Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P23193 (transcription elongation factor)
 739 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The human immunodeficiency virus type 1 (HIV-1) Tat protein strongly and specifically stimulates transcription elongation from the HIV-1 LTR and provides an important in vitro model system to study this process. Here we use protein-affinity chromatography to identify cellular factors involved in transcription elongation. A Tat-affinity column bound one transcription factor, Tat-SF1, efficiently and selectively. Tat-SF1 was identified originally as a Tat-specific coactivator, but we show it is a general transcription elongation factor. Our results also reveal the existence of an ATP-inactivatable general elongation factor (AIEF) required for Tat-SF1 activity and for which Tat can substitute functionally.
 ...
PMID:The HIV-1 Tat cellular coactivator Tat-SF1 is a general transcription elongation factor. 976 1

Prior phenotypic analysis of a vaccinia virus gene A18R mutant, Cts23, showed the synthesis of longer than wild type (Wt) length viral transcripts during the intermediate stage of infection, indicating that the A18R protein may act as a negative transcription elongation factor. The purpose of the work described here was to determine a biochemical activity for the A18R protein. Pulse-labeled transcription complexes established from intermediate virus promoters on bead-bound DNA templates were assayed for transcript release during an elongation step that contained nucleotides and various proteins. Pulse-labeled transcription complexes elongated in the presence of only nucleotides were unable to release nascent RNA. The addition of Wt extract during the elongation phase resulted in release of the nascent transcript, indicating that additional factors present in the Wt extract are capable of inducing transcript release. Extract from Cts23 or mock-infected cells was unable to induce release. The lack of release upon addition of Cts23 extract suggests that A18R is involved in release of nascent RNA. By itself, purified polyhistidine-tagged A18R protein (His-A18R) was unable to induce release; however, release did occur in the presence of purified His-A18R protein plus extract from either Cts23 or mock-infected cells. These data taken together indicate that A18R is necessary but not sufficient for release of nascent transcripts. We have also demonstrated that the combination of A18R protein and mock extract induces transcript release in an ATP-dependent manner, consistent with the fact that the A18R protein is an ATP-dependent helicase. Further analysis revealed that the release activity is not restricted to a vaccinia intermediate promoter but is observed using pulse-labeled transcription complexes initiated from all three viral gene class promoters. Therefore, we conclude that A18R and an as yet unidentified cellular factor(s) are required for the in vitro release of nascent RNA from a vaccinia virus transcription elongation complex.
 ...
PMID:Vaccinia virus gene A18R DNA helicase is a transcript release factor. 1062 2

Human immunodeficiency virus type 1 (HIV-1) Tat interacts with cyclin T1 (CycT1), a regulatory partner of CDK9 in the positive transcription elongation factor (P-TEFb) complex, and binds cooperatively with CycT1 to TAR RNA to recruit P-TEFb and promote transcription elongation. We show here that Tat also stimulates phosphorylation of affinity-purified core RNA polymerase II and glutathione S-transferase-C-terminal-domain substrates by CycT1-CDK9, but not CycH-CDK7, in vitro. Interestingly, incubation of recombinant Tat-P-TEFb complexes with ATP enhanced binding to TAR RNA dramatically, and the C-terminal half of CycT1 masked binding of Tat to TAR RNA in the absence of ATP. ATP incubation lead to autophosphorylation of CDK9 at multiple C-terminal Ser and Thr residues, and full-length CycT1 (amino acids 728) [CycT1(1-728)], but not truncated CycT1(1-303), was also phosphorylated by CDK9. P-TEFb complexes containing a catalytically inactive CDK9 mutant (D167N) bound TAR RNA weakly and independently of ATP, as did a C-terminal truncated CDK9 mutant that was catalytically active but unable to undergo autophosphorylation. Analysis of different Tat proteins revealed that the 101-amino-acid SF2 HIV-1 Tat was unable to bind TAR with CycT1(1-303) in the absence of phosphorylated CDK9, whereas unphosphorylated CDK9 strongly blocked binding of HIV-2 Tat to TAR RNA in a manner that was reversed upon autophosphorylation. Replacement of CDK9 phosphorylation sites with negatively charged residues restored binding of CycT1(1-303)-D167N-Tat, and rendered D167N a more potent inhibitor of transcription in vitro. Taken together, these results demonstrate that CDK9 phosphorylation is required for high-affinity binding of Tat-P-TEFb to TAR RNA and that the state of P-TEFb phosphorylation may regulate Tat transactivation in vivo.
 ...
PMID:CDK9 autophosphorylation regulates high-affinity binding of the human immunodeficiency virus type 1 tat-P-TEFb complex to TAR RNA. 1095 91

