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CDK9/cyclin complexes modulate endoderm induction by direct interaction with Mix.3/mixer.

ABSTRACT: Mix-related homeodomain proteins are involved in endoderm formation in the early vertebrate embryo. We used a yeast two-hybrid screen to identify proteins that interact with Mix.3/mixer to regulate endoderm induction. We demonstrate that cyclin-dependent kinase 9 (CDK9) interacts with the carboxyl terminal domain of Mix.3. CDK9 is the catalytic subunit of the PTEF-b transcription elongation complex that phosphorylates the C-terminal domain of RNA polymerase II to promote efficient elongation of nascent transcripts. Using whole embryo transcription reporter and animal pole explant assays, we show that Mix.3 activity is regulated by CDK9/cyclin complexes. Co-expression of cyclin T2 and cyclin K had different effects on Mix.3 transcriptional activity and endoderm induction. Our data suggest that binding of CDK9, and the recruitment of different cyclin partners, can modulate the endoderm-inducing activity of Mix.3 during embryonic development. Developmental Dynamics 238:1346-1357, 2009. (c) 2009 Wiley-Liss, Inc.

SUBMITTER: Zhu H

PROVIDER: S-EPMC2832930 | biostudies-literature | 2009 Jun

REPOSITORIES: biostudies-literature

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CDK9/cyclin complexes modulate endoderm induction by direct interaction with Mix.3/mixer.

Zhu Haiqing H Doherty Joanne R JR Kuliyev Emin E Mead Paul E PE

Developmental dynamics : an official publication of the American Association of Anatomists 20090601 6

Mix-related homeodomain proteins are involved in endoderm formation in the early vertebrate embryo. We used a yeast two-hybrid screen to identify proteins that interact with Mix.3/mixer to regulate endoderm induction. We demonstrate that cyclin-dependent kinase 9 (CDK9) interacts with the carboxyl terminal domain of Mix.3. CDK9 is the catalytic subunit of the PTEF-b transcription elongation complex that phosphorylates the C-terminal domain of RNA polymerase II to promote efficient elongation of ...[more]

PMID: 19347956

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Project description:BackgroundHIV-1 Tat activates RNA Polymerase II (RNAP II) elongation of the integrated provirus by recruiting a protein kinase known as P-TEFb to TAR RNA at the 5' end of nascent viral transcripts. The catalytic core of P-TEFb contains CDK9 and Cyclin T1 (CCNT1). A human endogenous complexome has recently been described - the set of multi-protein complexes in HeLa cell nuclei. We mined this complexome data set and identified 12 distinct multi-protein complexes that contain both CDK9 and CCNT1. We have termed these complexes CCAPs for CDK9/CCNT1-associated protein complexes. Nine CCAPs are novel, while three were previously identified as Core P-TEFb, the 7SK snRNP, and the Super-Elongation Complex. We have investigated the role of five newly identified CCAPs in Tat function and viral gene expression.ResultsWe examined five CCAPs that contain: 1) PPP1R10/TOX3/WDR82; 2) TTF2; 3) TPR; 4) WRNIP1; 5) FBXO11/CUL1/SKP1. SiRNA depletions of protein subunits of the five CCAPs enhanced Tat activation of an integrated HIV-1 LTR-Luciferase reporter in TZM-bl cells. Using plasmid transfection assays in HeLa cells, we also found that siRNA depletions of TTF2, FBXO11, PPP1R10, WDR82, and TOX3 enhanced Tat activation of an HIV-1 LTR-luciferase reporter, but the depletions did not enhance expression of an NF-κB reporter plasmid with the exception of PPP1R10. We found no evidence that depletion of CCAPs perturbed the level of CDK9/CCNT1 in the 7SK snRNP. We also found that the combination of siRNA depletions of both TTF2 and FBXO11 sensitized a latent provirus in Jurkat cells to reactivation by sub-optimal amounts of αCD3/CD28 antibodies.ConclusionsOur results identified five novel CDK9/CCNT1 complexes that are capable of negative regulation of HIV-1 Tat function and viral gene expression. Because siRNA depletions of CCAPs enhance Tat function, it is possible that these complexes reduce the level of CDK9 and CCNT1 available for Tat, similar to the negative regulation of Tat by the 7SK snRNP. Our results highlight the complexity in the biological functions of CDK9 and CCNT1.

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CDK9/cyclin complexes modulate endoderm induction by direct interaction with Mix.3/mixer.

Publications

CDK9/cyclin complexes modulate endoderm induction by direct interaction with Mix.3/mixer.

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