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Query: UNIPROT:P23193 (transcription elongation factor)
 739 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human T-lymphotropic virus type 1 (HTLV-1) encodes a transcriptional activator, Tax, whose function is essential for viral transcription and replication. Tax transactivates the viral long-terminal repeat through a series of protein-protein interactions which facilitate CREB and CBP/p300 binding. In addition, Tax dissociates transcription repressor histone deacetylase 1 interaction with the CREB response element. The subsequent events through which Tax interacts and communicates with RNA polymerase II and cyclin-dependent kinases (CDKs) are not clearly understood. Here we present evidence that Tax recruits positive transcription elongation factor b (P-TEFb) (CDK9/cyclin T1) to the viral promoter. This recruitment likely involves protein-protein interactions since Tax associates with P-TEFb in vitro as demonstrated by glutathione S-transferase fusion protein pull-down assays and in vivo as shown by co-immunoprecipitation assays. Functionally, small interfering RNA directed toward CDK9 inhibited Tax transactivation in transient assays. Consistent with these findings, the depletion of CDK9 from nuclear extracts inhibited Tax transactivation in vitro. Reconstitution of the reaction with wild-type P-TEFb, but not a kinase-dead mutant, recovered HTLV-1 transcription. Moreover, the addition of the CDK9 inhibitor flavopiridol blocked Tax transactivation in vitro and in vivo. Interestingly, we found that Tax regulates CDK9 kinase activity through a novel autophosphorylation pathway.
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PMID:Tax interacts with P-TEFb in a novel manner to stimulate human T-lymphotropic virus type 1 transcription. 1664 Dec 71

The formation of the Tat-protein/TAR-RNA complex is a crucial step in the regulation of human immunodeficiency virus (HIV)-gene expression. To obtain full-length viral transcripts the Tat/TAR complex has to recruit the positive transcription elongation factor complex (P-EFTb), which interacts with TAR through its cyclin T1 (CycT1) component. Mutational studies identified the TAR hexanucleotide loop as a crucial region for contacting CycT1. Interfering with the interaction between the Tat/CycT1 complex and the TAR-RNA is an attractive strategy for the design of anti-HIV drugs. Positively charged molecules, like aminoglycosides or peptidomimetics, bind the TAR-RNA, disrupting the Tat/TAR complex. Here, we investigate the complex between the HIV-2 TAR-RNA and a neooligoaminodeoxysaccharide by NMR spectroscopy. In contrast to other aminoglycosides, this novel aminoglycoside analogue contacts simultaneously the bulge residues required for Tat binding and the A35 residue of the hexanucleotide loop. Upon complex formation, the loop region undergoes profound conformational changes. The novel binding mode, together with the easy accessibility of derivatives for the neooligoaminodeoxysaccharide, could open the way to the design of a new class of TAR-RNA binders, which simultaneously inhibit the formation of both the Tat/TAR binary complex and the Tat/TAR/CycT1 ternary complex by obstructing both the bulge and loop regions of the RNA.
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PMID:TAR-RNA recognition by a novel cyclic aminoglycoside analogue. 1685 96

Cyclin K and the closely related cyclins T1, T2a, and T2b interact with cyclin-dependent kinase 9 (CDK9) forming multiple nuclear complexes, referred to collectively as positive transcription elongation factor b (P-TEFb). Through phosphorylation of the C-terminal domain of the RNA polymerase II largest subunit, distinct P-TEFb species regulate the transcriptional elongation of specific genes that play central roles in human physiology and disease development, including cardiac hypertrophy and human immunodeficiency virus-1 pathogenesis. We have determined the crystal structure of human cyclin K (residues 11-267) at 1.5 A resolution, which represents the first atomic structure of a P-TEFb subunit. The cyclin K fold comprises two typical cyclin boxes with two short helices preceding the N-terminal box. A prominent feature of cyclin K is an additional helix (H4a) in the first cyclin box that obstructs the binding pocket for the cell-cycle inhibitor p27(Kip1). Modeling of CDK9 bound to cyclin K provides insights into the structural determinants underlying the formation and regulation of this complex. A homology model of human cyclin T1 generated using the cyclin K structure as a template reveals that the two proteins have similar structures, as expected from their high level of sequence identity. Nevertheless, their CDK9-interacting surfaces display significant structural differences, which could potentially be exploited for the design of cyclin-targeted inhibitors of the CDK9-cyclin K and CDK9-cyclin T1 complexes.
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PMID:Crystal structure of human cyclin K, a positive regulator of cyclin-dependent kinase 9. 1716 70

