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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

DOI:http://dx.doi.org/10.7554/eLife.06535.025
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fig9: DOI:http://dx.doi.org/10.7554/eLife.06535.025

Mentions: We thank the reviewer for giving us an opportunity to explain this perplexing controversy. Indeed, in our own hands, knockdown of both CDK9 and CycT1 (∼10% CDK9 and 50% CycT1 remained in KD cells) caused only a partial reduction of the global pSer2 level (Author response image 2 Panel A; ∼30% remaining). However, when we looked at what was happening to the remaining CDK9 in the KD cells by measuring the signature 7SK snRNP component HEXIM1 and the signature Super Elongation Complex (SEC) components AFF4 and ELL2 that were associated with the immunoprecipitated CDK9 (Author response image 2 Panel B), we think we know the reason behind the partial pSer2 reduction. When the CDK9 signals were adjusted to a similar level (the CDK9 level in the KD cells was very low; and we had to use many times more the KD cells than the control in order to get a comparable CDK9 signal), it is obvious from the co-IP/Western analysis (Panel B) that there was a dramatic shift of P-TEFb from the 7SK snRNP to the SEC under the KD conditions. Since the vast majority of P-TEFb (up to 90% by some estimations) is normally sequestered in the 7SK snRNP, this shift basically has converted all the remaining P-TEFb into the active SEC complex. Thus, although the KD has significantly decreased the overall P-TEFb level in vivo, it failed to completely eliminate the active form of P-TEFb due to the drastic P-TEFb mobilization that was happening inside the cell, which allowed the detection of still a decent level of global pSer2 under these conditions.10.7554/eLife.06535.025Author response image 2.


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)

DOI:http://dx.doi.org/10.7554/eLife.06535.025
© Copyright Policy
Related In: Results  -  Collection

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

fig9: DOI:http://dx.doi.org/10.7554/eLife.06535.025
Mentions: We thank the reviewer for giving us an opportunity to explain this perplexing controversy. Indeed, in our own hands, knockdown of both CDK9 and CycT1 (∼10% CDK9 and 50% CycT1 remained in KD cells) caused only a partial reduction of the global pSer2 level (Author response image 2 Panel A; ∼30% remaining). However, when we looked at what was happening to the remaining CDK9 in the KD cells by measuring the signature 7SK snRNP component HEXIM1 and the signature Super Elongation Complex (SEC) components AFF4 and ELL2 that were associated with the immunoprecipitated CDK9 (Author response image 2 Panel B), we think we know the reason behind the partial pSer2 reduction. When the CDK9 signals were adjusted to a similar level (the CDK9 level in the KD cells was very low; and we had to use many times more the KD cells than the control in order to get a comparable CDK9 signal), it is obvious from the co-IP/Western analysis (Panel B) that there was a dramatic shift of P-TEFb from the 7SK snRNP to the SEC under the KD conditions. Since the vast majority of P-TEFb (up to 90% by some estimations) is normally sequestered in the 7SK snRNP, this shift basically has converted all the remaining P-TEFb into the active SEC complex. Thus, although the KD has significantly decreased the overall P-TEFb level in vivo, it failed to completely eliminate the active form of P-TEFb due to the drastic P-TEFb mobilization that was happening inside the cell, which allowed the detection of still a decent level of global pSer2 under these conditions.10.7554/eLife.06535.025Author response image 2.

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