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 DNA dependent protein kinase (DNA-PK) is a trimeric nuclear complex consisting of a large protein kinase and the Ku heterodimer that regulates kinase activity by its association with DNA. Recent findings have shown structural similarities between DNA-PK and a family of lipid and putative protein kinases (PIK family). DNA-PK is one of the PIK members known to be a protein kinase with clearly identified effector subunits. A broad range of observations link DNA-PK to dual roles in double strand DNA break (DSB) repair and transcription. Unlike its most closely related PIKs, DNA-PK is not required for activating cell cycle regulated DNA damage signalling mechanisms. Instead, the phenotypes and biochemical properties of DNA-PK are most consistent with functions in DSB repair and joining steps in recombination mechanisms. DNA-PK is associated with RNA polymerase II and RNA polymerase I transcription complexes, where it most frequently has a negative regulatory role.
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PMID:Functions of the DNA dependent protein kinase. 933 3

The C-terminal part of the largest subunit of eukaryotic RNA polymerase II is composed solely of the highly repeated consensus sequence Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7. This domain, called the C-terminal domain (CTD), is phosphorylated mostly at serine residues during transcription initiation, but the precise role of this phosphorylation remains controversial. Several protein kinases are able to phosphorylate this sequence in vitro. The aim of this work was to define the positions of the amino acids phosphorylated by four of these CTD kinases (transcription factor (TF) IIH-kinase, DNA-dependent protein kinase, and the mitogen-activated protein kinases ERK1 and ERK2) and to compare the specificity of these different protein kinases. We show that TFIIH kinase and the mitogen-activated protein kinases phosphorylate only serine 5 of the CTD sequence, whereas DNA-dependent protein kinase phosphorylates serines 2 and 7. Among the different CTD kinases, only TFIIH kinase is appreciably more active on two repeats of the consensus sequence than on one motif. These in vitro results can provide some clues to the nature of the protein kinases responsible for the in vivo phosphorylation of the RNA polymerase CTD. In particular, the ratio of phosphorylated serine to threonine observed in vivo cannot be explained if TFIIH kinase is the only protein kinase involved in the phosphorylation of the CTD.
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PMID:Characterization of the residues phosphorylated in vitro by different C-terminal domain kinases. 950 78

Ku is a DNA binding protein composed of 70 and 80 kDa subunits which was discovered as autoantigen in a patient with scleroderma-polymyositis overlap syndrome. Ku can bind to the end of DNA and also to some internal sequences. Ku-autoantigen acts as a potential transcription factor for several RNA polymerase II genes and RNA polymerase I gene. Ku is also associated with DNA-dependent protein kinase and involved in V(D)J recombination and DNA break repair mechanisms. Ku may be involved in replication, helicase activity and cell signaling. Therefore, Ku-autoantigen is a very important cellular factor which plays important role in the multiple cellular processes.
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PMID:Autoantigen Ku and its role in multiple cellular processes. 956 59

DNA-dependent protein kinase (DNA-PK) is involved in DNA repair but there is some evidence to suggest that it is also involved in regulating transcription. We used a pair of cell lines, SCVA2 and SC(8)-10, which are DNA-PK negative and positive respectively, in order to examine the effect of DNA-PK upon transcription. Initial experiments were performed using p53 as an activator of transcription because DNA-PK has been proposed as a candidate upstream activator of p53. It was found both in vivo and in vitro that efficient p53-dependent transcription required the presence of DNA-PK. However, phosphorylation of p53 by DNA-PK did not affect the DNA-binding ability of p53 nor its transcriptional activity when tested in vitro. Subsequent in vivo experiments suggested that a number of transcription activators functioned more efficiently in the presence of DNA-PK. Therefore DNA-PK may play a general role in regulation of transcription driven by RNA polymerase II. In addition, DNA-PK is shown to have no specific effect on p53-dependent transcription.
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PMID:Transcription by RNA polymerase II in DNA-PK deficient scid mouse cells. 1097 5

Ecteinascidin-743 (ET-743), an anti-tumor agent derived from the marine tunicate, Ecteinascidia turbinata, is active against various solid tumor cell lines, including soft tissue sarcoma, breast, ovarian, non-small-cell lung and prostate cancers and melanoma, and has a broad spectrum of anti-cancer activity in vivo. For reasons as yet unclear, sarcoma cell lines are exquisitely sensitive to ET-743. The drug has a unique mechanism of action that makes it a novel anti-tumor agent. ET-743 is a DNA-binding agent that covalently interacts with the minor groove of the DNA double helix to bend the molecule towards the major groove. Defects in DNA repair pathways have paradoxical effects on the anti-tumor activity of ET-743: loss of mismatch repair does not affect its toxicity; loss of DNA-dependent protein kinase activity enhances its toxicity; defects in transcription-coupled nucleotide excision repair confer resistance to ET-743. As a DNA repair capability appears to be necessary for at least one mechanism of ET-743-mediated cytotoxicity, the drug may interact with the DNA repair machinery to induce lethal strand breaks. One of the most novel aspects of ET-743 is its effect on RNA polymerase II-mediated gene transcription. ET-743 selectively inhibits activation of the multidrug resistance gene, while leaving constitutive gene expression relatively unaffected. Preliminary studies of other genes and transcriptional inducers indicate that ET-743 may be a more general inhibitor of activated, but not basal, transcription.
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PMID:ET-743: more than an innovative mechanism of action. 1217 91

