HIV-1 tat interaction with RNA polymerase IIC-terminal domain (CTD) and a dynamic association with CDK2 induce CTD phosphorylation and transcription from HIV-1 promoter
HIV-1 tat interaction with RNA polymerase IIC-terminal domain (CTD) and a dynamic association with CDK2 induce CTD phosphorylation and transcription from HIV-1 promoter
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DOI：
10.1074/jbc.m111349200
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发表时间：
2002-09-13
影响因子：
4.8
通讯作者：
Nekhai, S
中科院分区：
文献类型：
--
作者：
Deng, LW;Ammosova, T;Nekhai, S
人类免疫缺陷病毒I型（HIV-1）的达特蛋白通过RNA聚合酶II（RNAPII）促进转录延伸来激活病毒基因表达。在该过程中，达特通过激活与启动子复合物的一般转录因子（特别是CDK 7和CDK 9）相关的细胞周期依赖性激酶（CDK）来增强RNAPII的C末端结构域（CTD）的磷酸化。我们报道了一种与Tat相关的T细胞衍生激酶，它含有CDK 2。在这里，我们提供了进一步的证据表明，CDK 2参与Tat介导的CTD磷酸化和HIV-1的体外转录。Tat介导的CTD通过CDK 2磷酸化需要达特活化结构域中的半胱氨酸22和达特的氨基酸42-72。CDK 2磷酸化达特本身，显然是通过与达特的氨基酸15-24和36-49形成动态接触。此外，达特的氨基酸24-36和45-72与CTD相互作用。CDK 2与RNAPII相关，并在HIV-1长末端重复模板上组装的延伸复合物中发现。重组CDK 2/cyclin E刺激Tat依赖的HIV-1转录HeLa细胞核提取物中CDK 2/细胞周期蛋白E的免疫耗竭阻断了Tat依赖性转录。我们认为CDK 2是转录复合物的一部分，该复合物是Tat依赖性转录所需的，并且达特与CTD的相互作用以及达特与CDK 2/细胞周期蛋白E的动态关联通过CDK 2刺激CTD磷酸化。
Human immunodeficiency virus, type I (HIV-1), Tat protein activates viral gene expression through promoting transcriptional elongation by RNA polymerase II (RNAPII). In this process Tat enhances phosphorylation of the C-terminal domain (CTD) of RNAPII by activating cell cycle-dependent kinases (CDKs) associated with general transcription factors of the promoter complex, specifically CDK7 and CDK9. We reported a Tat-associated T-cell-derived kinase, which contained CDK2. Here, we provide further evidence that CDK2 is involved in Tat-mediated CTD phosphorylation and in HIV-1 transcription in vitro. Tat-mediated CTD phosphorylation by CDK2 required cysteine 22 in the activation domain of Tat and amino acids 42-72 of Tat. CDK2 phosphorylated Tat itself, apparently by forming dynamic contacts with amino acids 15-24 and 36-49 of Tat. Also, amino acids 24-36 and 45-72 of Tat interacted with CTD. CDK2 associated with RNAPII and was found in elongation complexes assembled on HIV-1 long-terminal repeat template. Recombinant CDK2/cyclin E stimulated Tat-dependent HIV-1 transcription in reconstituted transcription assay. Immunodepletion of CDK2/cyclin E in HeLa nuclear extract blocked Tat-dependent transcription. We suggest that CDK2 is part of a transcription complex that is required for Tat-dependent transcription and that interaction of Tat with CTD and a dynamic association of Tat with CDK2/cyclin E stimulated CTD phosphorylation by CDK2.