Pol II phosphorylation regulates a switch between transcriptional and splicing condensates
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA493945 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR7943783.fastq.gz | Fastqsanger.gz | |||
| SRR7943784.fastq.gz | Fastqsanger.gz | |||
| SRR7943785.fastq.gz | Fastqsanger.gz | |||
| SRR7943786.fastq.gz | Fastqsanger.gz |
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Project description:The synthesis of pre-mRNA by RNA polymerase II (Pol II) involves the formation of a transcription initiation complex, and a transition to an elongation complex. The large subunit of Pol II contains an intrinsically disordered C-terminal domain that is phosphorylated by cyclin-dependent kinases during the transition from initiation to elongation, thus influencing the interaction of the C-terminal domain with different components of the initiation or the RNA-splicing apparatus. Recent observations suggest that this model provides only a partial picture of the effects of phosphorylation of the C-terminal domain. Both the transcription-initiation machinery and the splicing machinery can form phase-separated condensates that contain large numbers of component molecules: hundreds of molecules of Pol II and mediator are concentrated in condensates at super-enhancers, and large numbers of splicing factors are concentrated in nuclear speckles, some of which occur at highly active transcription sites. Here we investigate whether the phosphorylation of the Pol II C-terminal domain regulates the incorporation of Pol II into phase-separated condensates that are associated with transcription initiation and splicing. We find that the hypophosphorylated C-terminal domain of Pol II is incorporated into mediator condensates and that phosphorylation by regulatory cyclin-dependent kinases reduces this incorporation. We also find that the hyperphosphorylated C-terminal domain is preferentially incorporated into condensates that are formed by splicing factors. These results suggest that phosphorylation of the Pol II C-terminal domain drives an exchange from condensates that are involved in transcription initiation to those that are involved in RNA processing, and implicates phosphorylation as a mechanism that regulates condensate preference.
2019-06-17 | GSE120656 | GEO
Project description:The synthesis of pre-mRNA by RNA polymerase II (Pol II) involves the formation of a transcription initiation complex, and a transition to an elongation complex1-4. The large subunit of Pol II contains an intrinsically disordered C-terminal domain that is phosphorylated by cyclin-dependent kinases during the transition from initiation to elongation, thus influencing the interaction of the C-terminal domain with different components of the initiation or the RNA-splicing apparatus5,6. Recent observations suggest that this model provides only a partial picture of the effects of phosphorylation of the C-terminal domain7-12. Both the transcription-initiation machinery and the splicing machinery can form phase-separated condensates that contain large numbers of component molecules: hundreds of molecules of Pol II and mediator are concentrated in condensates at super-enhancers7,8, and large numbers of splicing factors are concentrated in nuclear speckles, some of which occur at highly active transcription sites9-12. Here we investigate whether the phosphorylation of the Pol II C-terminal domain regulates the incorporation of Pol II into phase-separated condensates that are associated with transcription initiation and splicing. We find that the hypophosphorylated C-terminal domain of Pol II is incorporated into mediator condensates and that phosphorylation by regulatory cyclin-dependent kinases reduces this incorporation. We also find that the hyperphosphorylated C-terminal domain is preferentially incorporated into condensates that are formed by splicing factors. These results suggest that phosphorylation of the Pol II C-terminal domain drives an exchange from condensates that are involved in transcription initiation to those that are involved in RNA processing, and implicates phosphorylation as a mechanism that regulates condensate preference.
| S-EPMC6706314 | biostudies-literature
Project description:This SuperSeries is composed of the following subset Series: GSE30995: An Alternative Splicing Switch Regulates Embryonic Stem Cell Pluripotency and Reprogramming [RNA-Seq] GSE31006: An Alternative Splicing Switch Regulates Embryonic Stem Cell Pluripotency and Reprogramming [ChIP-Seq] GSE31007: An Alternative Splicing Switch Regulates Embryonic Stem Cell Pluripotency and Reprogramming [protein binding microarray] GSE31948: An Alternative Splicing Switch Regulates Embryonic Stem Cell Pluripotency and Reprogramming [AS microarray] Refer to individual Series
2011-09-14 | E-GEOD-30992 | biostudies-arrayexpress
Project description:The C-terminal domain of RPB1 (CTD) orchestrates transcription by recruiting regulators to RNA Pol II upon phosphorylation. Recent insights highlight CTD’s pivotal role in driving condensate formation on gene loci. Yet, the molecular mechanism behind how CTD-mediated recruitment of transcriptional regulators influences condensates formation remains unclear. Our study unveils that phosphorylation reversibly dissolves phase separation induced by the unphosphorylated CTD. Phosphorylated CTD, upon specific association with transcription regulatory proteins, forms distinct condensates from unphosphorylated CTD. Function studies demonstrate CTD variants with diverse condensation properties in vitro exhibit difference in promoter binding and mRNA co-processing in cells. Notably, varying CTD lengths lead to alternative splicing outcomes impacting cellular growth, linking the evolution of CTD variation/length with the complexity of splicing from yeast to human. These findings provide compelling evidence for a model wherein post-translational modification enables the transition of functionally specialized condensates, highlighting a co-evolution link between CTD condensation and splicing.