Transcriptional regulation of the human immunodeficiency virus type 1 (HIV-1) is a complex event that requires the cooperative action of both viral (e.g. Tat) and cellular (e.g. C/EBPbeta, NF-kappaB) factors. The HIV-1 Tat protein recruits the human positive transcription elongation factor P-TEFb, consisting of cdk9 and cyclin T1, to the HIV-1 transactivation response (TAR) region. In the absence of TAR, Tat activates the HIV-1 long terminal repeat (LTR) through its association with several cellular factors including C/EBPbeta. C/EBPbeta is a member of the CCAAT/enhancer-binding protein family of transcription factors and has been shown to be a critical transcriptional regulator of HIV-1 LTR. We examined whether Tat-C/EBPbeta association requires the presence of the P-TEFb complex. Using immunoprecipitation followed by Western blot, we demonstrated that C/EBPbeta-cyclin T1 association requires the presence of cdk9. Further, due to its instability, cdk9 was unable to physically interact with C/EBPbeta in the absence of cyclin T1 or Tat. Using kinase assays, we demonstrated that cdk9, but not a cdk9 dominant-negative mutant (cdk9-dn), phosphorylates C/EBPbeta. Our functional data show that co-transfection of C/EBPbeta and cdk9 leads to an increase in HIV-1 gene expression when compared to C/EBPbeta alone. Addition of C/EBP homologous protein (CHOP) inhibits C/EBPbeta transcriptional activity in the presence and absence of cdk9 and causes a delay in HIV-1 replication in T-cells. Together, our data suggest that Tat-C/EBPbeta association is mediated through cdk9, and that phosphorylated C/EBPbeta may influence AIDS progression by increasing expression of HIV-1 genes.
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PMID:C/EBPbeta regulates human immunodeficiency virus 1 gene expression through its association with cdk9. 1725 82

Positive transcription elongation factor b (P-TEFb) complexes, composed of cyclin-dependent kinase 9 (CDK9) and cyclin T1 or T2, are engaged by many cellular transcription regulators that activate or inhibit transcription from specific promoters. The related I-mfa (inhibitor of MyoD family a) and HIC (human I-mfa-domain-containing) proteins function in myogenic differentiation and embryonic development by participating in the Wnt signaling pathway. We report that I-mfa is a novel regulator of P-TEFb. Both HIC and I-mfa interact through their homologous I-mfa domains with cyclin T1 and T2 at two binding sites. One site is the regulatory histidine-rich domain that interacts with CDK9 substrates including RNA polymerase II. The second site contains a lysine and arginine-rich motif that is highly conserved between the two T cyclins. This site overlaps and includes the previously identified Tat/TAR recognition motif of cyclin T1 required for activation of human immunodeficiency virus type 1 (HIV-1) transcription. HIC and I-mfa can serve as substrates for P-TEFb. Their I-mfa domains also bind the activation domain of HIV-1 Tat and inhibit Tat- and P-TEFb-dependent transcription from the HIV-1 promoter. This transcriptional repression is cell-type specific and can operate via Tat and cyclin T1. Genomic and sequence comparisons indicate that the I-mf and HIC genes, as well as flanking genes, diverged from a duplicated chromosomal region. Our findings link I-mfa and HIC to viral replication, and suggest that P-TEFb is modulated in the Wnt signaling pathway.
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PMID:Developmental regulators containing the I-mfa domain interact with T cyclins and Tat and modulate transcription. 1728 77

