Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The RPB9 subunit of RNA polymerase II regulates transcription elongation activity and is required for the action of the transcription elongation factor, TFIIS. RPB9 comprises two zinc ribbon domains joined by a conserved linker region. The C-terminal zinc ribbon is similar in sequence to that found in TFIIS. To elucidate the relationship between the structure and transcription elongation function of RPB9, we initiated a mutagenesis study on the Saccharomyces cerevisiae homologue. The individual zinc ribbon domains, in isolation or in combination, could not stimulate transcription by a polymerase lacking RPB9, pol IIDelta9. Mutations in the N-terminal zinc ribbon had little effect on transcription activity. By contrast, mutations in the acidic loop that connects the second and third beta-strands of the C-terminal zinc ribbon were completely inactive for transcription. Interestingly, the analogous residues in TFIIS are also critical for elongation activity. A conserved charged stretch in the linker region (residues 89-95, DPTLPR) mediated the interaction with RNA polymerase II.
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PMID:Yeast RNA polymerase II subunit RPB9. Mapping of domains required for transcription elongation. 1064 77

The Rpb6 subunit of RNA polymerase II is one of the five subunits common to three forms of eukaryotic RNA polymerase. Deletion and truncation analyses of the rpb6 gene in the fission yeast Schizosaccharomyces pombe indicated that Rpb6, consisting of 142 amino acid residues, is an essential protein for cell viability, and the essential region is located in the C-terminal half between residues 61 and 139. After random mutagenesis, a total of 14 temperature-sensitive mutants were isolated, each carrying a single (or double in three cases and triple in one) mutation. Four mutants each carrying a single mutation in the essential region were sensitive to 6-azauracil (6AU), which inhibits transcription elongation by depleting the intracellular pool of GTP and UTP. Both 6AU sensitivity and temperature-sensitive phenotypes of these rpb6 mutants were suppressed by overexpression of TFIIS, a transcription elongation factor. In agreement with the genetic studies, the mutant RNA polymerases containing the mutant Rpb6 subunits showed reduced affinity for TFIIS, as measured by a pull-down assay of TFIIS-RNA polymerase II complexes using a fusion form of TFIIS with glutathione S-transferase. Moreover, the direct interaction between TFIIS and RNA polymerase II was competed by the addition of Rpb6. Taken together, the results lead us to propose that Rpb6 plays a role in the interaction between RNA polymerase II and the transcription elongation factor TFIIS.
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PMID:The Rpb6 subunit of fission yeast RNA polymerase II is a contact target of the transcription elongation factor TFIIS. 1064 12

The Elongin complex stimulates the rate of transcription elongation by RNA polymerase II by suppressing the transient pausing of the polymerase at many sites along the DNA template. Elongin is composed of a transcriptionally active A subunit and two small regulatory B and C subunits, the latter of which bind stably to each other to form a binary complex that interacts with Elongin A and strongly induces its transcriptional activity. To further understand the roles of Elongin in transcriptional regulation, we attempted to identify Elongin-related proteins. Here, we report on the cloning, expression, and characterization of human Elongin A2, a novel transcription elongation factor that exhibited 47% identity and 61% similarity to Elongin A. Biochemical studies have shown that Elongin A2 stimulates the rate of transcription elongation by RNA polymerase II and is capable of forming a stable complex with Elongin BC. However, in contrast to Elongin A, its transcriptional activity is not activated by Elongin BC. Northern blot analysis revealed that Elongin A2 mRNA was specifically expressed in the testis, suggesting that Elongin A2 may regulate the transcription of testis-specific genes.
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PMID:Identification and characterization of Elongin A2, a new member of the Elongin family of transcription elongation factors, specifically expressed in the testis. 1069 60

