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Query: UMLS:C0021051 (immunodeficiency)
 71,517 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Phosphorylation of the carboxyl-terminal domain (CTD) of the largest subunit of RNA polymerase II has been implicated as an important step in transcriptional regulation. Previously, we reported that a cellular CTD kinase, TAK, is targeted by the human immunodeficiency virus transactivator Tat. In the present study, we analyzed several other transactivators for the ability to interact with CTD kinases in vitro. The adenovirus E1A and herpes simplex virus VP16 proteins, but not other transactivators tested, were found to associate with a cellular kinase activity that hyperphosphorylates the CTD. The interaction is dependent upon a functional activation domain of E1A or VP16, suggesting that the interaction with a CTD kinase is relevant for the transactivation function of these proteins. The CTD kinase activities that interact with E1A and VP16 are related to each other but distinct from TAK. The Tat-, E1A- and VP16-associated CTD kinase activities detected in our assay also appear unrelated to MO15, the catalytic component of the CTD kinase activity of the general transcription factor TFIIH. Thus, this study has identified a novel interaction between viral transactivators and a cellular CTD kinase and suggests that at least two CTD kinases may mediate responses to viral transactivators.
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PMID:Viral transactivators specifically target distinct cellular protein kinases that phosphorylate the RNA polymerase II C-terminal domain. 860 64

We have studied the abilities of different transactivation domains to stimulate the initiation and elongation (postinitiation) steps of RNA polymerase II transcription in vivo. Nuclear run-on and RNase protection analyses revealed three classes of activation domains: Sp1 and CTF stimulated initiation (type I); human immunodeficiency virus type 1 Tat fused to a DNA binding domain stimulated predominantly elongation (type IIA); and VP16, p53, and E2F1 stimulated both initiation and elongation (type IIB). A quadruple point mutation of VP16 converted it from a type IIB to a type I activator. Type I and type IIA activators synergized with one another but not with type IIB activators. This observation implies that synergy can result from the concerted action of factors stimulating two different steps in transcription: initiation and elongation. The functional differences between activators may be explained by the different contacts they make with general transcription factors. In support of this idea, we found a correlation between the abilities of activators, including Tat, to stimulate elongation and their abilities to bind TFIIH.
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PMID:Three functional classes of transcriptional activation domain. 862 70

We investigated the role of TFIIH in transcription by RNA polymerase II (pol II) in vivo by microinjection of antibodies against this factor into Xenopus oocytes. Five different antibodies directed against four subunits of TFIIH were tested for effects on transcription of coinjected human immunodeficiency virus type 2 and c-myc templates. Each of these antibodies severely reduced the efficiency of elongation through human immunodeficiency virus type 2 and c-myc terminator elements. In contrast, an anti-TFIIB antibody did not inhibit elongation. Anti-TFIIH antibodies also had a much smaller inhibitory effect on total transcription than did anti-TFIIB or anti-pol II large subunit. Three inhibitors of TFIIH kinase activity, H-7, H-8, and dichlororibofuranosylbenzimidazole (DRB), inhibited elongation similarly to anti-TFIIH antibodies. These results strongly suggest a role for TFIIH in the stimulation of transcriptional elongation in vivo.
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PMID:TFIIH functions in regulating transcriptional elongation by RNA polymerase II in Xenopus oocytes. 866 44

The Tat protein is a transcriptional activator which is required for efficient human immunodeficiency virus 1 (HIV-1) gene expression Tat stimulates HIV-1 transcriptional elongation by increasing the processivity of RNA polymerase II. To address whether Tat-mediated effects on HIV-1 gene expression are due to modulation in the phosphorylation of the RNA polymerase II C-terminal domain (CTD), we developed a purification protocol to identify cellular kinases that are capable of binding to Tat and hyperphosphorylating the RNA polymerase II CTD. A 600 kDa protein complex with these properties was isolated, and specific components were identified using peptide microsequence analysis. This analysis indicated that proteins comprising the multi-subunit TFIIH complex, in addition to several novel factors, were associated with Tat using both in vitro and in vivo analysis. The Tat-associated kinase bound to the activation domain of Tat, and its ability to hyperphosphorylate RNA polymerase II was markedly stimulated by Tat. Furthermore, the addition of the Tat-associated kinase to in vitro transcription assays stimulated the ability of Tat to activate HIV-1 transcription. These results define a cellular kinase complex whose activity is modulated by Tat to result in activation of HIV-1 trancription.
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PMID:Purification of a Tat-associated kinase reveals a TFIIH complex that modulates HIV-1 transcription. 918 28

