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Open accessJournal ArticleDOI: 10.1038/S41467-021-21663-W

The methyltransferase SETD2 couples transcription and splicing by engaging mRNA processing factors through its SHI domain.

04 Mar 2021-Nature Communications (Springer Science and Business Media LLC)-Vol. 12, Iss: 1, pp 1443-1443
Abstract: Heterogeneous ribonucleoproteins (hnRNPs) are RNA binding molecules that are involved in key processes such as RNA splicing and transcription. One such hnRNP protein, hnRNP L, regulates alternative splicing (AS) by binding to pre-mRNA transcripts. However, it is unclear what factors contribute to hnRNP L-regulated AS events. Using proteomic approaches, we identified several key factors that co-purify with hnRNP L. We demonstrate that one such factor, the histone methyltransferase SETD2, specifically interacts with hnRNP L in vitro and in vivo. This interaction occurs through a previously uncharacterized domain in SETD2, the SETD2-hnRNP Interaction (SHI) domain, the deletion of which, leads to a reduced H3K36me3 deposition. Functionally, SETD2 regulates a subset of hnRNP L-targeted AS events. Our findings demonstrate that SETD2, by interacting with Pol II as well as hnRNP L, can mediate the crosstalk between the transcription and the splicing machinery.

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Topics: Ribonucleoprotein (62%), RNA splicing (58%), Alternative splicing (56%) ... read more
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8 results found


Open accessJournal ArticleDOI: 10.1126/SCIADV.ABG7444
Jia Ray Yu1, Jia Ray Yu2, Gary LeRoy2, Gary LeRoy1  +14 moreInstitutions (5)
14 Jul 2021-Science Advances
Abstract: Histone H3K27M is a driving mutation in diffuse intrinsic pontine glioma (DIPG), a deadly pediatric brain tumor. H3K27M reshapes the epigenome through a global inhibition of PRC2 catalytic activity and displacement of H3K27me2/3, promoting oncogenesis of DIPG. As a consequence, a histone modification H3K36me2, antagonistic to H3K27me2/3, is aberrantly elevated. Here, we investigate the role of H3K36me2 in H3K27M-DIPG by tackling its upstream catalyzing enzymes (writers) and downstream binding factors (readers). We determine that NSD1 and NSD2 are the key writers for H3K36me2. Loss of NSD1/2 in H3K27M-DIPG impedes cellular proliferation and tumorigenesis by disrupting tumor-promoting transcriptional programs. Further, we demonstrate that LEDGF and HDGF2 are the main readers mediating the protumorigenic effects downstream of NSD1/2-H3K36me2. Treatment with a chemically modified peptide mimicking endogenous H3K36me2 dislodges LEDGF/HDGF2 from chromatin and specifically inhibits the proliferation of H3K27M-DIPG. Our results indicate a functional pathway of NSD1/2-H3K36me2-LEDGF/HDGF2 as an acquired dependency in H3K27M-DIPG.

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Topics: Epigenome (52%), PRC2 (51%)

2 Citations


Open accessJournal ArticleDOI: 10.1016/J.JBC.2021.101075
Abstract: SETD2 is an important methyltransferase that methylates crucial substrates such as histone H3, tubulin, and STAT1 and also physically interacts with transcription and splicing regulators such as Pol II and various hnRNPs. Of note, SETD2 has a functionally uncharacterized extended N-terminal region, the removal of which leads to its stabilization. How this region regulates SETD2 half-life is unclear. Here we show that SETD2 consists of multiple long disordered regions across its length that cumulatively destabilize the protein by facilitating its proteasomal degradation. SETD2 disordered regions can reduce the half-life of the yeast homolog Set2 in mammalian cells as well as in yeast, demonstrating the importance of intrinsic structural features in regulating protein half-life. In addition to the shortened half-life, by performing fluorescence recovery after photobleaching assay we found that SETD2 forms liquid droplets in vivo, another property associated with proteins that contain disordered regions. The phase-separation behavior of SETD2 is exacerbated upon the removal of its N-terminal segment and results in activator-independent histone H3K36 methylation. Our findings reveal that disordered region-facilitated proteolysis is an important mechanism governing SETD2 function.

