Requirement for a kinase-specific chaperone pathway in the production of a Cdk9/cyclin T1 heterodimer responsible for P-TEFb-mediated tat stimulation of HIV-1 transcription

被引:124
作者
O'Keeffe, B [1 ]
Fong, Y [1 ]
Chen, D [1 ]
Zhou, S [1 ]
Zhou, Q [1 ]
机构
[1] Univ Calif Berkeley, Dept Mol & Cell Biol, Berkeley, CA 94720 USA
关键词
D O I
10.1074/jbc.275.1.279
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Tat activation of HIV-1 transcription is mediated by human transcription elongation factor P-TEFb, which interacts with Tat and phosphorylates the C-terminal domain of RNA polymerase II. The catalytic subunit of the P-TEFb complex, Cdk9, has been shown to interact with cyclin T and several other proteins of unknown identity, Consequently, the exact subunit composition of active P-TEFb has not been determined. Here we report the affinity purification and identification of the Cdk9 associated proteins. In addition to forming a heterodimer with cyclin T1, Cdk9 interacted with the molecular chaperone Hsp76 or a kinase-specific chaperone complex, Hsp90/Cdc37, to form two separate chaperone-Cdk9 complexes, Although the Cdk9/cyclin T1 dimer was exceptionally stable and produced slowly in the cell, free and unprotected Cdk9 appeared to be degraded rapidly, Several lines of evidence indicate the heterodimer of Cdk9/cyclin T1 to be the mature, active form of P-TEFb responsible for phosphorylation of the C-terminal domain of RNA polymerase II interaction with the Tat activation domain, and mediation of Tat activation of HIV-1 transcription. Pharmacological inactivation of Hsp90/Cdc37 function by geldanamycin revealed an essential role for the chaperone-Cdk9 complexes in generation of Cdk9/cyclin TI, Our data suggest a previously unrecognized chaperone-dependent pathway involving the sequential actions of Hsp70 and Hsp90/Cdc37 in the stabilization/folding of Cdk9 as well as the assembly of an active Cdk9/cyclin T1 complex responsible for P-TEFb-mediated Tat transactivation.
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页码:279 / 287
页数:9
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