Flavopiridol (L86-8275, HMR1275) is a cyclin-dependent kinase (Cdk) inhibitor in clinical trials as a cancer therapy that has been recently shown to block human immunodeficiency virus Tat transactivation and viral replication through inhibition of positive transcription elongation factor b (P-TEFb). Flavopiridol is the most potent P-TEFb inhibitor reported and the first Cdk inhibitor that is not competitive with ATP. We examined the ability of flavopiridol to inhibit P-TEFb (Cdk9/cyclin T1) phosphorylation of both RNA polymerase II and the large subunit of the 5, 6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) sensitivity-inducing factor and found that the IC(50) determined was directly related to the concentration of the enzyme. We concluded that the flavonoid associates with P-TEFb with 1:1 stoichiometry even at concentrations of enzyme in the low nanomolar range. These results indicate that the apparent lack of competition with ATP could be caused by a very tight binding of the drug. We developed a novel immobilized P-TEFb assay and demonstrated that the drug remains bound for minutes even in the presence of high salt. Flavopiridol remained bound in the presence of a 1000-fold excess of the commonly used inhibitor DRB, suggesting that the immobilized P-TEFb could be used in a simple screening assay that would allow the discovery or characterization of compounds with binding properties similar to flavopiridol. Finally, we compared the ability of flavopiridol and DRB to inhibit transcription in vivo using nuclear run-on assays and concluded that P-TEFb is required for transcription of most RNA polymerase II molecules in vivo.
 ...
PMID:Flavopiridol inactivates P-TEFb and blocks most RNA polymerase II transcription in vivo. 1143 68

Schizosaccharomyces pombe Cdk9/Pch1 protein kinase is a functional ortholog of the essential Saccharomyces cerevisiae Bur1/Bur2 kinase and a putative ortholog of metazoan P-TEFb (Cdk9/cyclin T). SpCdk9/Pch1 phosphorylates of the carboxyl-terminal domain (CTD) of the S. pombe transcription elongation factor Spt5, which consists of 18 tandem repeats of a nonapeptide of consensus sequence 1TPAWNSGSK9. We document the divalent cation dependence and specificity of SpCdk9/Pch1, its NTP dependence and specificity, the dependence of Spt5-CTD phosphorylation on the number of tandem nonamer repeats, and the specificity for phosphorylation of the Spt5-CTD on threonine at position 1 within the nonamer element. SpCdk9/Pch1 also phosphorylates the CTD heptaptide repeat array of the largest subunit of S. pombe RNA polymerase II (consensus sequence YSPTSPS) and does so exclusively on serine. SpCdk9/Pch1 catalyzes autophosphorylation of the kinase and cyclin subunits of the kinase complex. The distribution of phosphorylation sites on SpCdk9 (86% Ser(P), 11% Thr(P), 3% Tyr(P)) is distinct from that on Pch1 (2% Ser(P), 98% Thr(P)). We conducted a structure-guided mutational analysis of SpCdk9, whereby a total of 29 new mutations of 12 conserved residues were tested for in vivo function by complementation of a yeast bur1Delta mutant. We identified many lethal and conditional mutations of side chains implicated in binding ATP and the divalent cation cofactor, phosphoacceptor substrate recognition, and T-loop dynamics. We surmise that the lethality of the of T212A mutation in the T-loop reflects an essential phosphorylation event, insofar as the conservative T212S change rescued wild-type growth; the phosphomimetic T212E change rescued growth at 30 degrees C; and the effects of mutating the T-loop threonine were phenocopied by mutations in the three conserved arginines predicted to chelate the phosphate on the T-loop threonine.
 ...
PMID:Characterization of the Schizosaccharomyces pombe Cdk9/Pch1 protein kinase: Spt5 phosphorylation, autophosphorylation, and mutational analysis. 1290 90

Macronucleus of Tetrahymena divides amitotically, although in a microtubule-dependent fashion. Besides the localization study and pharmacological study of macronuclear microtubules, mechanism of the macronuclear division is poorly understood. A biochemical search for microtubule-associated protein was attempted from the isolated macronucleus. Improvement on macronucleus isolation method and microtubule coprecipitation assay led to the cloning of p138, a new homologue of transcription elongation factor FACT (facilitates chromatin transcription) 140kDa subunit. DNase treatment test of macronuclear extract and the sequence of p138 suggested that p138 is associated with chromosome in the macronucleus. The release tests of p138 from microtubules indicated that p138 is released from microtubules in the presence of ATP but not in the presence of AMP-PNP. Together, the results suggest a novel function of FACT homologue, that p138 interacts with both microtubules and chromosome.
 ...
PMID:Identification and molecular cloning of Tetrahymena 138-kDa protein, a transcription elongation factor homologue that interacts with microtubules in vitro. 1501 45

