Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019693 (HIV)
 170,526 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Tat protein mediates transactivation of human immunodeficiency virus type 1 (HIV-1), which results in more-efficient transcript elongation. Since phosphorylation of C-terminal domain (CTD) of RNA polymerase II correlates with its enhanced processivity, we studied the properties of a Tat-associated CTD kinase derived from mitogenically stimulated human primary T lymphocytes (TTK). TTK binds to full-length Tat and specifically phosphorylates CTD and CDK2. This dual kinase activity is characteristic of CDK-activating kinase (CAK). The CTD kinase activity is induced upon mitogenic stimulation of primary T lymphocytes. Fractionation of T-cell lysate demonstrates that Tat-associated CTD kinase activity elutes in two peaks. About 60% of Tat-associated CTD kinase copurifies with CDK2 kinase activity and contains the CAK components CDK7 and cyclin H. The rest of Tat-associated kinase is free of CDK2 kinase activity and the CAK components and thus may represent a novel CTD kinase. The kinase activities of TTK are blocked by the adenosine analog 5,6-dichloro-1-beta-D-ribofuranosyl-benzimidazole (DRB) as well as by the kinase inhibitor H8 at concentrations known to block transcript elongation. Importantly, the Tat-associated kinase markedly induced CAK. We suggest that the mechanism of Tat-mediated processive transcription of the HIV-1 promoter includes a Tat-associated CAK activator.
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PMID:A human primary T-lymphocyte-derived human immunodeficiency virus type 1 Tat-associated kinase phosphorylates the C-terminal domain of RNA polymerase II and induces CAK activity. 931 22

During transcription of mRNA genes, there is a correlation between the phosphorylation state of the C-terminal domain (CTD) of the large subunit of RNA polymerase II (RNAP II) and the ability of the RNAP II complex to processively transcribe the gene. To examine the involvement of CTD phosphorylation in modulation of RNAP II function, we have analyzed the ability of a known CTD kinase, human Cdk8, to modulate HIV-1 LTR-driven gene expression upon directed targeting to a promoter-proximal nascent RNA element. The results indicated that Cdk8, when localized to an RNA element, activates gene expression in a catalysis-dependent manner. Also, Cdk8 targeted to RNA was observed to act in a synergystic manner with DNA-targeted Sp1 but not with DNA-targeted HIV-1 Tat, suggesting that RNA-targeted Cdk8 acts on similar rate limiting post-initiation events as Tat. As recent observations suggest that Tat/TAR-mediated transcription of the proviral genome of HIV depends on specific phosphorylation of RNAP II in its CTD by the Tat-associated kinase (TAK/p-TEFb/Cdk9), our results indicate that Cdk8 shares with Cdk9 the ability to modulate transcription upon targeting to a nascent RNA element.
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PMID:Targeting of CDK8 to a promoter-proximal RNA element demonstrates catalysis-dependent activation of gene expression. 968 96

HIV-1 Tat activates transcription through binding to human cyclin T1, a regulatory subunit of the TAK/P-TEFb CTD kinase complex. Here we show that the cyclin domain of hCycT1 is necessary and sufficient to interact with Tat and promote cooperative binding to TAR RNA in vitro, as well as mediate Tat transactivation in vivo. A Tat:TAR recognition motif (TRM) was identified at the carboxy-terminal edge of the cyclin domain, and we show that hCycT1 can interact simultaneously with Tat and CDK9 on TAR RNA in vitro. Alanine-scanning mutagenesis of the hCycT1 TRM identified residues that are critical for the interaction with Tat and others that are required specifically for binding of the complex to TAR RNA. Interestingly, we find that the interaction between Tat and hCycT1 requires zinc as well as essential cysteine residues in both proteins. Cloning and characterization of the murine CycT1 protein revealed that it lacks a critical cysteine residue (C261) and forms a weak, zinc-independent complex with HIV-1 Tat that greatly reduces binding to TAR RNA. A point mutation in mCycT1 (Y261C) restores high-affinity, zinc-dependent binding to Tat and TAR in vitro, and rescues Tat transactivation in vivo. Although overexpression of hCycT1 in NIH3T3 cells strongly enhances transcription from an integrated proviral promoter, we find that this fails to overcome all blocks to productive HIV-1 infection in murine cells.
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PMID:The interaction between HIV-1 Tat and human cyclin T1 requires zinc and a critical cysteine residue that is not conserved in the murine CycT1 protein. 983 4

