Splicing modulators elicit global translational repression by condensate-prone proteins translated from introns
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PROVIDER: PRJNA530968 | ENA |
REPOSITORIES: ENA
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Project description:Chemical splicing modulators that bind to the spliceosome have provided an attractive venue for cancer treatment. Splicing modulators induce accumulation and subsequent translation of a subset of intron-retained mRNAs. Yet, the biological effect of proteins containing translated intron sequences remains unclear. Here we identified a number of truncated proteins generated upon treatment with the splicing modulator spliceostatin A (SSA) using genome-wide ribosome profiling and bio-orthogonal non-canonical amino-acid tagging (BONCAT) mass spectrometry. A subset of these truncated proteins has intrinsically disordered regions, forms insoluble cellular condensates, and triggers the proteotoxic stress response through JNK phosphorylation, thereby inhibiting the mTORC1 pathway. In turn, this reduces global translation. These findings indicate that creating an overburden of condensate-prone proteins derived from introns represses translation and prevents further production of harmful truncated proteins. This mechanism appears to contribute to the antiproliferative and proapoptotic activity of splicing modulators.
2021-03-29 | GSE129305 | GEO
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2009-08-09 | E-GEOD-14461 | biostudies-arrayexpress
Project description:Removal of introns by pre-mRNA splicing is a critical and in some cases rate-limiting step in mammalian gene expression. Deep sequencing of mouse embryonic stem cell RNA revealed many specific internal introns that are significantly more abundant than the other introns within poly(A) selected transcripts; we classify these as “detained” introns (DIs). We identified thousands of DIs flanking both constitutive and alternatively spliced exons in human and mouse cell lines. Drug inhibition of Clk SR-protein kinase activity triggered rapid splicing changes in a specific set of DIs, about half of which showed increased splicing and half increased intron detention, altering the transcript pool of over 300 genes. These data suggest a widespread mechanism by which a nuclear detained pool of mostly processed pre-mRNAs can be rapidly mobilized in response to stress or homeostatic autoregulation. v6.5 mouse embryonic stem cells were untreated, treated with the Clk kinase inhibitor KH-CB19, or treated with DMSO as a negative control. Untreated cells were harvested and a single replicate was sequenced using a custom, ligation-based, stranded library preparation protocol. Treated cells were harvested at time 0 and at 2 hours post-treatment, and poly(A)-selected RNA-seq libraries were made from biological duplicates for each treatment/time, barcoded, and sequenced by strand-specific, paired-end sequencing using the Illumina TruSeq kit.
2015-01-15 | E-GEOD-57231 | biostudies-arrayexpress
Project description:Intron retention (IR) is an alternative splicing event where mRNAs containing unspliced introns are exported to the cytoplasm and then either translated, giving rise to new protein isoforms, or degraded via the nonsense-mediated decay pathway. However, unspliced introns are also seen in nuclear RNA. Some of these are spliced at a low rate but do eventually become fully spliced and their respective mRNAs exported, while others stay retained and are recognized by nuclear decay machineries. IR has been shown to be regulated during granulocyte and neuronal differentiation and in some cancers, and a subset of the nuclear retained introns, called detained introns, have been implicated in growth control pathways. Despite many findings on IR and its biological significance, it is still not clear whether an incompletely spliced intron observed is a true retained intron that is exported as an mRNA or is a product of slow splicing that remains in the nucleus. A better understanding is needed of the molecular events that reduce intron excision rate and determine the release of an RNA for export. We have found that some genes in mouse embryonic stem cells (mESCs) produce RNAs that are highly associated with the high molecular-weight nuclear pellet (chromatin), rather than the soluble nucleoplasm or cytoplasm. This chromatin-associated RNA is polyadenylated, usually contains one or more incompletely spliced introns, and is often extensively bound by polypyrimidine tract-binding protein 1 (PTBP1). We hypothesize that PTBP1 inhibits splicing of these introns, and this causes sequestration of the transcript on chromatin. We are studying the role of PTBP1 in inhibiting intron excision and anchoring RNAs in the nuclear and chromatin compartments of mESCs.
