Ontology highlight
ABSTRACT:
PROVIDER: PRJNA260842 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR1561935.fastq.gz | Fastqsanger.gz | |||
| SRR1561936.fastq.gz | Fastqsanger.gz | |||
| SRR1561937.fastq.gz | Fastqsanger.gz | |||
| SRR1561938.fastq.gz | Fastqsanger.gz | |||
| SRR1561939.fastq.gz | Fastqsanger.gz |
Items per page: 5 1 - 5 of 5 |
Similar Datasets
Project description:CDK9 is the kinase subunit of P-TEFb that enables RNA polymerase (Pol) II to transit from promoter-proximal pausing to productive elongation. Although considerable interest exists in CDK9 as a therapeutic target, little progress has been made due to the lack of highly selective inhibitors. Here, we describe the development of i-CDK9 as such an inhibitor that potently suppresses CDK9 phosphorylation of substrates and causes genome-wide Pol II pausing. While most genes experience reduced expression, MYC and other primary response genes increase expression upon sustained i-CDK9 treatment. Essential for this increase, the bromodomain protein BRD4 captures P-TEFb from 7SK snRNP to deliver to target genes and also enhances CDK9’s activity and resistance to inhibition. Because the i-CDK9-induced MYC expression and binding to P-TEFb compensate for P-TEFb’s loss of activity, only the simultaneous inhibition of CDK9 and MYC can efficiently induce growth arrest and apoptosis of cancer cells, suggesting the potential of a combinatorial treatment strategy. We used microarrays to examine the global impact on gene expression by imhibiting CDK9 at different time durations. HeLa cell lines treated with CDK9 inhibitor at different time points
2015-06-17 | E-GEOD-60952 | biostudies-arrayexpress
Project description:Expression profiling of HeLa cell line treated with CDK9 inhibitor
| PRJNA260843 | ENA
Project description:Chromatin is highly condensed and transcriptionally repressed during mitosis. Although it is established that some general transcription factors are inactivated by phosphorylation at mitosis, many details of mitotic transcriptional repression and its underlying mechanisms are largely unknown. Here, we provide evidence that as cells enter mitosis, genes with transcriptionally engaged RNA Polymerase II (Pol II) can continue transcription until the end of the gene to clear Pol II from mitotic chromatin. Using ChIP-Seq, we find that the transcriptional reinitiation process is globally impaired in early mitosis (prophase/prometaphase), with loss of TFIIB occupancy and nucleosome-free regions at promoters. Pretreatment of nocodazole-arrested mitotic cells with the P-TEFb inhibitor flavopiridol prevents the release of promoter-proximal engaged Pol II. Global nascent RNA sequencing and RNA fluorescence in situ hybridization (FISH) of individual genes demonstrate the existence of transcriptionally engaged Pol II in early mitosis. Chemical and mutational inhibition of P-TEFb in mitosis leads to delays in the progression of cell division. Together, our study reveals a novel mechanism for mitotic transcriptional repression whereby transcriptionally engaged Pol II can progress into productive elongation and finish transcription to allow proper cellular division. ChIP-Seq of Pol II of different forms, TFIIB, H3K4me3 in human HeLa cells at different cell cycle stages. ChIP-Seq of Pol II in HeLa mitotic cells with or without CDK9 inhibitor flavopiridol pretreatment. Nascent RNA-seq in asynchronous and arrested mitotic cells.
2015-10-30 | E-GEOD-71848 | biostudies-arrayexpress
Project description:CDK9 is the kinase subunit of P-TEFb that enables RNA polymerase (Pol) II to transit from promoter-proximal pausing to productive elongation. Although considerable interest exists in CDK9 as a therapeutic target, little progress has been made due to the lack of highly selective inhibitors. Here, we describe the development of i-CDK9 as such an inhibitor that potently suppresses CDK9 phosphorylation of substrates and causes genome-wide Pol II pausing. While most genes experience reduced expression, MYC and other primary response genes increase expression upon sustained i-CDK9 treatment. Essential for this increase, the bromodomain protein BRD4 captures P-TEFb from 7SK snRNP to deliver to target genes and also enhances CDK9’s activity and resistance to inhibition. Because the i-CDK9-induced MYC expression and binding to P-TEFb compensate for P-TEFb’s loss of activity, only the simultaneous inhibition of CDK9 and MYC can efficiently induce growth arrest and apoptosis of cancer cells, suggesting the potential of a combinatorial treatment strategy. ChIP-seq of Pol II in HeLa cells before or after i-CDk9 treatment
2015-06-17 | E-GEOD-60953 | biostudies-arrayexpress
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:Release of promoter-proximal paused RNA polymerase II (Pol II) during early elongation is a critical step in transcriptional regulation in metazoan cells. Paused Pol II release is thought to require the kinase activity of cyclin-dependent kinase 9 (CDK9) for the phosphorylation of DRB sensitivity-inducing factor, negative elongation factor, and C-terminal domain (CTD) serine-2 of Pol II. We found that Pol II-associated factor 1 (PAF1) is a critical regulator of paused Pol II release, that positive transcription elongation factor b (P-TEFb) directly regulates the initial recruitment of PAF1 complex (PAF1C) to genes, and that the subsequent recruitment of CDK12 is dependent on PAF1C. These findings reveal cooperativity among P-TEFb, PAF1C, and CDK12 in pausing release and Pol II CTD phosphorylation. Comparison of the chromatin occupancy of [1] PAF1, CDC73, LEO1, CTR9, total Pol II, and CTD serine 2-phosphorylated Pol II by ChIP-seq in THP1 cells; [2] PAF1, Pol II, Pol II (ser-5p), CDK12, and CDK9 by ChIP-seq in control and PAF1 knockdown cells; [3] LEO1 and Pol II by ChIP-seq in control and flavopiridol treated THP1 cells.