Elongin is a transcription elongation factor that was first identified in mammalian systems and is composed of the three subunits, elongin A, B, and C. Sequence homologues of elongin A and elongin C, but not elongin B, were identified in the yeast genome. Neither yeast elongin A nor C sequence homologues was required for cell viability. The two gene products could be purified from yeast as a complex. A recombinant form of the complex, which could only be produced in bacteria if the gene products were co-expressed, was purified over several chromatographic steps. The complex did not stimulate transcription elongation by yeast RNA polymerase II. Using limited proteolysis, the N-terminal 144 residues of yeast elongin A were shown to be sufficient for interaction with yeast elongin C. The purified complex of yeast elongin C/elongin A(1-143) was analyzed using circular dichroism and nuclear magnetic spectroscopy. These studies revealed that yeast elongin A is unfolded but undergoes a dramatic modification of its structure in the presence of elongin C, and that elongin C forms a stable dimer in the absence of elongin A.
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PMID:Elongin from Saccharomyces cerevisiae. 1075 24

The activation of the HIV-1 long terminal repeat (LTR) by the viral transcriptional transactivator Tat is an essential step in the viral replication cycle. To increase the processivity of RNA polymerase II, Tat interacts with the positive transcription elongation factor b (P-TEFb) and cyclin-dependent kinase (CDK)-activating kinase (CAK). In this study, we demonstrate that a pseudo-substrate peptide for CDK7, mC2p, inhibits HIV-1 replication as well as Tat transactivation. Specifically, mC2p blocks only the activity of CAK and not that of P-TEFb. Moreover, mC2p inhibits Tat transactivation and HIV replication. Therefore, the activation of CDK7 by Tat is considered a critical step of Tat transactivation and mC2p and related compounds represent potential candidates for novel anti-HIV therapeutics.
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PMID:HIV-1 replication is inhibited by a pseudo-substrate peptide that blocks Tat transactivation. 1079 93

TFIIS is a transcription elongation factor that consists of three domains. We have previously solved the structures of domains II and III, which stimulate arrested polymerase II elongation complexes in order to resume transcription. Domain I is conserved in evolution from yeast to human species and is homologous to the transcription factors elongin A and CRSP70. Domain I also interacts with the transcriptionally active RNA polymerase II holoenzyme and therefore, may have a function unrelated to the previously described transcription elongation activity of TFIIS. We have solved the structure of domain I of yeast TFIIS using NMR spectroscopy. Domain I is a compact four-helix bundle that is structurally independent of domains II and III of the TFIIS. Using the yeast structure as a template, we have modeled the homologous domains from elongin A and CRSP70 and identified a conserved positively charged patch on the surface of all three proteins, which may be involved in conserved functional interactions with the transcriptional machinery.
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PMID:Structure of a conserved domain common to the transcription factors TFIIS, elongin A, and CRSP70. 1081 49

Eukaryotic mRNA synthesis is catalyzed by multisubunit RNA polymerase II and proceeds through multiple stages referred to as preinitiation, initiation, elongation, and termination. Over the past 20 years, biochemical studies of eukaryotic mRNA synthesis have largely focused on the preinitiation and initiation stages of transcription. These studies led to the discovery of the class of general initiation factors (TFIIB, TFIID, TFIIE, TFIIF, and TFIIH), which function in intimate association with RNA polymerase II and are required for selective binding of polymerase to its promoters, formation of the open complex, and synthesis of the first few phosphodiester bonds of nascent transcripts. Recently, biochemical studies of the elongation stage of eukaryotic mRNA synthesis have led to the discovery of several cellular proteins that have properties expected of general elongation factors and that have been found to play unanticipated roles in human disease. Among these candidate general elongation factors are the positive transcription elongation factor b (P-TEFb), eleven-nineteen lysine-rich in leukemia (ELL), Cockayne syndrome complementation group B (CSB), and elongin proteins, which all function in vitro to expedite elongation by RNA polymerase II by suppressing transient pausing or premature arrest by polymerase through direct interactions with the elongation complex. Despite their similar activities in elongation, the P-TEFb, ELL, CSB, and elongin proteins appear to play roles in a diverse collection of human diseases, including human immunodeficiency virus-1 infection, acute myeloid leukemia, Cockayne syndrome, and the familial cancer predisposition syndrome von Hippel-Lindau disease. here we review our current understanding of the P-TEFb, ELL, CSB, and elongin proteins, their mechanisms of action, and their roles in human disease.
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PMID:Transcription elongation and human disease. 1087 52

