Browsing by Subject "RNA-binding proteins"

Now showing 1 - 5 of 5
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    Comparative Analysis of Alternative Splicing Profiles in Th Cell Subsets Reveals Extensive Cell Type–Specific Effects Modulated by a Network of Transcription Factors and RNA-Binding Proteins
    (American Association of Immunologists, 2021-09-28) Mir, Quoseena; Lakshmipati, Deepak K.; Ulrich, Benjamin J.; Kaplan, Mark H.; Janga, Sarath Chandra; Biomedical Engineering and Informatics, Luddy School of Informatics, Computing, and Engineering
    Alternative splicing (AS) plays an important role in the development of many cell types; however, its contribution to Th subsets has been clearly defined. In this study, we compare mice naive CD4+ Th cells with Th1, Th2, Th17, and T regulatory cells and observed that the majority of AS events were retained intron, followed by skipped-exon events, with at least 1200 genes across cell types affected by AS events. A significant fraction of the AS events, especially retained intron events from the 72-h time point, were no longer observed 2 wk postdifferentiation, suggesting a role for AS in early activation and differentiation via preferential expression of specific isoforms required during T cell activation, but not for differentiation or effector function. Examining the protein consequence of the exon-skipping events revealed an abundance of structural proteins encoding for intrinsically unstructured peptide regions, followed by transmembrane helices, β strands, and polypeptide turn motifs. Analyses of expression profiles of RNA-binding proteins (RBPs) and their cognate binding sites flanking the discovered AS events revealed an enrichment for specific RBP recognition sites in each of the Th subsets. Integration with publicly available chromatin immunoprecipitation sequencing datasets for transcription factors support a model wherein lineage-determining transcription factors impact the RBP profile within the differentiating cells, and this differential expression contributes to AS of the transcriptome via a cascade of cell type-specific posttranscriptional rewiring events.
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    Dissecting the expression landscape of RNA-binding proteins in human cancers
    (2014-01) Kechavarzi, Bobak; Janga, Sarath Chandra
    Background RNA-binding proteins (RBPs) play important roles in cellular homeostasis by controlling gene expression at the post-transcriptional level. Results We explore the expression of more than 800 RBPs in sixteen healthy human tissues and their patterns of dysregulation in cancer genomes from The Cancer Genome Atlas project. We show that genes encoding RBPs are consistently and significantly highly expressed compared with other classes of genes, including those encoding regulatory components such as transcription factors, miRNAs and long non-coding RNAs. We also demonstrate that a set of RBPs, numbering approximately 30, are strongly upregulated (SUR) across at least two-thirds of the nine cancers profiled in this study. Analysis of the protein–protein interaction network properties for the SUR and non-SUR groups of RBPs suggests that path length distributions between SUR RBPs is significantly lower than those observed for non-SUR RBPs. We further find that the mean path lengths between SUR RBPs increases in proportion to their contribution to prognostic impact. We also note that RBPs exhibiting higher variability in the extent of dysregulation across breast cancer patients have a higher number of protein–protein interactions. We propose that fluctuating RBP levels might result in an increase in non-specific protein interactions, potentially leading to changes in the functional consequences of RBP binding. Finally, we show that the expression variation of a gene within a patient group is inversely correlated with prognostic impact. Conclusions Overall, our results provide a roadmap for understanding the impact of RBPs on cancer pathogenesis.
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    Long Non-Coding RNA Expression Levels Modulate Cell-Type-Specific Splicing Patterns by Altering Their Interaction Landscape with RNA-Binding Proteins
    (MDPI, 2019-08-06) Porto, Felipe Wendt; Daulatabad, Swapna Vidhur; Janga, Sarath Chandra; BioHealth Informatics, School of Informatics and Computing
    Recent developments in our understanding of the interactions between long non-coding RNAs (lncRNAs) and cellular components have improved treatment approaches for various human diseases including cancer, vascular diseases, and neurological diseases. Although investigation of specific lncRNAs revealed their role in the metabolism of cellular RNA, our understanding of their contribution to post-transcriptional regulation is relatively limited. In this study, we explore the role of lncRNAs in modulating alternative splicing and their impact on downstream protein-RNA interaction networks. Analysis of alternative splicing events across 39 lncRNA knockdown and wildtype RNA-sequencing datasets from three human cell lines-HeLa (cervical cancer), K562 (myeloid leukemia), and U87 (glioblastoma)-resulted in the high-confidence (false discovery rate (fdr) < 0.01) identification of 11,630 skipped exon events and 5895 retained intron events, implicating 759 genes to be impacted at the post-transcriptional level due to the loss of lncRNAs. We observed that a majority of the alternatively spliced genes in a lncRNA knockdown were specific to the cell type. In tandem, the functions annotated to the genes affected by alternative splicing across each lncRNA knockdown also displayed cell-type specificity. To understand the mechanism behind this cell-type-specific alternative splicing pattern, we analyzed RNA-binding protein (RBP)-RNA interaction profiles across the spliced regions in order to observe cell-type-specific alternative splice event RBP binding preference. Despite limited RBP binding data across cell lines, alternatively spliced events detected in lncRNA perturbation experiments were associated with RBPs binding in proximal intron-exon junctions in a cell-type-specific manner. The cellular functions affected by alternative splicing were also affected in a cell-type-specific manner. Based on the RBP binding profiles in HeLa and K562 cells, we hypothesize that several lncRNAs are likely to exhibit a sponge effect in disease contexts, resulting in the functional disruption of RBPs and their downstream functions. We propose that such lncRNA sponges can extensively rewire post-transcriptional gene regulatory networks by altering the protein-RNA interaction landscape in a cell-type-specific manner.