2015-12-11 | E-GEOD-62171 | biostudies-arrayexpress
Project description:CDK9 is a critical kinase required for the productive transcription of protein-coding genes by RNA polymerase II (pol II) in higher eukaryotes. Phosphorylation of targets including the elongation factor SPT5 and the carboxyl-terminal domain (CTD) of RNA pol II allow the polymerase to pass an early elongation checkpoint (EEC), which is encountered soon after initiation. In addition to halting RNA polymerase II at the EEC, CDK9 inhibition also 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, suggesting that CDK9 and PP2A, working together, regulate the coupling of elongation and transcription termination to RNA maturation. Our phosphoproteomic analyses, using either DRB or an analog-sensitive CDK9 cell line confirm the splicing factor SF3B1 as an additional key target of this kinase. CDK9 inhibition causes loss of interaction of splicing and export factors with SF3B1, suggesting that CDK9 also helps to co-ordinates coupling of splicing and export to transcription.
2022-08-22 | PXD026720 | Pride
Project description:Targeted treatment for triple-negative breast cancer (TNBC) remains an elusive clinical challenge. Cyclin-dependent kinase 9 (CDK9) is a transcriptional regulator shown to promote growth of multiple cancers, including TNBC, and represents a potential new therapeutic target. CDK9 increases RNA Polymerase II (Pol II) activity, which facilitates sustained expression of short-lived oncogenic and anti-apoptotic proteins. However, pre-clinical evaluation of CDK9 as a therapeutic target has been hampered by the poor selectivity of existing CDK9 inhibitors (CDK9i). Here, we report a novel CDK9i; D11-8, which exhibits high potency against CDK9 (Ki = 8 nM) and displays remarkable selectivity over other CDKs and human kinases. D11-8 inhibited TNBC cell line proliferation,, induced G2/M cell cycle arrest, and increased apoptosis. Mechanistically, D11-8 inhibited CDK9; reducing Pol II phosphorylation, down-regulating expression of proto-oncogene MYC and anti-apoptotic marker MCL1, and dramatically inducing Pol II promoter-proximal pausing at MYC, G2/M checkpoint, and E2F2 target genes. D11-8 was further validated for TNBC in vivo, and inhibited TNBC cell line tumour growth without toxicity. To further elucidate the cancer cell specificity of D11-8, normal breast tissue was collected from women undergoing reduction mammoplasty surgery and treated with D11-8 ex vivo as patient-derived explants. High doses of D11-8 (2.7 µM) had no effect on the proliferative capacity of normal breast epithelial cells (Ki67 positivity), and tissues appeared histologically normal. Collectively, these data demonstrate that D11-8 effectively inhibits CDK9 in TNBC cell lines, resulting in growth inhibition without short-term toxicity.
2023-11-25 | GSE184335 | GEO
Project description:MM1.S cells are an aggressive dexamethasone sensitive multiple myeloma cell line whose transcritional program is driven by deregulated c-Myc activity. We present ChIP-seq analysis of key transcritional regulators that are implicated the c-Myc transcriptional network in MM1.S cells treated with vehicle or 500nM JQ1. Brd4, Cdk9, cMyc, Max, Med1, RNA Pol II, and the chromatin modifications H3K4me3 and H3K27Ac were profiled in MM1.S cells treated with 500nM JQ1 for 24hr
2012-11-30 | E-GEOD-42161 | biostudies-arrayexpress
Project description:Transcription steps are marked by different modifications of the C-terminal domain of RNA polymerase II (RNAPII). Phosphorylation of Ser5 and Ser7 by cyclin-dependent kinase 7 (CDK7) as part of TFIIH marks initiation, whereas phosphorylation of Ser2 by CDK9 marks elongation. These processes are thought to take place in localized transcription foci in the nucleus, known as M-bM-^@M-^XM-bM-^@M-^Xtranscription factories,M-bM-^@M-^YM-bM-^@M-^Y but it has been argued that the observed clusters/foci are mere fixation or labeling artifacts. We show that transcription factories exist in living cells as distinct foci by live-imaging fluorescently labeled CDK9, a kinase known to associate with active RNAPII. These foci were observed in different cell types derived from CDK9-mCherry knock-in mice. We show that these foci are very stable while highly dynamic in exchanging CDK9. Chromatin immunoprecipitation (ChIP) coupled with deep sequencing (ChIP-seq) data show that the genome-wide binding sites of CDK9 and initiating RNAPII overlap on transcribed genes. Immunostaining shows that CDK9- mCherry foci colocalize with RNAPII-Ser5P, much less with RNAPII-Ser2P, and not with CDK12 (a kinase reported to be involved in the Ser2 phosphorylation) or with splicing factor SC35. In conclusion, transcription factories exist in living cells, and initiation and elongation of transcripts takes place in different nuclear compartments. Examination of genome occupancy of CDK9 and RNAPII that was performed by ChIP-seq in the MEL cell line as described (Soler et al. 2010, 2011) using CDK9 C20 antibody (Santa Cruz Biotechnology, C20, sc-484) and RNA Pol II antibody (Santa Cruz Biotechnology, N20, sc899),
2013-05-13 | E-GEOD-46849 | biostudies-arrayexpress