Streptococcus mutans is an important pathogen in the initiation of dental caries as the bacterium remains metabolically active when the environment becomes acidic. The mechanisms underlying this ability to survive and proliferate at low pH remain an area of intense investigation. Differential two-dimensional electrophoretic proteome analysis of S. mutans grown at steady state in continuous culture at pH 7.0 or pH 5.0 enabled the resolution of 199 cellular and extracellular protein spots with altered levels of expression. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry identified 167 of these protein spots. Sixty-one were associated with stress-responsive pathways involved in DNA replication, transcription, translation, protein folding and proteolysis. The 61 protein spots represented isoforms or cleavage products of 30 different proteins, of which 25 were either upregulated or uniquely expressed during acid-tolerant growth at pH 5.0. Among the unique and upregulated proteins were five that have not been previously identified as being associated with acid tolerance in S. mutans and/or which have not been studied in any detail in oral streptococci. These were the single-stranded DNA-binding protein, Ssb, the transcription elongation factor, GreA, the RNA exonuclease, polyribonucleotide nucleotidyltransferase (PnpA), and two proteinases, the ATP-binding subunit, ClpL, of the Clp family of proteinases and a proteinase encoded by the pep gene family with properties similar to the dipeptidase, PepD, of Lactobacillus helveticus. The identification of these and other differentially expressed proteins associated with an acid-tolerant-growth phenotype provides new information on targets for mutagenic studies that will allow the future assessment of their physiological significance in the survival and proliferation of S. mutans in low pH environments.
 ...
PMID:Stress-responsive proteins are upregulated in Streptococcus mutans during acid tolerance. 1513 96

Cdk7 performs two essential but distinct functions as a CDK-activating kinase (CAK) required for cell-cycle progression and as the RNA polymerase II (Pol II) CTD kinase of general transcription factor IIH. To investigate the substrate specificity underlying this dual function, we created an analog-sensitive (AS) Cdk7 able to use bulky ATP derivatives. Cdk7-AS-cyclin H-Mat1 phosphorylates approximately 10-15 endogenous polypeptides in nuclear extracts. We identify seven of these as known and previously unknown Cdk7 substrates that define two classes: proteins such as Pol II and transcription elongation factor Spt5, recognized efficiently only by the fully activated Cdk7 complex, through sequences surrounding the site of phosphorylation; and CDKs, targeted equivalently by all active forms of Cdk7, dependent on substrate motifs remote from the phosphoacceptor residue. Thus, Cdk7 accomplishes dual functions in cell-cycle control and transcription not through promiscuity but through distinct, stringent modes of substrate recognition.
 ...
PMID:Dichotomous but stringent substrate selection by the dual-function Cdk7 complex revealed by chemical genetics. 1632 5

Promoter clearance and transcriptional processivity in eukaryotic cells are fundamentally regulated by the phosphorylation of the carboxy-terminal domain of RNA polymerase II (RNAPII). One of the kinases that essentially performs this function is P-TEFb (positive transcription elongation factor b), which is composed of cyclin-dependent kinase 9 (CDK9) associated with members of the cyclin T family. Here we show that cellular GCN5 and P/CAF, members of the GCN5-related N-acetyltransferase family of histone acetyltransferases, regulate CDK9 function by specifically acetylating the catalytic core of the enzyme and, in particular, a lysine that is essential for ATP coordination and the phosphotransfer reaction. Acetylation markedly reduces both the kinase function and transcriptional activity of P-TEFb. In contrast to unmodified CDK9, the acetylated fraction of the enzyme is specifically found in the insoluble nuclear matrix compartment. Acetylated CDK9 associates with the transcriptionally silent human immunodeficiency virus type 1 provirus; upon transcriptional activation, it is replaced by the unmodified form, which is involved in the elongating phase of transcription marked by Ser2-phosphorylated RNAPII. Given the conservation of the CDK9 acetylated residues in the catalytic task of virtually all CDK proteins, we anticipate that this mechanism of regulation might play a broader role in controlling the function of other members of this kinase family.
 ...
PMID:Acetylation of conserved lysines in the catalytic core of cyclin-dependent kinase 9 inhibits kinase activity and regulates transcription. 1825 Jan 57

Mycoplasmas are parasitic bacteria with small genomes. Since parasitic bacteria need to adapt themselves to their hosts, there is a possibility that some genes evolved under species-specific constraint. We assume that Ureaplasma parvum has candidate genes that evolved in a species-specific manner in its genome. Here we examined synonymous-to-nonsynonymous substitution ratios (omega) of the 143 mycoplasma-orthologous genes of Ureaplasma and other mycoplasmas using branch models. As a result, the model allowing for Ureaplasma branch-specific omega in addition to omega of other mycoplasmas was significantly supported in 16 genes. First, the Ureaplasma-specific model was significantly supported in the genes encoding a transcription elongation factor and a transcription terminator factor, suggesting that transcription-related genes of Ureaplasma have evolved in a unique manner compared to those of other mycoplasmas. Second, the Ureaplasma-specific model was significantly supported in the gene encoding uracil-DNA glycosylase. In addition, the omega value of the gene in the Ureaplasma lineage was approximately 30-fold lower than those of other lineages, suggesting that uracil-DNA glycosylase of Ureaplasma evolved under stronger functional constraint than those of other mycoplasmas. Finally, three glycolytic genes of Ureaplasma were suggested to have evolved under relaxed selection. Among mycoplasmas, only Ureaplasma has urease and synthesizes ATPs via hydrolysis of urea. This raises the possibility that Ureaplasma does not need a glycolysis pathway for ATP synthesis. This unique energy-producing system may be related to the Ureaplasma-specific evolution of the glycolytic genes.
 ...
PMID:Detection of the genes evolving under Ureaplasma-specific selection. 1841 24


1 2 Next >>