Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1051052 | ENA |
REPOSITORIES: ENA
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Project description:Transcription and splicing are intricately linked processes, with emerging evidence highlighting the involvement of splicing factors mediating their coupling. U1 small nuclear ribonucleoprotein particle (U1 snRNP), a key splicing factor, not only serves as the initial component of the spliceosome but also plays a crucial role in preventing premature cleavage and polyadenylation, facilitating long-distance transcription elongation. Here, we show that U1 snRNP regulates alternative promoter usage through inhibition of premature polyadenylation. Inhibiting U1 snRNP with antisense oligonucleotides or introducing a premature polyadenylation leads to a significant decrease in downstream promoter activity in genes with premature polyadenylation sites in between two promoters. Conversely, restoring U1 snRNP activity or inhibiting premature polyadenylation sites using a gene-specific U1 snRNP rescues downstream promoter activity. Mechanistically, U1 snRNP inhibition correlates with reduced chromatin accessibility and serine 5 phosphorylation levels of RNA polymerase II at downstream promoters. Our findings propose a model where U1 snRNP facilitates productive transcription from upstream promoters, triggering downstream promoter activation by destabilizing nucleosomes and promoting promoter escape.
2025-05-19 | GSE284682 | GEO
Project description:Transcription and splicing are intricately linked processes, with emerging evidence highlighting the involvement of splicing factors mediating their coupling. U1 small nuclear ribonucleoprotein particle (U1 snRNP), a key splicing factor, not only serves as the initial component of the spliceosome but also plays a crucial role in preventing premature cleavage and polyadenylation, facilitating long-distance transcription elongation. Here, we show that U1 snRNP regulates alternative promoter usage through inhibition of premature polyadenylation. Inhibiting U1 snRNP with antisense oligonucleotides or introducing a premature polyadenylation leads to a significant decrease in downstream promoter activity in genes with premature polyadenylation sites in between two promoters. Conversely, restoring U1 snRNP activity or inhibiting premature polyadenylation sites using a gene-specific U1 snRNP rescues downstream promoter activity. Mechanistically, U1 snRNP inhibition correlates with reduced chromatin accessibility and serine 5 phosphorylation levels of RNA polymerase II at downstream promoters. Our findings propose a model where U1 snRNP facilitates productive transcription from upstream promoters, triggering downstream promoter activation by destabilizing nucleosomes and promoting promoter escape.
2025-05-19 | GSE284680 | GEO
Project description:Transcription and splicing are inherently intertwined. Accumulating evidence support a role of splicing factors in mediating this coupling. U1 small nuclear ribonucleoprotein particle (U1 snRNP), a key splicing factor, acts as the initial building block of the spliceosome, interacting with nascent pre-mRNA at 5' splice sites. In addition to its role in splicing, U1 snRNP is crucial for preventing premature cleavage and polyadenylation, enabling long-distance transcription elongation. Here, we show that U1snRNP can regulate the usage of alternative promoters through its role in inhibiting premature polyadenylation. Using antisense oligonucleotides to inhibit U1 snRNP, we first observed a markedly decrease in downstream promoter activity at the newly synthesized RNA level. Interestingly, U1 snRNP inhibition selectively impacts downstream promoters of genes featuring premature polyadenylation sites located between two promoters. Overexpressing a wild-type U1 snRNP or a gene-specific U1 snRNP designed to inhibit premature polyadenylation sites restores downstream promoter activity. Conversely, introducing a premature polyadenylation site between two promoters reduces downstream promoter activity. Exploring the underlying molecular mechanisms, we identified a substantial decrease in serine 5 phosphorylation levels in newly recruited RNA polymerase II at downstream promoters following U1 snRNP inhibition, and reduced chromatin accessibility in the vicinity of downstream promoters. Overall, our model is consistent with productive transcription from upstream promoters triggering downstream promoter activation in the presence of U1 snRNP by destabilizing nucleosomes and promoting promoter escape at downstream promoters.
2025-05-19 | GSE252391 | GEO
Project description:Transcription and splicing are inherently intertwined. Accumulating evidence support a role of splicing factors in mediating this coupling. U1 small nuclear ribonucleoprotein particle (U1 snRNP), a key splicing factor, acts as the initial building block of the spliceosome, interacting with nascent pre-mRNA at 5' splice sites. In addition to its role in splicing, U1 snRNP is crucial for preventing premature cleavage and polyadenylation, enabling long-distance transcription elongation. Here, we show that U1 snRNP can regulate the usage of alternative promoters through its role in inhibiting premature polyadenylation. Using antisense oligonucleotides to inhibit U1 snRNP, we first observed a markedly decrease in downstream promoter activity at the newly synthesized RNA level. Interestingly, U1 snRNP inhibition selectively impacts downstream promoters of genes featuring premature polyadenylation sites located between two promoters. Overexpressing a wild-type U1 snRNP or a gene-specific U1 snRNP designed to inhibit premature polyadenylation sites restores downstream promoter activity. Conversely, introducing a premature polyadenylation site between two promoters reduces downstream promoter activity. Exploring the underlying molecular mechanisms, we identified a substantial decrease in serine 5 phosphorylation levels in newly recruited RNA polymerase II at downstream promoters following U1 snRNP inhibition, and reduced chromatin accessibility in the vicinity of downstream promoters. Overall, our model is consistent with productive transcription from upstream promoters triggering downstream promoter activation in the presence of U1 snRNP by destabilizing nucleosomes and promoting promoter escape at downstream promoters.