The human immunodeficiency virus encodes the transcriptional transactivator Tat, which binds to the transactivation response (TAR) RNA stem-loop in the viral long terminal repeat (LTR) and increases rates of elongation rather than initiation of transcription by RNA polymerase II (Pol II). In this study, we demonstrate that Tat binds directly to the cyclin-dependent kinase 7 (CDK7), which leads to productive interactions between Tat and the CDK-activating kinase (CAK) complex and between Tat and TFIIH. Tat activates the phosphorylation of the carboxy-terminal domain (CTD) of Pol II by CAK in vitro. The ability of CAK to phosphorylate the CTD can be inhibited specifically by a CDK7 pseudosubstrate peptide that also inhibits transcriptional activation by Tat in vitro and in vivo. We conclude that the phosphorylation of the CTD by CAK is essential for Tat transactivation. Our data identify a cellular protein that interacts with the activation domain of Tat, demonstrate that this interaction is critical for the function of Tat, and provide a mechanism by which Tat increases the processivity of Pol II.
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PMID:The HIV transactivator TAT binds to the CDK-activating kinase and activates the phosphorylation of the carboxy-terminal domain of RNA polymerase II. 933 27

Most of the genes involved in the pathogenesis of the DNA replication and repair syndromes have now been cloned, and our understanding of the basis for the pleiotropic phenotype associated with many of these syndromes has rapidly and dramatically expanded. The elucidation of the specific interactions between proteins that comprise the transcription factor complex TFIIH raises the possibility that nucleotide excision repair, RNA polymerase II transcription, and cell cycle control are connected. Defects in the XPB, XPD, and XPG genes can result in three different syndromes, xeroderma pigmentosum, Cockayne syndrome, or trichothiodystrophy, depending on the specific mutation involved. The recent cloning of the genes involved in Bloom syndrome (BLM) and Werner syndrome (WRN) show that both are DNA and RNA helicases with homology to each other and to other DExH box helicases, yet the mechanism by which defects in these genes cause such different phenotypes is not yet understood. The ataxia-telangiectasia gene (ATM) is involved in a variety of signal transduction pathways that regulate the cellular response to normal proliferative stimuli as well as the response to DNA damage, and the disruption of these signal transduction pathways provides an explanation for ataxia-telangiectasia characteristics such as ionizing radiation sensitivity, immunodeficiency, and infertility. Although the first Fanconi anemia gene (FAC) was cloned over 5 years ago, and a second Fanconi anemia gene (FAA) was cloned in 1996, the biochemical function of Fanconi anemia proteins largely remains a mystery. The recent construction of mutant mouse strains for several of these diseases should help unlock the difficult puzzle of the pathogenesis of these syndromes.
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PMID:Disorders of DNA replication and repair. 942 94

Human immunodeficiency virus, type 1 (HIV-1) Tat protein activates transcription from the HIV-1 long terminal repeat. Tat interacts with TFIIH and Tat-associated kinase (a transcription elongation factor P-TEFb) and requires the carboxyl-terminal domain of the largest subunit of RNA polymerase II (pol II) for transactivation. We developed a stepwise RNA pol II walking approach and used Western blotting to determine the role of TFIIH and P-TEFb in HIV-1 transcription elongation. Our results demonstrate the new findings that P-TEFb is a component of the preinitiation complex and travels with the elongating RNA pol II, whereas TFIIH is released from the elongation complexes before the trans-activation responsive region RNA is synthesized. Our results suggest that TFIIH and P-TEFb are involved in the clearance of promoter-proximal pausing of RNA pol II on the HIV-1 long terminal repeat at different stages.
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PMID:Tat-associated kinase (P-TEFb): a component of transcription preinitiation and elongation complexes. 1006 4