Human immunodeficiency virus type 1 (HIV-1) transcriptional transactivator (Tat) recruits the positive transcription elongation factor b (P-TEFb) to the viral promoter. Consisting of cyclin dependent kinase 9 (Cdk9) and cyclin T1, P-TEFb phosphorylates RNA polymerase II and the negative transcription elongation factor to stimulate the elongation of HIV-1 genes. A major fraction of nuclear P-TEFb is sequestered into a transcriptionally inactive 7SK small nuclear ribonucleoprotein (snRNP) by the coordinated actions of the 7SK small nuclear RNA (snRNA) and hexamethylene bisacetamide (HMBA) induced protein 1 (HEXIM1). In this study, we demonstrate that Tat prevents the formation of and also releases P-TEFb from the 7SK snRNP in vitro and in vivo. This ability of Tat depends on the integrity of its N-terminal activation domain and stems from the high affinity interaction between Tat and cyclin T1, which allows Tat to directly displace HEXIM1 from cyclin T1. Furthermore, we find that in contrast to the Tat-independent activation of the HIV-1 promoter, Tat-dependent HIV-1 transcription is largely insensitive to the inhibition by HEXIM1. Finally, primary blood lymphocytes display a reduced amount of the endogenous 7SK snRNP upon HIV-1 infection. All these data are consistent with the model that Tat not only recruits but also increases the active pool of P-TEFb for efficient HIV-1 transcription.
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PMID:Tat competes with HEXIM1 to increase the active pool of P-TEFb for HIV-1 transcription. 1734 62

The C-terminal domain (CTD) of RNA polymerase II is phosphorylated during the transcription cycle by three cyclin-dependent kinases (CDKs): CDK7, CDK8, and CDK9. CDK9 and its interacting cyclin T partners belong to the positive transcription elongation factor b (P-TEFb) complexes, which phosphorylate the CTD to promote transcription elongation. We report that Arabidopsis thaliana CDK9-like proteins, CDKC;1 and CDKC;2, and their interacting cyclin T partners, CYCT1;4 and CYCT1;5, play important roles in infection with Cauliflower mosaic virus (CaMV). cdkc;2 and cyct1;5 knockout mutants are highly resistant and cdkc;2 cyct1;5 double mutants are extremely resistant to CaMV. The mutants respond normally to other types of plant viruses that do not replicate by reverse transcription. Expression of a reporter gene driven by the CaMV 35S promoter is markedly reduced in the cdkc;2 and cyct1;5 mutants, indicating that the kinase complexes are important for transcription from the viral promoter. Loss of function of CDKC;1/CDKC;2 or CYCT1;4/CYCT1;5 results in complete resistance to CaMV as well as altered leaf and flower growth, trichome development, and delayed flowering. These results establish Arabidopsis CDKC kinase complexes as important host targets of CaMV for transcriptional activation of viral genes and critical regulators of plant growth and development.
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PMID:Roles of Arabidopsis cyclin-dependent kinase C complexes in cauliflower mosaic virus infection, plant growth, and development. 1746 59

The positive transcription elongation factor b (P-TEFb) is an essential regulator of viral gene expression during the life cycle of human immunodeficiency virus type 1 (HIV-1). Its cyclin T1 subunit forms a ternary complex with the viral transcriptional transactivator (Tat) protein and the transactivation response (TAR) RNA element thereby activating cyclin dependent kinase 9 (Cdk9), which stimulates transcription at the level of chain elongation. We report the structure of the cyclin box domain of human cyclin T1 at a resolution of 2.67 A. The structure was obtained by crystallographic analysis of a fusion protein composed of cyclin T1 linked to the transactivator protein Tat from equine infectious anemia virus (EIAV), which is functionally and structurally related to HIV-1 Tat. The conserved cyclin box domain of cyclin T1 exhibits structural features for interaction with physiological binding partners such as Cdk9. A recognition site for Cdk/Cyclin substrates is partly covered by a cyclin T-specific insert, suggesting specific interactions with regulatory factors. The previously identified Tat/TAR recognition motif (TRM) forms a C-terminal helix that is partly occluded in the cyclin box repeat interface, while cysteine 261 is accessible to form an intermolecular zinc finger with Tat. Residues of the TRM contribute to a positively charged groove that may directly attract RNA molecules. The EIAV Tat protein instead appeared undefined from the electron density map suggesting that it is highly disordered. Functional experiments confirmed the TAR binding properties of the fusion protein and suggested residues on the second cyclin box repeat to contribute to Tat stimulated transcription.
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PMID:Cyclin box structure of the P-TEFb subunit cyclin T1 derived from a fusion complex with EIAV tat. 1754 Apr 6