2025-05-19 | GSE252389 | GEO
Project description:Transcription and splicing are inherently intertwined. Accumulating evidence support a role of splicing factors in mediating this coupling. U1 small nuclear ribonucleoprotein particle (U1 snRNP), a key splicing factor, acts as the initial building block of the spliceosome, interacting with nascent pre-mRNA at 5' splice sites. In addition to its role in splicing, U1 snRNP is crucial for preventing premature cleavage and polyadenylation, enabling long-distance transcription elongation. Here, we show that U1 snRNP can regulate the usage of alternative promoters through its role in inhibiting premature polyadenylation. Using antisense oligonucleotides to inhibit U1 snRNP, we first observed a markedly decrease in downstream promoter activity at the newly synthesized RNA level. Interestingly, U1 snRNP inhibition selectively impacts downstream promoters of genes featuring premature polyadenylation sites located between two promoters. Overexpressing a wild-type U1 snRNP or a gene-specific U1 snRNP designed to inhibit premature polyadenylation sites restores downstream promoter activity. Conversely, introducing a premature polyadenylation site between two promoters reduces downstream promoter activity. Exploring the underlying molecular mechanisms, we identified a substantial decrease in serine 5 phosphorylation levels in newly recruited RNA polymerase II at downstream promoters following U1 snRNP inhibition, and reduced chromatin accessibility in the vicinity of downstream promoters. Overall, our model is consistent with productive transcription from upstream promoters triggering downstream promoter activation in the presence of U1 snRNP by destabilizing nucleosomes and promoting promoter escape at downstream promoters.
2025-05-19 | GSE252387 | GEO
Project description:U1 snRNP regulates alternative promoter activity by inhibiting premature polyadenylation
| PRJNA1060322 | ENA
Project description:CDK9 is a critical kinase required for the productive transcription of protein-coding genes by RNA polymerase II (pol II). As part of P-TEFb, CDK9 phosphorylates the carboxyl-terminal domain (CTD) of pol II and elongation factors, including SPT5, which allows pol II to elongate past the early elongation checkpoint (EEC) encountered soon after initiation. We show that, in addition to halting pol II at the EEC, loss of CDK9 activity causes premature termination of transcription across the last exon, loss of polyadenylation factors from chromatin, and loss of polyadenylation of nascent transcripts. Inhibition of the phosphatase PP2A abrogates the premature termination and loss of polyadenylation caused by CDK9 inhibition, indicating that this kinase/phosphatase pair regulates transcription elongation and RNA processing at the end of protein-coding genes. Our phosphoproteomic analyses after CDK9 inhibition, using either DRB or an analog-sensitive CDK9 cell line, confirm the splicing factor SF3B1 as an additional key target of this kinase. These results emphasize the important roles that CDK9 plays in coupling transcription elongation and termination to RNA maturation downstream of the EEC. As part of this project, we characterized the interactome of SF3B1 in HeLa cells in duplicate.
2022-08-22 | PXD033694 | Pride
Project description:In eukaryotes, U1 small nuclear ribonucleoprotein (snRNP) forms spliceosomes in equal stoichiometry with U2, U4, U5 and U6 snRNPs; however, its abundance in human far exceeds that of the other snRNPs. Here we used antisense morpholino oligonucleotide to U1 snRNA to achieve functional U1 snRNP knockdown in HeLa cells, and identified accumulated unspliced pre-mRNAs by genomic tiling microarrays. In addition to inhibiting splicing, U1 snRNP knockdown caused premature cleavage and polyadenylation in numerous pre-mRNAs at cryptic polyadenylation signals, frequently in introns near (<5 kilobases) the start of the transcript. This did not occur when splicing was inhibited with U2 snRNA antisense morpholino oligonucleotide or the U2-snRNP-inactivating drug spliceostatin A unless U1 antisense morpholino oligonucleotide was also included. We further show that U1 snRNA–pre-mRNA base pairing was required to suppress premature cleavage and polyadenylation from nearby cryptic polyadenylation signals located in introns. These findings reveal a critical splicing-independent function for U1 snRNP in protecting the transcriptome, which we propose explains its overabundance.
2010-09-29 | GSE24179 | GEO
Project description:The pervasive nature of RNA polymerase II (Pol II) transcription requires efficient termination. A key player in this process is the cleavage and polyadenylation (CPA) factor PCF11, which directly binds to the Pol II C-terminal domain and dismantles elongating Pol II from DNA in vitro. We demonstrate that PCF11-mediated termination is essential for vertebrate development. A range of genomic analyses, including: mNET-seq, 3’ mRNA-seq, chromatin RNA-seq and ChIP-seq, reveals that PCF11 enhances transcription termination and stimulates early polyadenylation genome-wide. PCF11 binds preferentially between closely spaced genes, where it prevents transcriptional interference and downstream gene silencing. Notably, PCF11 is sub-stoichiometric to the CPA complex. Low levels of PCF11 are maintained by an auto-regulatory mechanism involving premature termination of its own transcript, and are important for normal development. Both in human cell culture and during zebrafish development, PCF11 selectively attenuates the expression of other transcriptional regulators by premature CPA and termination.
2019-02-28 | GSE127256 | GEO
Project description:Functional inhibition of core spliceosomal machinery activates intronic premature cleavage and polyadenylation of pre-mRNAs
| PRJNA1195049 | ENA