Cyclin T1 has been identified recently as a regulatory subunit of CDK9 and as a component of the transcription elongation factor P-TEFb. Cyclin T1/CDK9 complexes phosphorylate the carboxy terminal domain (CTD) of RNA polymerase II (RNAP II) in vitro. Here we report that the levels of cyclin T1 are dramatically upregulated by two independent signaling pathways triggered respectively by PMA and PHA in primary human peripheral blood lymphocytes (PBLs). Activation of these two pathways in tandem is sufficient for PBLs to enter and progress through the cell cycle. However, the expression of cyclin T1 is not growth and/or cell cycle regulated in other cell types, indicating that regulation of cyclin T1 expression is dependent on tissue-specific signaling pathways. Upregulation of cyclin T1 in stimulated PBLs results in induction of the CTD kinase activity of the cyclin T1/CDK9 complex, which in turn correlates directly with phosphorylation of RNAP II in vivo, linking for the first time activation of the cyclin T1/ CDK9 pair with phosphorylation of RNAP II in vivo. In addition, we report here that endogenous CDK9 and cyclin T1 complexes associate with HIV-1 generated Tat in relevant cells and under physiological conditions (HIV-1 infected T cells). This, together with our results showing that HIV-1 replication in stimulated PBLs correlates with the levels of cyclin T1 protein and associated CTD kinase activity, suggests that the cyclin T1/CDK9 pair is one of the HIV-1 required host cellular cofactors generated during T cell activation.
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PMID:Upregulation of cyclin T1/CDK9 complexes during T cell activation. 987 25

Actinomycin D and alpha-amanitin are commonly used to inhibit transcription. Unexpectedly, however, the transcription of the human immunodeficiency virus (HIV-1) long terminal repeats (LTR) is shown to be activated at the level of elongation, in human and murine cells exposed to these drugs, whereas the Rous sarcoma virus LTR, the human cytomegalovirus immediate early gene (CMV), and the HSP70 promoters are repressed. Activation of the HIV LTR is independent of the NFkappaB and TAR sequences and coincides with an enhanced average phosphorylation of the C-terminal domain (CTD) from the largest subunit of RNA polymerase II. Both the HIV-1 LTR activation and the bulk CTD phosphorylation enhancement are prevented by several CTD kinase inhibitors, including 5, 6-dichloro-1-beta-D-ribofuranosylbenzimidazole. The efficacies of the various compounds to block CTD phosphorylation and transcription in vivo correlate with their capacities to inhibit the CDK9/PITALRE kinase in vitro. Hence, the positive transcription elongation factor, P-TEFb, is likely to contribute to the average CTD phosphorylation in vivo and to the activation of the HIV-1 LTR induced by actinomycin D.
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PMID:The transcriptional inhibitors, actinomycin D and alpha-amanitin, activate the HIV-1 promoter and favor phosphorylation of the RNA polymerase II C-terminal domain. 1034 61

HIV-1 gene expression relies upon a complex machinery that is primarily controlled by two viral regulatory proteins, Tat and Rev. Rev is involved in regulating post-transcriptional events of HIV-1 gene expression. The Tat protein transactivates transcription from the HIV-1 5' long terminal repeat (LTR) and acts in synergy with specific cellular factors. Recently, it has been shown that one set of these cellular factors is a protein kinase activity termed TAK (Tat-associated kinase), which activates transcription by hyperphosphorylation of the carboxyl-terminal domain (CTD) of the large subunit of RNA polymerase II. TAK also enhances transcription of HIV-2, together with the retroviral transactivator, Tat-2. The TAK activity appears to be related to the CTD kinase P-TEFb, which stabilizes transcription elongation of many genes and was originally isolated from Drosophila extracts. Both TAK and P-TEFb contain at least two subunits: the cyclin-dependent kinase, CDK9 (PITALRE), the catalytic subunit, and the regulatory subunit, cyclin T1. CDK9 and cyclin T1 are ubiquitous factors that affects many cellular processes, including cell differentiation and apoptosis. The involvement of TAK in HIV-1 and HIV-2 gene expression is an important aspect in the biology of these two retroviruses, and may lead to the development of novel antiretroviral drugs and/or gene therapy approaches for the treatment of patients with AIDS.
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PMID:Regulatory functions of Cdk9 and of cyclin T1 in HIV tat transactivation pathway gene expression. 1053 59

Activation of the HIV-1 promoter by the virally encoded Tat protein is characterized by efficient processive transcription, mediated by host cell factors that are tethered to the promoter with the Tat-TAR RNA complex. Importantly, viral gene activation has been shown to be stimulated in mitogenically induced cells, although the link between cell cycle regulation and viral gene activation is unclear. We reported a Tat-associated CAK/CTD kinase from mitogenically induced primary human T-cells (TTK) (S. Nekhai et al., 1997, J. Virol. 71, 7436-7441). Here, biological activity of the kinase has been studied by direct microinjection at the individual-cell level. The TTK-dependent Tat response is maximal during G1 phase as shown by co-injection with Tat protein in cells synchronized at the various stages of the cell cycle. The cell cycle dependence of the Tat response was confirmed by inhibiting G0 --> G1 progression with the expression of dominant negative mutant Ras(Asn17) or the cyclin-dependent kinase CDK4. The results support a mechanism whereby transactivation of the HIV promoter is regulated by cell growth signal transduction pathways that target the Tat cofactor.
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PMID:Cell cycle-dependent stimulation of the HIV-1 promoter by Tat-associated CAK activator. 1063 11