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Topics: Protein degradation (58%), Histone H3 (53%), Fluorescence recovery after photobleaching (52%) ... read more

1 Citations


Open accessJournal ArticleDOI: 10.1016/J.NEURON.2021.08.008
15 Sep 2021-Neuron
Abstract: Neuronal alternative splicing is a key gene regulatory mechanism in the brain. However, the spliceosome machinery is insufficient to fully specify splicing complexity. In considering the role of the epigenome in activity-dependent alternative splicing, we and others find the histone modification H3K36me3 to be a putative splicing regulator. In this study, we found that mouse cocaine self-administration caused widespread differential alternative splicing, concomitant with the enrichment of H3K36me3 at differentially spliced junctions. Importantly, only targeted epigenetic editing can distinguish between a direct role of H3K36me3 in splicing and an indirect role via regulation of splice factor expression elsewhere on the genome. We targeted Srsf11, which was both alternatively spliced and H3K36me3 enriched in the brain following cocaine self-administration. Epigenetic editing of H3K36me3 at Srsf11 was sufficient to drive its alternative splicing and enhanced cocaine self-administration, establishing the direct causal relevance of H3K36me3 to alternative splicing of Srsf11 and to reward behavior.

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Topics: Alternative splicing (68%), RNA splicing (63%), Spliceosome (58%) ... read more

1 Citations


Open accessJournal ArticleDOI: 10.3389/FCELL.2021.739780
Kenya Bonitto1, Kirthana Sarathy1, Kaiser Atai1, Mithun Mitra1  +1 moreInstitutions (1)
Abstract: Many of the cells in our bodies are quiescent, that is, temporarily not dividing. Under certain physiological conditions such as during tissue repair and maintenance, quiescent cells receive the appropriate stimulus and are induced to enter the cell cycle. The ability of cells to successfully transition into and out of a quiescent state is crucial for many biological processes including wound healing, stem cell maintenance, and immunological responses. Across species and tissues, transcriptional, epigenetic, and chromosomal changes associated with the transition between proliferation and quiescence have been analyzed, and some consistent changes associated with quiescence have been identified. Histone modifications have been shown to play a role in chromatin packing and accessibility, nucleosome mobility, gene expression, and chromosome arrangement. In this review, we critically evaluate the role of different histone marks in these processes during quiescence entry and exit. We consider different model systems for quiescence, each of the most frequently monitored candidate histone marks, and the role of their writers, erasers and readers. We highlight data that support these marks contributing to the changes observed with quiescence. We specifically ask whether there is a quiescence histone "code," a mechanism whereby the language encoded by specific combinations of histone marks is read and relayed downstream to modulate cell state and function. We conclude by highlighting emerging technologies that can be applied to gain greater insight into the role of a histone code for quiescence.

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Topics: Histone code (65%), Histone methylation (57%), Nucleosome (54%) ... read more

Open accessPosted ContentDOI: 10.1101/2021.05.22.445248
Saikat Bhattacharya1, Suman Wang1, Divya Reddy2, Siyuan Shen1  +8 moreInstitutions (3)
23 May 2021-bioRxiv
Abstract: The RNA recognition motif (RRM) binds to nucleic acids as well as proteins. More than one such domain is found in the pre-mRNA processing hnRNP proteins. While the mode of RNA recognition by RRMs is known, the molecular basis of their protein interaction remains obscure. Here we describe the mode of interaction between hnRNP L and LL with the methyltransferase SETD2. We demonstrate that for the interaction to occur, a leucine pair within a highly conserved stretch of SETD2 insert their side chains in hydrophobic pockets formed by hnRNP L RRM2. Notably, the structure also highlights that RRM2 can form a ternary complex with SETD2 and RNA. Remarkably, mutating the leucine pair in SETD2 also results in its reduced interaction with other hnRNPs. Importantly, the similarity that the mode of SETD2-hnRNP L interaction shares with other related protein-protein interactions reveals a conserved design by which splicing regulators interact with one another.

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Topics: RNA splicing (55%), RNA recognition motif (55%), RNA (53%)

References
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65 results found


Open accessJournal ArticleDOI: 10.1038/NMETH.1923
01 Apr 2012-Nature Methods
Abstract: As the rate of sequencing increases, greater throughput is demanded from read aligners. The full-text minute index is often used to make alignment very fast and memory-efficient, but the approach is ill-suited to finding longer, gapped alignments. Bowtie 2 combines the strengths of the full-text minute index with the flexibility and speed of hardware-accelerated dynamic programming algorithms to achieve a combination of high speed, sensitivity and accuracy.

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27,973 Citations


Open accessJournal ArticleDOI: 10.1093/BIOINFORMATICS/BTP616
01 Jan 2010-Bioinformatics
Abstract: Summary: It is expected that emerging digital gene expression (DGE) technologies will overtake microarray technologies in the near future for many functional genomics applications. One of the fundamental data analysis tasks, especially for gene expression studies, involves determining whether there is evidence that counts for a transcript or exon are significantly different across experimental conditions. edgeR is a Bioconductor software package for examining differential expression of replicated count data. An overdispersed Poisson model is used to account for both biological and technical variability. Empirical Bayes methods are used to moderate the degree of overdispersion across transcripts, improving the reliability of inference. The methodology can be used even with the most minimal levels of replication, provided at least one phenotype or experimental condition is replicated. The software may have other applications beyond sequencing data, such as proteome peptide count data. Availability: The package is freely available under the LGPL licence from the Bioconductor web site (http://bioconductor.org).

