P-TEFb function in ncRNA transcription
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ABSTRACT:
PROVIDER: PRJNA483416 | ENA |
REPOSITORIES: ENA
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Project description:Many ncRNAs serve as regulatory molecules in various physiological pathways. Distinct from protein-coding RNA expression, ncRNA expression is regulated solely by transcription and RNA processing/stability. Transcriptional regulation in ncRNA genes is thus important to be understood and is yet to be known completely. Previously, we identified that a subset of mammalian ncRNA genes is transcriptionally regulated by RNA Pol II promoter-proximal pausing and in a tissue-specific manner. In this study, human stimulus-inducible ncRNA genes were monitored to assess the function of P-TEFb, a master Pol II pausing regulator for protein-coding genes, in ncRNA transcription. Our findings indicate that the expression of many ncRNA genes is regulated by P-TEFb. Interestingly, a biphasic characteristic of P-TEFb inhibition in serum responsive ncRNA genes was observed: S2 phospho-Pol II was largely increased in the TSS (–300 to +300) whereas overall, it was decreased in the gene body (> +350) upon chemical inhibition of P-TEFb. In addition, the three representative ncRNAs, MALAT1, NEAT1, and XIST, were further analyzed for determining P-TEFb association. Taken together, we propose the transcriptional activation of many human ncRNAs utilizing pausing and releasing Pol II and the regulatory mechanism of transcriptional elongation of these genes requiring the function of P-TEFb.
2018-07-31 | GSE117848 | GEO
Project description:The P-TEFb complex promotes transcription elongation by releasing paused RNA polymerase II. P-TEFb itself is known to be inactivated through binding to the non-coding RNA 7SK but there is only limited information about mechanisms regulating their association. Here, we show that cells deficient in the RNA-binding protein hnRNP R, a known 7SK interactor, exhibit increased transcription due to phosphorylation of RNA polymerase II. Intriguingly, loss of hnRNP R promotes the release of P-TEFb from 7SK, accompanied by enhanced hnRNP A1 binding to 7SK. Additionally, we found that hnRNP R interacts with BRD4, and that hnRNP R depletion increases BRD4 binding to the P-TEFb component CDK9. Finally, CDK9 is stabilized upon loss of hnRNP R and its association with Cyclin K is enhanced. Together, our results indicate that hnRNP R negatively regulates transcription by modulating the activity and stability of the P-TEFb complex, exemplifying the multimodal regulation of P-TEFb by an RNA-binding protein.
2022-07-21 | GSE202720 | GEO
Project description:RUF6 is a ncRNA gene family that is transcribed by RNA Polymerase III but actively regulates the Pol II-transcribed var virulence gene family. Understanding how RUF6 ncRNA connect to downstream effectors is lacking. We developed an RNA-directed proteomic discovery (ChIRP-MS) protocol to identify in vivo RUF6 ncRNA protein interactions. The RUF6 ncRNA interactome was purified with biotinylated antisense oligonucleotides. Quantitative label-free mass spectrometry identified several unique proteins linked to gene transcription including. Affinity purification of Pf-DDX5 identified proteins originally found by our RUF6-ChIRP protocol, validating the technique’s robustness for identifying ncRNA interactomes in P. falciparum. Our work identifies a RUF6 ncRNA protein complex that interacts with RNA Pol II to sustain var gene expression.
2022-11-15 | PXD036801 | Pride
Project description:DNA damage response (DDR) involves dramatic transcriptional alterations, the mechanisms of which remain obscure. Here we show that following DNA damage, the RNA-binding motif protein 7 (RBM7) stimulates RNA polymerase II (Pol II) elongation and increases cell viability by activating the positive transcription elongation factor b (P-TEFb). This is mediated by DNA damage-enhanced binding of RBM7 to 7SK, the scaffold of the 7SK small nuclear ribonucleoprotein (snRNP) that inhibits P-TEFb. As a result, P-TEFb relocates from 7SK snRNP to chromatin to activate transcription of key DDR genes and multiple classes of non-coding RNAs. By alleviating the inhibition of P-TEFb, RBM7 thus promotes Pol II elongation to activate a transcriptional response that is crucial for cell fate upon genotoxic insult. Our work highlights a novel paradigm in stress-dependent control of Pol II pause release, and offers the promise of combating cancer by RBM7 and P-TEFb antagonists in combination with established DNA damage-inducing chemotherapeutics.
