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Compensatory induction of MYC expression by sustained CDK9 inhibition via a BRD4-dependent mechanism.

Lu H, Xue Y, Xue Y, Yu GK, Arias C, Lin J, Fong S, Faure M, Weisburd B, Ji X, Mercier A, Sutton J, Luo K, Gao Z, Zhou Q - Elife (2015)

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.

ABSTRACT
CDK9 is the kinase subunit of positive transcription elongation factor b (P-TEFb) that enables RNA polymerase (Pol) II's transition from promoter-proximal pausing to productive elongation. Although considerable interest exists in CDK9 as a therapeutic target, little progress has been made due to 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 simultaneously inhibiting CDK9 and MYC/BRD4 can efficiently induce growth arrest and apoptosis of cancer cells, suggesting the potential of a combinatorial treatment strategy.

No MeSH data available.


Related in: MedlinePlus

Examination of the purity and concentrations of WT and ∆PID BRD4 used in the CDK9 kinase assay.DOI:http://dx.doi.org/10.7554/eLife.06535.015
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fig4s5: Examination of the purity and concentrations of WT and ∆PID BRD4 used in the CDK9 kinase assay.DOI:http://dx.doi.org/10.7554/eLife.06535.015

Mentions: To this end, the ability of recombinant CDK9-CycT1 (Invitrogen) to phosphorylate a synthetic Pol II CTD peptide (termed CDK7tide; Bio-Synthesis, Inc.) was measured in the presence of either WT BRD4 or a BRD4 mutant lacking the C-terminal P-TEFb-interacting domain (∆PID). The BRD4 proteins were affinity-purified from transfected HEK293T cells under highly stringent conditions to strip away their associated factors (Figure 4—figure supplement 5). While WT BRD4 was able to increase CDK9 kinase activity dose dependently up to 2.2-fold, ∆PID lacked this ability (Figure 4I), indicating that the physical interaction between BRD4 and P-TEFb was essential for the elevated CDK9 activity. It is worth pointing out that different from a recent study reporting that BRD4 is an atypical kinase that can directly phosphorylate the Pol II CTD on Ser2 (Devaiah et al., 2012), neither WT nor ∆PID BRD4 alone was able to cause phosphorylation of the CTD peptide in the absence of P-TEFb (Figure 4I).


Compensatory induction of MYC expression by sustained CDK9 inhibition via a BRD4-dependent mechanism.

Lu H, Xue Y, Xue Y, Yu GK, Arias C, Lin J, Fong S, Faure M, Weisburd B, Ji X, Mercier A, Sutton J, Luo K, Gao Z, Zhou Q - Elife (2015)

Examination of the purity and concentrations of WT and ∆PID BRD4 used in the CDK9 kinase assay.DOI:http://dx.doi.org/10.7554/eLife.06535.015
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4490784&req=5

fig4s5: Examination of the purity and concentrations of WT and ∆PID BRD4 used in the CDK9 kinase assay.DOI:http://dx.doi.org/10.7554/eLife.06535.015
Mentions: To this end, the ability of recombinant CDK9-CycT1 (Invitrogen) to phosphorylate a synthetic Pol II CTD peptide (termed CDK7tide; Bio-Synthesis, Inc.) was measured in the presence of either WT BRD4 or a BRD4 mutant lacking the C-terminal P-TEFb-interacting domain (∆PID). The BRD4 proteins were affinity-purified from transfected HEK293T cells under highly stringent conditions to strip away their associated factors (Figure 4—figure supplement 5). While WT BRD4 was able to increase CDK9 kinase activity dose dependently up to 2.2-fold, ∆PID lacked this ability (Figure 4I), indicating that the physical interaction between BRD4 and P-TEFb was essential for the elevated CDK9 activity. It is worth pointing out that different from a recent study reporting that BRD4 is an atypical kinase that can directly phosphorylate the Pol II CTD on Ser2 (Devaiah et al., 2012), neither WT nor ∆PID BRD4 alone was able to cause phosphorylation of the CTD peptide in the absence of P-TEFb (Figure 4I).

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.

ABSTRACT
CDK9 is the kinase subunit of positive transcription elongation factor b (P-TEFb) that enables RNA polymerase (Pol) II's transition from promoter-proximal pausing to productive elongation. Although considerable interest exists in CDK9 as a therapeutic target, little progress has been made due to 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 simultaneously inhibiting CDK9 and MYC/BRD4 can efficiently induce growth arrest and apoptosis of cancer cells, suggesting the potential of a combinatorial treatment strategy.

No MeSH data available.


Related in: MedlinePlus