Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Expression profiling of HeLa cell line treated with CDK9 inhibitor

ABSTRACT: 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.

ORGANISM(S): Homo sapiens

PROVIDER: GSE60952 | GEO | 2015/06/17

SECONDARY ACCESSION(S): PRJNA260843

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Expression profiling of HeLa cell line treated with CDK9 inhibitor

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

Compensatory induction of MYC expression by sustained CDK9 inhibition via a BRD4-dependent mechanism: Pol II occupancy profiling by ChIP-Seq in HeLa cell line in the presence or absence of CDK9 inhibition

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

2015-06-17 | GSE60953 | GEO

JMJD5 couples with CDK9 to release the paused RNA polymerase II

Project description:More than 30% of genes in higher eukaryotes are regulated by RNA Polymerase II (Pol II) promoter proximal pausing. Pausing is released by the positive transcription elongation factor complex (P-TEFb). However, the exact mechanism by which this occurs and whether phosphorylation of the carboxyl-terminal domain of Pol II is involved in the process remains unknown. We previously reported that JMJD5 could generate tailless nucleosomes at position +1 from transcription start sites (TSS) thus perhaps enable progression of Pol II. Here we find that knockout of JMJD5 leads to accumulation of nucleosomes at position +1. Absence of JMJD5 also results in loss of or lowered transcription of a large number of genes. Interestingly, we found that phosphorylation, by CDK9, of Ser2 within two neighboring heptad repeats in the carboxy-terminal domain of Pol II together with phosphorylation of Ser5 within the second repeat, HR-Ser2p(1,2)-Ser5p(2) for short, allows Pol II to bind JMJD5 via engagement of the N-terminal domain of JMJD5. We suggest that these events bring JMJD5 near the nucleosome at position +1, thus allowing JMJD5 to clip histones on this nucleosome, a phenomenon that may contribute to release of Pol II pausing.

2020-06-27 | GSE153322 | GEO

RNA Polymerase II-associated factor 1 regulates the release and phosphorylation of paused RNA Polymerase II

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

The pause-initiation limit restricts transcription activation in human cells

Project description:Eukaryotic gene transcription is often controlled at the level of RNA polymerase II (Pol II) pausing in the promoter-proximal region. Pausing Pol II limits the frequency of transcription initiation (‘pause-initiation limit’), predicting that the pause duration must be decreased for transcriptional activation. To test this prediction, we conducted a genome-wide kinetic analysis of the heat shock response in human cells. We show that the pause-initiation limit restricts transcriptional activation at most genes. Gene activation generally requires the activity of the P-TEFb kinase CDK9, which decreases the duration of Pol II pausing and thereby enables an increase in the productive initiation frequency. The transcription of enhancer elements is generally not pause-limited and can be activated without CDK9 activity. Our results define the kinetics of Pol II transcriptional regulation in human cells at all gene classes during a natural transcription response.

2019-07-10 | GSE123980 | GEO

Integrative Proteomics Identifies MePCE as a Substrate of CDK9 for Releasing P-TEFb from 7SK snRNP Under Genotoxic Stress

Project description:The dynamic phosphorylation process regulated by the positive transcription elongation factor b (P-TEFb), a CDK9-cyclin T heterodimer, is critical for transcription elongation of numerous mammalian genes. Most P-TEFb in cells are normally sequestered in the 7SK snRNP, and their release from this complex is well-controlled to maintain proper cell homeostasis and growth. The comprehensive profiling of CDK9 substrates is essential for understanding diverse functions of P-TEFb in controlling gene expression. Here, integrative proteomics methodologies are developed to globally identify the potential substrates of CDK9 in vivo. Our results show that the 7SK methylphosphate capping enzyme MePCE is directly phosphorylated by CDK9 within the 7SK snRNP, which is known to exhibit little kinase activity toward other known CDK9 substrates. Upon ultraviolet radiation, MePCE becomes phosphorylated, leading to the dissociation of 7SK snRNP and release of P-TEFb to activate transcription. Meanwhile, the phosphorylation of MePCE does not affect its methyltransferase activity toward 7SK RNA. Our data suggest that even though CDK9 in 7SK snRNP has little kinase activity toward external substrates, it can phosphorylate the snRNP integral component MePCE, which is required to release P-TEFb. This novel inside-to-outside mode of P-TEFb activation may serve as a critical checkpoint in regulating gene transcription under stressful conditions.

2024-07-03 | PXD053627 |

Selective inhibition of CDK9 in triple negative breast cancer

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

RBM7 activates P-TEFb for a pro-survival transcriptional response to DNA damage

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

TFIID enables RNA polymerase II promoter-proximal pausing

Project description:RNA polymerase II (pol II) transcribes all protein-coding and many non-coding RNAs in the human genome. Pol II transcription initiation is governed by the Pre-Initiation Complex (PIC), which contains TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH, pol II, and Mediator. After initiation, pol II enzymes typically pause after transcribing less than 100 bases, and paused polymerases represent a common regulatory intermediate. Accordingly, paused pol II has been implicated in enhancer function, development and homeostasis, and diseases ranging from cancer to viral pathogenesis. Precisely how pol II promoter-proximal pausing is enforced and regulated remains unclear; however, protein complexes such as NELF and DSIF increase pausing whereas the activity of CDK9 (P-TEFb complex) correlates with pause release. To address specific mechanistic questions about pol II pausing and its regulation, we reconstituted human pol II promoter-proximal pausing in vitro, entirely with purified factors (no extracts). As expected, NELF and DSIF increased pol II pausing in vitro, whereas P-TEFb promoted pause release. Unexpectedly, the PIC alone was sufficient to reconstitute pol II pausing, suggesting that pausing is an inherent property of the PIC. In agreement, pol II pausing was lost upon replacement of the TFIID complex with TATA-binding protein (TBP); moreover, pausing was dependent upon TFIID subunits TAF1 and TAF2. TAF1/2 bind genomic DNA downstream of the pol II initiation site, invoking a “complex interaction” model for pausing. Consistent with this model, PRO-Seq experiments revealed increased transcription upon acute depletion (t=60 min) of TAF1 and TAF2 in human cells, and pol II pausing was disrupted at thousands of genes. Similar results were obtained in TAF1-depleted Drosophila S2 cells. Collectively, these data establish the general transcription factor TFIID as a genome-wide regulator of pol II promoter-proximal pausing.

2020-05-28 | GSE132764 | GEO

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data