2018-06-01 | E-MTAB-6475 | biostudies-arrayexpress
Project description:Mediator is regarded a general co-activator of RNA-Polymerase II dependent transcription but not much is known about its function and regulation in mouse pluripotent embryonic stem cells (mESC). One means of controlling Mediator function is provided by binding of the Cdk8 module (Med12, Cdk8, Ccnc and Med13) to Mediator. Here we report that the Cdk8 module subunit Med12 operates together with PRC1 to silence developmental key genes in the pluripotent state. At the molecular level, PRC1 is required to assemble ncRNA containing Med12-Mediator complexes at promoters of repressed genes. In the course of cellular differentiation the H2A-ubiquitin binding protein Zrf1 abrogates PRC1-Med12 binding and facilitates the recruitment of Cdk8 into Mediator. Remodeling of the Mediator-associated protein complex converts Mediator into a transcriptional enhancer that mediates ncRNA-dependent activation of Polycomb target genes
2016-05-24 | GSE73352 | GEO
Project description:We showed that pharmacological inhibition of P-TEFb attenuated cyst development in ADPKD mouse models. Genome-wide analyses demonstrated that P-TEFb inhibition abrogated transcriptional pause release and suppressed a pathologic gene expression program during cystogenesis. This work revealed a mechanism by which aberrant activation of P-TEFb-mediated transcription elongation promotes cystogenesis in ADPKD.
2019-06-20 | GSE128521 | GEO
Project description:Releasing promoter-proximally-paused RNA polymerase II (RNAPII) by the positive transcription elongation factor b (P-TEFb) is a central regulatory step of eukaryotic mRNA synthesis. The nuclear activity of P-TEFb is controlled mainly by the 7SK small nuclear RNP (snRNP) which sequesters active P-TEFb into inactive 7SK/P-TEFb snRNP. Here we demonstrate that under normal culture conditions, the lack of 7SK snRNP has only a minor impact on global RNAPII transcription and promoter-proximal pausing without detectable consequences on cell growth and proliferation. However, upon ultraviolet (UV) light-induced DNA damage, cells lacking 7SK have a defective transcriptional response and highly reduced viability. Both UV-induced release of ‘lesion-scanning’ polymerases and activation of key early responsive genes are compromised in the absence of 7SK. We show that proper induction of 7SK-dependent UV-responsive genes, namely efficient release of paused RNAPII and recruitment of key elongation factors, requires P-TEFb activity directly mobilized from the nucleoplasmic 7SK/P-TEFb snRNP.
2021-03-30 | GSE147055 | GEO
Project description:Castration resistant prostate cancer (CRPC) is a lethal disease1-4. Aberrant activation of the androgen receptor (AR) becomes a central mechanism contributing to the resistance of endocrine therapies2,3. Here we demonstrate that non-coding RNAs transcribed from the AR bound-enhancers RNAs (AR-eRNAs) are upregulated in human CRPC cells in vitro, xenografts in vivo and patient tissues. Expression of a subset of genes with elevated AR-eRNAs, including TLE1 and HTR3A, is inversely correlated with biochemical recurrence-free survival of CRPC patients. We identify aan HIV-1 TAR-like (TAR-L) motif in AR-eRNAs of AR target genes including KLK3 (or PSA) and TMPRSS2. The TAR-L motif is important for these eRNAs to bind to CYCLIN T1 of the positive transcription elongation factor b (P-TEFb) complex. Knockdown of PSA eRNA diminishes RNA polymerase II (Pol II) serine-2 (Ser-2) phosphorylation at the PSA promoter. The TAR-L motif in KLK3 eRNA is crucial for effective transcription of PSA mRNA. Together, wWe demonstrate a P-TEFb activation function of eRNA and reveal aberrant eRNA expression as a functional indicator of AR abnormality in CRPC. Our results also suggest that eRNAs as amay be a potential target for CRPC therapy. Androgen receptor (AR) binding sites in human prostate cancer cell lines, LNCaP and C4-2, were studied using ChIP-seq. ChIP enriched and input DNA were sequenced using Illumina HiSeq 2000.
2016-02-12 | E-GEOD-55031 | biostudies-arrayexpress
Project description:The transcription factor T-bet induces differentiation of CD4+ T cells into the Th1 lineage and also allows for a degree of functional plasticity. Here, we show that T-bet acts through super-enhancers to recruit the elongation factor P-TEFb. Th1-specific genes are poised for activation in Th2 cells and P-TEFb recruitment activates transcriptional elongation. T-bet also induces extensive P-TEFb binding at super-enhancers, where it acts to stimulate enhancer RNA transcription. P-TEFb inhibition selectively blocks activation of lineage-specific genes and reverses Th1-associated retinitis pathology. T-bet-mediated recruitment of P-TEFb to super-enhancers at otherwise poised genes provides a model for how lineage-specifying factors promote differentiation towards specific cell fates whilst maintaining a degree of functional plasticity. Transcriptional profiling of mouse Th1 and Th2 cells treated with JQ1, which inhibits P-TEFb recruitment, or the P-TEFb inhibitor Flavopiridol vs DMSO conrol treated cells. Goal was to identify genes which are sensitive to P-TEFb inhibition.
2016-06-08 | GSE62485 | GEO
Project description:We deep sequenced wildtype and Tbx5-mutant mouse atria, identifying ~2600 novel Tbx5-dependent ncRNAs. Tbx5-dependent ncRNA transcription provided a quantitative metric of Tbx5-dependent enhancer activity, correlating with target gene expression.
2023-08-02 | GSE239805 | GEO