Positive transcription elongation factor (P-TEFb), which is composed of CDK9 and cyclin T1, plays an important role in cellular and viral gene expression. Our lab has recently demonstrated that P-TEFb is required for Tax transactivation of the viral long terminal repeat (LTR). P-TEFb is found in two major complexes: the inactive form, which is associated with inhibitory subunits 7SK snRNA and HEXIM1, and the active form, which is associated with, at least in part, Brd4. In this study, we analyzed the effect of Brd4 on human T-lymphotropic virus type 1 (HTLV-1) transcription. Overexpression of Brd4 repressed Tax transactivation of the HTLV-1 LTR in a dose-dependent manner. In vitro binding studies suggest that Tax and Brd4 compete for binding to P-TEFb through direct interaction with cyclin T1. Tax interacts with cyclin T1 amino acids 426 to 533, which overlaps the region responsible for Brd4 binding. In vivo, overexpression of Tax decreased the amount of 7SK snRNA associated with P-TEFb and stimulates serine 2 phosphorylation of the RNA polymerase II carboxyl-terminal domain, suggesting that Tax regulates the functionality of P-TEFb. Our results suggest the possibility that Tax may compete and functionally substitute for Brd4 in P-TEFb regulation.
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PMID:Modulation of the Brd4/P-TEFb interaction by the human T-lymphotropic virus type 1 tax protein. 1768 63

HIV-1 transcription is essential for the virus replication cycle. HIV-1 Tat is a viral transactivator that strongly stimulates the processivity of RNA polymerase II (RNAPII) via recruitment of the cyclin T1/CDK9 positive transcription elongation factor, which phosphorylates the C-terminal domain (CTD) of RNAPII. Consistently, HIV-1 replication in transformed cells is very sensitive to direct CDK9 inhibition. Thus, CDK9 could be a potential target for anti-HIV-1 therapy. A clearer understanding of the requirements for CDK9 activity in primary human T cells is needed to assess whether the CDK9-dependent step in HIV-1 transcription can be targeted clinically. We have investigated the effects of limiting CDK9 activity with recombinant lentiviruses expressing a dominant-negative form of CDK9 (HA-dnCDK9) in peripheral blood lymphocytes (PBLs) and other cells. Our results show that direct inhibition of CDK9 potently inhibits HIV-1 replication in single-round infection assays with little to undetectable effects on RNAPII transcription, RNA synthesis, proliferation and viability. In PBLs purified from multiple donors, direct inhibition of CDK9 activity blocks HIV-1 replication/transcription but does not prevent T-cell activation, as determined via measurement of cell surface and cell cycle entry and progression markers, and DNA synthesis. We have also compared the effects of HA-dnCDK9 to flavopiridol (FVP), a general CDK inhibitor that potently inhibits CDK9. In contrast to HA-dnCDK9, FVP interferes with key cellular processes at concentrations that inhibit HIV-1 replication with potency similar to HA-dnCDK9. In particular, FVP inhibits several T-cell activation markers and DNA synthesis in primary PBLs at the minimal concentrations required to inhibit HIV-1 replication. Our results imply that small pharmacological compounds targeting CDK9 with enhanced selectivity could be developed into effective anti-HIV-1 therapeutic drugs.
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PMID:Direct inhibition of CDK9 blocks HIV-1 replication without preventing T-cell activation in primary human peripheral blood lymphocytes. 1794 27


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