2020-11-18 | GSE159993 | GEO
Project description:Removal of introns by pre-mRNA splicing is a critical and in some cases rate-limiting step in mammalian gene expression. Deep sequencing of mouse embryonic stem cell RNA revealed many specific internal introns that are significantly more abundant than the other introns within poly(A) selected transcripts; we classify these as “detained” introns (DIs). We identified thousands of DIs flanking both constitutive and alternatively spliced exons in human and mouse cell lines. Drug inhibition of Clk SR-protein kinase activity triggered rapid splicing changes in a specific set of DIs, about half of which showed increased splicing and half increased intron detention, altering the transcript pool of over 300 genes. These data suggest a widespread mechanism by which a nuclear detained pool of mostly processed pre-mRNAs can be rapidly mobilized in response to stress or homeostatic autoregulation.
2015-01-15 | GSE57231 | GEO
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2023-10-11 | GSE208345 | GEO
Project description:Spliceosomal introns are ubiquitous non-coding RNAs typically destined for rapid debranching and degradation. Here, we describe 34 excised Saccharomyces cerevisiae introns that, although rapidly degraded in log-phase growth, accumulate as linear RNAs under either saturated-growth conditions or other stresses that cause prolonged inhibition of TORC1, a key integrator of growth signaling. Introns that become stabilized remain associated with components of the spliceosome and differ from other spliceosomal introns in having a short distance between their lariat branch point and 3´ splice site, which is necessary and sufficient for their stabilization. Deletion of these unusual introns is disadvantageous in saturated conditions and causes aberrantly high growth rates of yeast chronically challenged with the TORC1 inhibitor rapamycin. Reintroduction of native or engineered stable introns suppresses this aberrant rapamycin response. Thus, excised introns function within the TOR growth-signaling network of S. cerevisiae, and more generally, excised spliceosomal introns can have biological functions.
2018-10-29 | GSE121765 | GEO
Project description:Local Translation in Nuclear Condensate Amyloid-bodies
| PRJNA692199 | ENA
Project description:Identification of eukaryotic mRNAs that are translated at reduced cap binding complex eIF4F concentrations using a cDNA microarray. Although most eukaryotic mRNAs need a functional cap binding complex eIF4F for efficient 5' end-dependent scanning to initiate translation, picornaviral, hepatitis C viral, and a few cellular RNAs have been shown to be translated by internal ribosome entry, a mechanism that can operate in the presence of low levels of functional eIF4F. To identify cellular mRNAs that can be translated when eIF4F is depleted or in low abundance and that, therefore, may contain internal ribosome entry sites, mRNAs that remained associated with polysomes were isolated from human cells after infection with poliovirus and were identified by using a cDNA microarray. Approximately 200 of the 7000 mRNAs analyzed remained associated with polysomes under these conditions. Among the gene products encoded by these polysome-associated mRNAs were immediate-early transcription factors, kinases, and phosphatases of the mitogen-activated protein kinase pathways and several protooncogenes, including c-myc and Pim-1. In addition, the mRNA encoding Cyr61, a secreted factor that can promote angiogenesis and tumor growth, was selectively mobilized into polysomes when eIF4F concentrations were reduced, although its overall abundance changed only slightly. Subsequent tests confirmed the presence of internal ribosome entry sites in the 5' noncoding regions of both Cyr61 and Pim-1 mRNAs. Overall, this study suggests that diverse mRNAs whose gene products have been implicated in a variety of stress responses, including inflammation, angiogenesis, and the response to serum, can use translational initiation mechanisms that require little or no intact cap binding protein complex eIF4F. This study is described more fully in Johannes G et al.(1999) Proc Natl Acad Sci U S A 96:13118-23.
2002-06-27 | E-GEOD-62 | biostudies-arrayexpress