2024-02-09 | GSE252260 | GEO
Project description:The C-terminal domain of RPB1 (CTD) orchestrates transcription by recruiting regulators to RNA Pol II upon phosphorylation. Recent insights highlight CTD’s pivotal role in driving condensate formation on gene loci. Yet, the molecular mechanism behind how CTD-mediated recruitment of transcriptional regulators influences condensates formation remains unclear. Our study unveils that phosphorylation reversibly dissolves phase separation induced by the unphosphorylated CTD. Phosphorylated CTD, upon specific association with transcription regulatory proteins, forms distinct condensates from unphosphorylated CTD. Function studies demonstrate CTD variants with diverse condensation properties in vitro exhibit difference in promoter binding and mRNA co-processing in cells. Notably, varying CTD lengths lead to alternative splicing outcomes impacting cellular growth, linking the evolution of CTD variation/length with the complexity of splicing from yeast to human. These findings provide compelling evidence for a model wherein post-translational modification enables the transition of functionally specialized condensates, highlighting a co-evolution link between CTD condensation and splicing.
2024-02-09 | GSE252258 | GEO
Project description:This SuperSeries is composed of the following subset Series: GSE31485: CTCF promotes RNA pol II pausing and links DNA methylation to alternative splicing [ChIP-Seq] GSE31486: CTCF promotes RNA pol II pausing and links DNA methylation to alternative splicing [RNA-Seq] Refer to individual Series
2011-10-05 | E-GEOD-31278 | biostudies-arrayexpress
Project description:Gene activation is thought to involve a multistep process whereby transcription factors bind to distal enhancer sites and recruit the Mediator complex which contacts the pre-initiating RNA Polymerase II (Pol II) complex assembled at the start site of the gene. The interaction of Mediator and Pol II has yet to be observed in the nucleus of living cells and the dynamics of this interaction are not yet elucidated. Here we use quantitative live cell super-resolution and light sheet imaging to study the organization and dynamics of endogenous Mediator and Pol II directly in living mouse embryonic stem cells. In addition to forming transient clusters with average lifetimes of 11.1 (± 0.9) s, and 12.1 (± 1.4) s respectively, Mediator and Pol II also form large and stable clusters in stem cells (~15 stable clusters per cell). The large and stable Mediator and Pol II clusters gradually disappear within hours after induction of stem cell differentiation. Mediator and Pol II colocalize in the large clusters. Inhibition of Brd4 bromodomains necessary for enhancer association eliminates both Mediator and Pol II stable clusters, and inhibition of transcription elongation selectively eliminates stable Pol II but not stable Mediator clusters. Tracking of Mediator and Pol II stable clusters suggests they are chromatin associated and they coalesce upon contact, a property associated with phase separated droplets. We conclude that Mediator and Pol II associate in diffraction-sized condensates with a defined lifetime dependent on active transcription in living stem cells.
2018-07-22 | GSE115436 | GEO
Project description:Material properties of biomolecular condensates regulates RNA splicing and tumorigenesis
| PRJNA504562 | ENA
Project description:It remains largely unclear how material properties of biomolecular condensates impact physiological and pathological processes such as cancer. Here we show that AKAP95, a nuclear protein involved in gene regulation, plays an important role in tumorigenesis through directly regulating mRNA splicing. We show that AKAP95, through an intrinsic disordered region (IDR), forms liquid-like droplets in vitro and dynamic foci in nucleus. We have also identified key residues in IDR whose mutations to different residues perturb AKAP95 condensation in opposite directions. Importantly, the biological activity of AKAP95 in splice regulation is affected by either disruption of liquid condensation or enhancing the gelation of the assembly. We then show that the ability of AKAP95 in overcoming oncogene-induced senescence requires AKAP95 in an appropriate window of liquidity. These results thus link phase separation to tumorigenesis and uncover an important role of regulating the material property of protein condensates in disease including cancer.
2020-05-06 | GSE122308 | GEO
Project description:Biomolecular condensates play important roles in diverse biological processes. Many viruses form biomolecular condensates which have been implicated in various functions critical for productive infection of host cells. The adenovirus L1-52/55 kilodalton protein (52K) was recently shown to form viral biomolecular condensates that coordinate viral genome packaging and capsid assembly. Although critical for packaging, we do not know how viral condensates are regulated during adenovirus infection. Here we show that phosphorylation of serine residues 28 and 75 within the N-terminal intrinsically disordered region of 52K modulates viral condensates in vitro and in cells, promoting liquid-like properties over condensate hardening. Furthermore, we demonstrate that phosphorylation of 52K promotes viral genome packaging and production of infectious progeny particles. Collectively, our findings provide insights into how viral condensate properties are regulated and maintained in a state conducive to their function in viral progeny production. In addition, our findings have implications for antiviral strategies aimed at targeting the regulation of viral biomolecular condensates to limit viral multiplication.
2023-12-06 | PXD046971 | Pride