Distinct layers of BRD4-PTEFb reveal bromodomain-independent function in transcriptional regulation

Bin Zheng; Sarah Gold; Marta Iwanaszko; Benjamin Charles Howard; Lu Wang; Ali Shilatifard

doi:10.1016/j.molcel.2023.06.032

Distinct layers of BRD4-PTEFb reveal bromodomain-independent function in transcriptional regulation

Bin Zheng, Sarah Gold, Marta Iwanaszko, Benjamin Charles Howard, Lu Wang^*, Ali Shilatifard^*

^*Corresponding author for this work

Biochemistry and Molecular Genetics

Research output: Contribution to journal › Article › peer-review

21 Scopus citations

Abstract

The BET family protein BRD4, which forms the CDK9-containing BRD4-PTEFb complex, is considered to be a master regulator of RNA polymerase II (Pol II) pause release. Because its tandem bromodomains interact with acetylated histone lysine residues, it has long been thought that BRD4 requires these bromodomains for its recruitment to chromatin and transcriptional regulatory function. Here, using rapid depletion and genetic complementation with domain deletion mutants, we demonstrate that BRD4 bromodomains are dispensable for Pol II pause release. A minimal, bromodomain-less C-terminal BRD4 fragment containing the PTEFb-interacting C-terminal motif (CTM) is instead both necessary and sufficient to mediate Pol II pause release in the absence of full-length BRD4. Although BRD4-PTEFb can associate with chromatin through acetyl recognition, our results indicate that a distinct, active BRD4-PTEFb population functions to regulate transcription independently of bromodomain-mediated chromatin association. These findings may enable more effective pharmaceutical modulation of BRD4-PTEFb activity.

Original language	English (US)
Pages (from-to)	2896-2910.e4
Journal	Molecular cell
Volume	83
Issue number	16
DOIs	https://doi.org/10.1016/j.molcel.2023.06.032
State	Published - Aug 17 2023

Funding

We thank Brianna Monroe for illustrating the findings of this study in the model. We want to thank Emily Rendleman, Sheetal Ganesan, Cassy Philips, and Jacob Zeidner for initial NGS sequencing and Yue He for her participation in the cloning for GFP-tagged ΔBD and C. We also want to thank Drs. Edwin Smith, Marc Morgan, Yuki Aoi and Karen Adelman for in-depth discussions and suggestions for the study. We are grateful for all the current and past Shilatifard lab members for their support. L.W. laboratory is supported by NIH grant R35GM146979. Studies in the Shilatifard lab are supported by generous funding through the Outstanding Investigator Award mechanism by the National Cancer Institute grant R35-CA197569 .

Keywords

BRD4
JQ1
PTEFb
Pol II
bromodomains
dBET6
elongation
pause release
rapid depletion

ASJC Scopus subject areas

Molecular Biology
Cell Biology

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10.1016/j.molcel.2023.06.032

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title = "Distinct layers of BRD4-PTEFb reveal bromodomain-independent function in transcriptional regulation",

abstract = "The BET family protein BRD4, which forms the CDK9-containing BRD4-PTEFb complex, is considered to be a master regulator of RNA polymerase II (Pol II) pause release. Because its tandem bromodomains interact with acetylated histone lysine residues, it has long been thought that BRD4 requires these bromodomains for its recruitment to chromatin and transcriptional regulatory function. Here, using rapid depletion and genetic complementation with domain deletion mutants, we demonstrate that BRD4 bromodomains are dispensable for Pol II pause release. A minimal, bromodomain-less C-terminal BRD4 fragment containing the PTEFb-interacting C-terminal motif (CTM) is instead both necessary and sufficient to mediate Pol II pause release in the absence of full-length BRD4. Although BRD4-PTEFb can associate with chromatin through acetyl recognition, our results indicate that a distinct, active BRD4-PTEFb population functions to regulate transcription independently of bromodomain-mediated chromatin association. These findings may enable more effective pharmaceutical modulation of BRD4-PTEFb activity.",

keywords = "BRD4, JQ1, PTEFb, Pol II, bromodomains, dBET6, elongation, pause release, rapid depletion",

author = "Bin Zheng and Sarah Gold and Marta Iwanaszko and Howard, {Benjamin Charles} and Lu Wang and Ali Shilatifard",

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AU - Gold, Sarah

AU - Iwanaszko, Marta

AU - Howard, Benjamin Charles

AU - Wang, Lu

AU - Shilatifard, Ali

PY - 2023/8/17

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N2 - The BET family protein BRD4, which forms the CDK9-containing BRD4-PTEFb complex, is considered to be a master regulator of RNA polymerase II (Pol II) pause release. Because its tandem bromodomains interact with acetylated histone lysine residues, it has long been thought that BRD4 requires these bromodomains for its recruitment to chromatin and transcriptional regulatory function. Here, using rapid depletion and genetic complementation with domain deletion mutants, we demonstrate that BRD4 bromodomains are dispensable for Pol II pause release. A minimal, bromodomain-less C-terminal BRD4 fragment containing the PTEFb-interacting C-terminal motif (CTM) is instead both necessary and sufficient to mediate Pol II pause release in the absence of full-length BRD4. Although BRD4-PTEFb can associate with chromatin through acetyl recognition, our results indicate that a distinct, active BRD4-PTEFb population functions to regulate transcription independently of bromodomain-mediated chromatin association. These findings may enable more effective pharmaceutical modulation of BRD4-PTEFb activity.

AB - The BET family protein BRD4, which forms the CDK9-containing BRD4-PTEFb complex, is considered to be a master regulator of RNA polymerase II (Pol II) pause release. Because its tandem bromodomains interact with acetylated histone lysine residues, it has long been thought that BRD4 requires these bromodomains for its recruitment to chromatin and transcriptional regulatory function. Here, using rapid depletion and genetic complementation with domain deletion mutants, we demonstrate that BRD4 bromodomains are dispensable for Pol II pause release. A minimal, bromodomain-less C-terminal BRD4 fragment containing the PTEFb-interacting C-terminal motif (CTM) is instead both necessary and sufficient to mediate Pol II pause release in the absence of full-length BRD4. Although BRD4-PTEFb can associate with chromatin through acetyl recognition, our results indicate that a distinct, active BRD4-PTEFb population functions to regulate transcription independently of bromodomain-mediated chromatin association. These findings may enable more effective pharmaceutical modulation of BRD4-PTEFb activity.

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KW - pause release

KW - rapid depletion

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Distinct layers of BRD4-PTEFb reveal bromodomain-independent function in transcriptional regulation

Abstract

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