HIV-1 Tat is able to form a ternary complex with P/CAF and p300 and increase the affinity for CDK9/P-TEFb CTD kinase complex. Our previous study demonstrated that Tat binds to p300/CBP in the minimal HAT domain (aa 1253-1790) and that the interaction results in a change of conformation on p300/CBP. Here, we show that the Tat-p300 interaction increases the HAT activity of p300 on histone H4 that is associated with nucleosomal DNA and not with free histones. Nucleosomal histone H4 was acetylated on lysines 8, 12, and 16. Acetylation of H4 was inhibited by Lys-coenzyme A (CoA), a selective inhibitor of p300 acetyltransferase activity. Unexpectedly, we also found that Tat could autoacetylate itself, which was specific to lysine residues 41 and 71. Peptides lacking these two lysines could not enhance the HAT activity of p300. Comparison of the sequences of Tat with other HIV-1 clades and HAT containing transcription factors indicated sequence identity in the acetyl-CoA binding motif A, KGXG. Furthermore, when utilizing an in vitro transcription assay, as well as a Tat mutant virus, we found that ectopic expression of only wild-type Tat in the presence of p300, and not a lysine 41 Tat mutant, could activate HIV-1 chromatin DNA, as evidenced by the absence of HIV-1 virion antigen. Therefore, transcription of integrated viral DNA in vivo requires the HAT activity of coactivators that are modulated by Tat to derepress the HIV-1 chromatin structure and aid in activated transcription.
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PMID:Enhancement of the p300 HAT activity by HIV-1 Tat on chromatin DNA. 1168 53

HIV-1 gene expression is regulated by a viral transactivator protein (Tat) which induces transcriptional elongation of HIV-1 long tandem repeat (LTR). This induction requires hyperphosphorylation of the C-terminal domain (CTD) repeats of RNA polymerase II (Pol II). To achieve CTD hyperphosphorylation, Tat stimulates CTD kinases associated with general transcription factors of the promoter complex, specifically TFIIH-associated CDK7 and positive transcription factor b-associated CDK9 (cyclin-dependent kinase 9). Other studies indicate that Tat may bind an additional CTD kinase that regulates the target-specific phosphorylation of RNA Pol II CTD. We previously reported that Tat-associated T-cell-derived kinase (TTK), purified from human primary T-cells, stimulates Tat-dependent transcription of HIV-1 LTR in vivo [Nekhai, Shukla, Fernandez, Kumar and Lamb (2000) Virology 266, 246-256]. In the work presented here, we characterized the components of TTK by biochemical fractionation and the function of TTK in transcription assays in vitro. TTK uniquely co-purified with CDK2 and not with either CDK9 or CDK7. Tat induced the TTK-associated CDK2 kinase to phosphorylate CTD, specifically at Ser-2 residues. The TTK fraction restored Tat-mediated transcription activation of HIV-1 LTR in a HeLa nuclear extract immunodepleted of CDK9, but not in the HeLa nuclear extract double-depleted of CDK9 and CDK7. Direct microinjection of the TTK fraction augmented Tat transactivation of HIV-1 LTR in human primary HS68 fibroblasts. The results argue that TTK-associated CDK2 may function to maintain target-specific phosphorylation of RNA Pol II that is essential for Tat transactivation of HIV-1 promoter. They are also consistent with the observed cell-cycle-specific induction of viral gene transactivation.
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PMID:HIV-1 Tat-associated RNA polymerase C-terminal domain kinase, CDK2, phosphorylates CDK7 and stimulates Tat-mediated transcription. 1204 28

Phosphorylation of RNA polymerase II carboxy-terminal domain (CTD) in hepta-repeats YSPTSPS regulates eukaryotic transcription. Whereas Ser5 is phosphorylated in the initiation phase, Ser2 phosphorylation marks the elongation state. Here we show that the positive transcription elongation factor P-TEFb is a Ser5 CTD kinase that is unable to create Ser2/Ser5 double phosphorylations, while it exhibits fourfold higher activity on a CTD substrate pre-phosphorylated at Ser7 compared with the consensus hepta-repeat or the YSPTSPK variant. Mass spectrometry reveals an equal number of phosphorylations to the number of hepta-repeats provided, yet the mechanism of phosphorylation is distributive despite the repetitive nature of the substrate. Inhibition of P-TEFb activity is mediated by two regions in Hexim1 that act synergistically on Cdk9 and Cyclin T1. HIV-1 Tat/TAR abrogates Hexim1 inhibition to stimulate transcription of viral genes but does not change the substrate specificity. Together, these results provide insight into the multifaceted pattern of CTD phosphorylation.
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PMID:Serine-7 but not serine-5 phosphorylation primes RNA polymerase II CTD for P-TEFb recognition. 2258 4


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