We report that the chromatin-specific transcription elongation factor FACT functions in conjunction with the RNA polymerase II CTD kinase P-TEFb to alleviate transcription inhibition by DSIF (DRB sensitivity-inducing factor) and NELF (negative elongation factor). We find that the kinase activity of TFIIH is dispensable for this activity, demonstrating that TFIIH-mediated CTD phosphorylation is not involved in the regulation of FACT and DSIF/NELF activities. Thus, we propose a novel transcriptional regulatory network in which DSIF/NELF inhibition of transcription is prevented by P-TEFb in cooperation with FACT. This study uncovers a novel role for FACT in the regulation of transcription on naked DNA that is independent of its activities on chromatin templates. In addition, this study reveals functional differences between P-TEFb and TFIIH in the regulation of transcription.
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PMID:FACT relieves DSIF/NELF-mediated inhibition of transcriptional elongation and reveals functional differences between P-TEFb and TFIIH. 1091 1

Tat stimulation of human immunodeficiency virus type 1 (HIV-1) transcription requires Tat-dependent recruitment of human positive transcription elongation factor b (P-TEFb) to the HIV-1 promoter and the formation on the trans-acting response element (TAR) RNA of a P-TEFb-Tat-TAR ternary complex. We show here that the P-TEFb heterodimer of Cdk9-cyclin T1 is intrinsically incapable of forming a stable complex with Tat and TAR due to two built-in autoinhibitory mechanisms in P-TEFb. Both mechanisms exert little effect on the P-TEFb-Tat interaction but prevent the P-TEFb-Tat complex from binding to TAR RNA. The first autoinhibition arises from the unphosphorylated state of Cdk9, which establishes a P-TEFb conformation unfavorable for TAR recognition. Autophosphorylation of Cdk9 overcomes this inhibition by inducing conformational changes in P-TEFb, thereby exposing a region in cyclin T1 for possible TAR binding. An intramolecular interaction between the N- and C-terminal regions of cyclin T1 sterically blocks the P-TEFb-TAR interaction and constitutes the second autoinhibitory mechanism. This inhibition is relieved by the binding of the C-terminal region of cyclin T1 to the transcription elongation factor Tat-SF1 and perhaps other cellular factors. Upon release from the intramolecular interaction, the C-terminal region also interacts with RNA polymerase II and is required for HIV-1 transcription, suggesting its role in bridging the P-TEFb-Tat-TAR complex and the basal elongation apparatus. These data reveal novel control mechanisms for the assembly of a multicomponent transcription elongation complex at the HIV-1 promoter.
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PMID:Relief of two built-In autoinhibitory mechanisms in P-TEFb is required for assembly of a multicomponent transcription elongation complex at the human immunodeficiency virus type 1 promoter. 1091 73

RNA polymerase II lacking the Rpb9 subunit uses alternate transcription initiation sites in vitro and in vivo and is unable to respond to the transcription elongation factor TFIIS in vitro. Here, we show that RPB9 has a synthetic phenotype with the TFIIS gene. Disruption of RPB9 in yeast also resulted in sensitivity to 6-azauracil, which is a phenotype linked to defects in transcription elongation. Expression of the TFIIS gene on a high-copy plasmid partially suppressed the 6-azauracil sensitivity of Deltarpb9 cells. We set out to determine the relevant cellular role of yeast Rpb9 by assessing the ability of 20 different site-directed and deletion mutants of RPB9 to complement the initiation and elongation defects of Deltarpb9 cells in vivo. Rpb9 is composed of two zinc ribbons. The N-terminal zinc ribbon restored the wild-type pattern of initiation start sites, but was unable to complement the growth defects associated with defects in elongation. Most of the site-directed mutants complemented the elongation-specific growth phenotypes and reconstituted the normal pattern of transcription initiation sites. The anti-correlation between the growth defects of cells disrupted for RPB9 and the selection of transcription start sites suggests that this is not the primary cellular role for Rpb9. Genome-wide transcription profiling of Deltarpb9 cells revealed only a few changes, predominantly in genes related to metabolism.
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PMID:RNA polymerase II subunit Rpb9 regulates transcription elongation in vivo. 1093 84


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