 recordsLinked to original sources

g.nome, A Transparent Bioinformatics Pipeline that Enables Differential Expression and Alternative Splicing Analysis by Non-Computational Biologists

Reproducibility and accessibility are cardinal principles in the rapidly evolving field of bioinformatics. As the collection of biological data grows, proper use of pipelines to analyze datasets can become a bottleneck restricting efficient analysis. Biologists who collect data and test hypotheses may not have strong computational backgrounds and may not be able to fully understand the underlying strengths and weaknesses of computational approaches or fully exploit their data. Some data may be misunderstood and, perhaps more importantly, critical findings may remain unobserved. High throughput RNA sequencing (RNAseq) has advanced our understanding of transcriptomics across diverse applications. Here we introduce g.nome, a bioinformatics platform that integrates contemporary tools necessary for independent analysis. A user-friendly graphical interface simplifies running jobs and allows simplified analysis of different datasets by non-bioinformaticians. g.nome was used to analyze the consequences of localizing the critical RNAi factor argonaute (AGO) to nuclei of colorectal cancer cell line HCT116. Analysis using the pipeline facilitated the straightforward identification of splicing changes and the prioritization of these splicing changes for validation and further experimental analysis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=114 SRC="FIGDIR/small/652286v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@f0fa7org.highwire.dtl.DTLVardef@ccac5borg.highwire.dtl.DTLVardef@147c331org.highwire.dtl.DTLVardef@5fe66c_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

Nuclear Argonaute:miRNA complexes recognize target sequences within chromatin and silence gene expression

RNA interference (RNAi) in mammalian cells involves recognition of mRNA in the cytoplasm and inhibition of translation. Both protein RNAi factors and miRNAs, however, are present in mammalian cell nuclei. It is unclear how this nuclear localization affects endogenous gene expression. Here, we use chimeric eCLIP to identify complexes of Argonaute 2 (AGO2) and miRNAs. We identify the most abundant miRNAs associated with chromatin and their chromatin-associated RNA targets. Chimeric eCLIP revealed that High mobility group AT-Hook A (HMGA2) was the most compelling target for miRNA-mediated gene binding. There are four confirmed let-7 miRNA sites within the 3-UTR in the cytoplasm or nucleus and three within chromatin-associated RNA. The expression of mature HMGA2 mRNA was repressed by let-7 in the cytoplasm and nucleus. let-7 had little effect splicing or transcription. Our data validate chimeric eCLIP as a powerful method for experimentally identifying promising miRNA:RNA interactions. Rather than a solely cytoplasmic event, binding of RNAi factors to mRNA targets may begin in the nucleus through a mechanism that can reduce RNA levels in both the cytoplasm and the nucleus. miRNA-mediated silencing of mRNAs may be influenced by both nuclear and cytoplasmic interactions.

biochemistry↗