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    MS0621, a novel small-molecule modulator of Ewing sarcoma chromatin accessibility, interacts with an RNA-associated macromolecular complex and influences RNA splicing
    (Frontiers Media, 2023-01-30) Vital, Tamara; Wali, Aminah; Butler, Kyle V.; Xiong, Yan; Foster, Joseph P., II; Marcel, Shelsa S.; McFadden, Andrew W.; Nguyen, Valerie U.; Bailey, Benton M.; Lamb, Kelsey N.; James, Lindsey I.; Frye, Stephen V.; Mosely, Amber L.; Jin, Jian; Pattenden, Samantha G.; Davis, Ian J.; Biochemistry and Molecular Biology, School of Medicine
    Ewing sarcoma is a cancer of children and young adults characterized by the critical translocation-associated fusion oncoprotein EWSR1::FLI1. EWSR1::FLI1 targets characteristic genetic loci where it mediates aberrant chromatin and the establishment of de novo enhancers. Ewing sarcoma thus provides a model to interrogate mechanisms underlying chromatin dysregulation in tumorigenesis. Previously, we developed a high-throughput chromatin-based screening platform based on the de novo enhancers and demonstrated its utility in identifying small molecules capable of altering chromatin accessibility. Here, we report the identification of MS0621, a molecule with previously uncharacterized mechanism of action, as a small molecule modulator of chromatin state at sites of aberrant chromatin accessibility at EWSR1::FLI1-bound loci. MS0621 suppresses cellular proliferation of Ewing sarcoma cell lines by cell cycle arrest. Proteomic studies demonstrate that MS0621 associates with EWSR1::FLI1, RNA binding and splicing proteins, as well as chromatin regulatory proteins. Surprisingly, interactions with chromatin and many RNA-binding proteins, including EWSR1::FLI1 and its known interactors, were RNA-independent. Our findings suggest that MS0621 affects EWSR1::FLI1-mediated chromatin activity by interacting with and altering the activity of RNA splicing machinery and chromatin modulating factors. Genetic modulation of these proteins similarly inhibits proliferation and alters chromatin in Ewing sarcoma cells. The use of an oncogene-associated chromatin signature as a target allows for a direct approach to screen for unrecognized modulators of epigenetic machinery and provides a framework for using chromatin-based assays for future therapeutic discovery efforts.
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    Seten: a tool for systematic identification and comparison of processes, phenotypes, and diseases associated with RNA-binding proteins from condition-specific CLIP-seq profiles
    (Cold Spring Harbor Laboratory Press, 2017-06) Budak, Gungor; Srivastava, Rajneesh; Janga, Sarath Chandra; BioHealth Informatics, School of Informatics and Computing
    RNA-binding proteins (RBPs) control the regulation of gene expression in eukaryotic genomes at post-transcriptional level by binding to their cognate RNAs. Although several variants of CLIP (crosslinking and immunoprecipitation) protocols are currently available to study the global protein-RNA interaction landscape at single-nucleotide resolution in a cell, currently there are very few tools that can facilitate understanding and dissecting the functional associations of RBPs from the resulting binding maps. Here, we present Seten, a web-based and command line tool, which can identify and compare processes, phenotypes, and diseases associated with RBPs from condition-specific CLIP-seq profiles. Seten uses BED files resulting from most peak calling algorithms, which include scores reflecting the extent of binding of an RBP on the target transcript, to provide both traditional functional enrichment as well as gene set enrichment results for a number of gene set collections including BioCarta, KEGG, Reactome, Gene Ontology (GO), Human Phenotype Ontology (HPO), and MalaCards Disease Ontology for several organisms including fruit fly, human, mouse, rat, worm, and yeast. It also provides an option to dynamically compare the associated gene sets across data sets as bubble charts, to facilitate comparative analysis. Benchmarking of Seten using eCLIP data for IGF2BP1, SRSF7, and PTBP1 against their corresponding CRISPR RNA-seq in K562 cells as well as randomized negative controls, demonstrated that its gene set enrichment method outperforms functional enrichment, with scores significantly contributing to the discovery of true annotations. Comparative performance analysis using these CRISPR control data sets revealed significantly higher precision and comparable recall to that observed using ChIP-Enrich. Seten's web interface currently provides precomputed results for about 200 CLIP-seq data sets and both command line as well as web interfaces can be used to analyze CLIP-seq data sets. We highlight several examples to show the utility of Seten for rapid profiling of various CLIP-seq data sets.