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Topics: Bioconductor (64%)

21,575 Citations


Open accessJournal ArticleDOI: 10.1093/BIOINFORMATICS/BTS635
01 Jan 2013-Bioinformatics
Abstract: Motivation Accurate alignment of high-throughput RNA-seq data is a challenging and yet unsolved problem because of the non-contiguous transcript structure, relatively short read lengths and constantly increasing throughput of the sequencing technologies. Currently available RNA-seq aligners suffer from high mapping error rates, low mapping speed, read length limitation and mapping biases. Results To align our large (>80 billon reads) ENCODE Transcriptome RNA-seq dataset, we developed the Spliced Transcripts Alignment to a Reference (STAR) software based on a previously undescribed RNA-seq alignment algorithm that uses sequential maximum mappable seed search in uncompressed suffix arrays followed by seed clustering and stitching procedure. STAR outperforms other aligners by a factor of >50 in mapping speed, aligning to the human genome 550 million 2 × 76 bp paired-end reads per hour on a modest 12-core server, while at the same time improving alignment sensitivity and precision. In addition to unbiased de novo detection of canonical junctions, STAR can discover non-canonical splices and chimeric (fusion) transcripts, and is also capable of mapping full-length RNA sequences. Using Roche 454 sequencing of reverse transcription polymerase chain reaction amplicons, we experimentally validated 1960 novel intergenic splice junctions with an 80-90% success rate, corroborating the high precision of the STAR mapping strategy. Availability and implementation STAR is implemented as a standalone C++ code. STAR is free open source software distributed under GPLv3 license and can be downloaded from http://code.google.com/p/rna-star/.

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Topics: MRNA Sequencing (57%)

20,172 Citations


Journal ArticleDOI: 10.1038/85686
Abstract: We describe a largely unbiased method for rapid and large-scale proteome analysis by multidimensional liquid chromatography, tandem mass spectrometry, and database searching by the SEQUEST algorithm, named multidimensional protein identification technology (MudPIT). MudPIT was applied to the proteome of the Saccharomyces cerevisiae strain BJ5460 grown to mid-log phase and yielded the largest proteome analysis to date. A total of 1,484 proteins were detected and identified. Categorization of these hits demonstrated the ability of this technology to detect and identify proteins rarely seen in proteome analysis, including low-abundance proteins like transcription factors and protein kinases. Furthermore, we identified 131 proteins with three or more predicted transmembrane domains, which allowed us to map the soluble domains of many of the integral membrane proteins. MudPIT is useful for proteome analysis and may be specifically applied to integral membrane proteins to obtain detailed biochemical information on this unwieldy class of proteins.

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Topics: Proteome (65%), Proteomics (55%), Integral membrane protein (55%) ... read more

4,699 Citations


Open accessJournal ArticleDOI: 10.1038/NATURE07509
Eric T. Wang1, Rickard Sandberg2, Rickard Sandberg1, Shujun Luo3  +6 moreInstitutions (4)
27 Nov 2008-Nature
Abstract: Through alternative processing of pre-messenger RNAs, individual mammalian genes often produce multiple mRNA and protein isoforms that may have related, distinct or even opposing functions. Here we report an in-depth analysis of 15 diverse human tissue and cell line transcriptomes on the basis of deep sequencing of complementary DNA fragments, yielding a digital inventory of gene and mRNA isoform expression. Analyses in which sequence reads are mapped to exon-exon junctions indicated that 92-94% of human genes undergo alternative splicing, 86% with a minor isoform frequency of 15% or more. Differences in isoform-specific read densities indicated that most alternative splicing and alternative cleavage and polyadenylation events vary between tissues, whereas variation between individuals was approximately twofold to threefold less common. Extreme or 'switch-like' regulation of splicing between tissues was associated with increased sequence conservation in regulatory regions and with generation of full-length open reading frames. Patterns of alternative splicing and alternative cleavage and polyadenylation were strongly correlated across tissues, suggesting coordinated regulation of these processes, and sequence conservation of a subset of known regulatory motifs in both alternative introns and 3' untranslated regions suggested common involvement of specific factors in tissue-level regulation of both splicing and polyadenylation.

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4,242 Citations