Tat stimulates human immunodeficiency virus type 1 (HIV-1) transcriptional elongation by recruitment of the human transcription elongation factor P-TEFb, consisting of Cdk9 and cyclin T1, to the HIV-1 promoter via cooperative binding to the nascent HIV-1 transactivation response RNA element. The Cdk9 kinase activity has been shown to be essential for P-TEFb to hyperphosphorylate the carboxy-terminal domain (CTD) of RNA polymerase II and mediate Tat transactivation. Recent reports have shown that Tat can also interact with the multisubunit transcription factor TFIIH complex and increase the phosphorylation of CTD by the Cdk-activating kinase (CAK) complex associated with the core TFIIH. These observations have led to the proposal that TFIIH and P-TEFb may act sequentially and in a concerted manner to promote phosphorylation of CTD and increase polymerase processivity. Here, we show that under conditions in which a specific and efficient interaction between Tat and P-TEFb is observed, only a weak interaction between Tat and TFIIH that is independent of critical amino acid residues in the Tat transactivation domain can be detected. Furthermore, immunodepletion of CAK under high-salt conditions, which allow CAK to be dissociated from core-TFIIH, has no effect on either basal HIV-1 transcription or Tat activation of polymerase elongation in vitro. Therefore, unlike the P-TEFb kinase activity that is essential for Tat activation of HIV-1 transcriptional elongation, the CAK kinase associated with TFIIH appears to be dispensable for Tat function.
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PMID:Tat activates human immunodeficiency virus type 1 transcriptional elongation independent of TFIIH kinase. 1008 52

This study describes a potential new function of hnRNP U as an RNA polymerase (Pol II) elongation inhibitor. We demonstrated that a subfraction of human hnRNP U is associated with the Pol II holoenzyme in vivo and as such recruited to the promoter as part of the preinitiation complex. hnRNP U, however, appears to dissociate from the Pol II complex at the early stage of transcription and is therefore absent from the elongating Pol II complex. When tested in the human immunodeficiency virus type 1 transcription system, hnRNP U inhibits elongation rather than initiation of transcription by Pol II. This inhibition requires the carboxy-terminal domain (CTD) of Pol II. We showed that hnRNP U can bind TFIIH in vivo under certain conditions and inhibit TFIIH-mediated CTD phosphorylation in vitro. We find that the middle domain of hnRNP U is sufficient to mediate its Pol II association and its inhibition of TFIIH-mediated phosphorylation and Pol II elongation. The abilities of hnRNP U to inhibit TFIIH-mediated CTD phosphorylation and its Pol II association are necessary for hnRNP U to mediate the repression of Pol II elongation. Based on these observations, we suggest that a subfraction of hnRNP U, as a component of the Pol II holoenzyme, may downregulate TFIIH-mediated CTD phosphorylation in the basal transcription machinery and repress Pol II elongation. With such functions, hnRNP U might provide one of the mechanisms by which the CTD is maintained in an unphosphorylated state in the Pol II holoenzyme.
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PMID:hnRNP U inhibits carboxy-terminal domain phosphorylation by TFIIH and represses RNA polymerase II elongation. 1049 Jun 22

Tat stimulates human immunodeficiency virus type 1 (HIV-1) transcriptional elongation by recruitment of carboxyl-terminal domain (CTD) kinases to the HIV-1 promoter. Using an immobilized DNA template assay, we have analyzed the effect of Tat on kinase activity during the initiation and elongation phases of HIV-1 transcription. Our results demonstrate that cyclin-dependent kinase 7 (CDK7) (TFIIH) and CDK9 (P-TEFb) both associate with the HIV-1 preinitiation complex. Hyperphosphorylation of the RNA polymerase II (RNAP II) CTD in the HIV-1 preinitiation complex, in the absence of Tat, takes place at CTD serine 2 and serine 5. Analysis of preinitiation complexes formed in immunodepleted extracts suggests that CDK9 phosphorylates serine 2, while CDK7 phosphorylates serine 5. Remarkably, in the presence of Tat, the substrate specificity of CDK9 is altered, such that the kinase phosphorylates both serine 2 and serine 5. Tat-induced CTD phosphorylation by CDK9 is strongly inhibited by low concentrations of 5, 6-dichloro-1-beta-D-ribofuranosylbenzimidazole, an inhibitor of transcription elongation by RNAP II. Analysis of stalled transcription elongation complexes demonstrates that CDK7 is released from the transcription complex between positions +14 and +36, prior to the synthesis of transactivation response (TAR) RNA. In contrast, CDK9 stays associated with the complex through +79. Analysis of CTD phosphorylation indicates a biphasic modification pattern, one in the preinitiation complex and the other between +36 and +79. The second phase of CTD phosphorylation is Tat-dependent and TAR-dependent. These studies suggest that the ability of Tat to increase transcriptional elongation may be due to its ability to modify the substrate specificity of the CDK9 complex.
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PMID:Tat modifies the activity of CDK9 to phosphorylate serine 5 of the RNA polymerase II carboxyl-terminal domain during human immunodeficiency virus type 1 transcription. 1086 64


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