Ku is an abundant nuclear protein with an essential function in the repair of DNA double-strand breaks. Various observations suggest that Ku also interacts with the cellular transcription machinery, although the mechanism and significance of this interaction are not well understood. In the present study, we investigated the subnuclear distribution of Ku in normally growing human cells by using confocal microscopy, chromatin immunoprecipitation, and protein immunoprecipitation. All three approaches indicated association of Ku with RNA polymerase II (RNAP II) elongation sites. This association occurred independently of the DNA-dependent protein kinase catalytic subunit and was highly selective. There was no detectable association with the initiating isoform of RNAP II or with the general transcription initiation factors. In vitro protein-protein interaction assays demonstrated that the association of Ku with elongation proteins is mediated, in part, by a discrete C-terminal domain in the Ku80 subunit. Functional disruption of this interaction with a dominant-negative mutant inhibited transcription in vitro and in vivo and suppressed cell growth. These results suggest that association of Ku with transcription sites is important for maintenance of global transcription levels. Tethering of double-strand break repair proteins to defined subnuclear structures may also be advantageous in maintenance of genome stability.
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PMID:Subnuclear localization of Ku protein: functional association with RNA polymerase II elongation sites. 1239 Nov 74

DNA-dependent protein kinase represses RNA polymerase I (Pol I) transcription in vitro. To investigate the mechanism underlying transcriptional repression, we compared Pol I transcription in extracts from cells that either contain or lack the catalytic subunit of DNA-PK (DNA-PKcs). ATP-dependent repression of Pol I transcription was observed in extracts from DNA-PKcs-containing but not -deficient cells, required templates with free DNA ends, and was overcome by exogenous SL1, the factor that nucleates initiation complex formation. Order-of-addition experiments demonstrate that DNA-PKcs does not inactivate component(s) of the Poll transcription machinery. Instead, phosphorylated Ku protein competes with SL1 for binding to the rDNA promoter and, as a consequence, prevents initiation complex formation. The results reveal a novel mechanism of transcriptional regulation by DNA-PK. Once targeted to DNA, autophosphorylated Ku may displace positive- or negative-acting factors from their target sites, thereby repressing or activating transcription in a gene-specific manner.
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PMID:Mechanism of inhibition of RNA polymerase I transcription by DNA-dependent protein kinase. 1253 May 33

We have shown that a constitutively active Galpha13 (Galpha13Q226L) induces differentiation in P19 embryonic carcinoma cells to an endodermal phenotype. In this report, we demonstrate that Ku, a heterodimer of p80 (Ku80) and p70 (Ku70), is upregulated in P19 cells overexpressing Galpha13Q226L. Ku is the regulatory subunit of the DNA-dependent protein kinase and is primarily involved in DNA repair and recombination. Ku80 also is a somatostatin receptor. We show that while overexpression of Ku80 drastically reduced P19 cell proliferation, it was not sufficient to induce endodermal differentiation. However, coexpression of Galpha13Q226L and an antisense Ku80 abrogated the retarded growth rate and endodermal differentiation observed in cells expressing only Galpha13Q226L. Overexpression of Galpha13Q226L or Ku80 downregulated RNA polymerase I-mediated transcriptional activity and overexpression of antisense Ku80 restored the activity to control level. These results suggest that Ku80 is required for Galpha13-mediated endodermal differentiation in P19 cells.
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PMID:Ku80 is required but not sufficient for Galpha13-mediated endodermal differentiation in P19 embryonic carcinoma cells. 1535 36

The DNA-dependent protein kinase catalytic subunit (DNA-PKcs) regulates the non-homologous end-joining pathway of DNA double-strand break repair in mammalian cells. The ability of DNA-PKcs to sense and respond to different terminal DNA structures is postulated to be important for its regulatory function. It is unclear whether discrimination occurs at the time of formation of the initial protein-DNA complex or later, at the time of formation of a paired, or synaptic complex between opposing DNA ends. To gain further insight into the mechanism of regulation, we characterized the binding of DNA-PKcs to immobilized DNA fragments that cannot undergo synapsis. Results showed that DNA-PKcs strongly discriminates between different terminal structures at the time of initial complex formation. Although Ku protein stabilizes DNA-PKcs binding overall, it is not required for discrimination between terminal structures. Base mispairing, temperature and the presence of an interstrand linkage influence the stability of the initial complex in a manner that suggests a requirement for DNA unwinding, reminiscent of the 'open complex' model of RNA polymerase-promoter DNA interaction. ATP and a nonhydrolyzable ATP analog also influence the stability of the DNA-PKcs*DNA complex, apparently by an allosteric mechanism that does not require DNA-PKcs autophosphorylation.
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PMID:Terminal DNA structure and ATP influence binding parameters of the DNA-dependent protein kinase at an early step prior to DNA synapsis. 1648 83

BRCA1-associated RING domain protein BARD1, along with its heterodimeric partner BRCA1, plays important roles in cellular response to DNA damage. Immediate cellular response to genotoxic stress is mediated by a family of phosphoinositide 3-kinase-related protein kinases, such as ataxia-telangiectasia mutated (ATM), ATM and Rad3-related, and DNA-dependent protein kinase. ATM-mediated phosphorylation of BRCA1 enhances the DNA damage checkpoint functions of BRCA1, but how BARD1 is regulated during DNA damage signaling has not been examined. Here, we report that BARD1 undergoes phosphorylation upon ionizing radiation or UV radiation and identify Thr(714) as the in vivo BARD1 phosphorylation site. Importantly, DNA damage functions of BARD1 (i.e., inhibition of pre-mRNA polyadenylation and degradation of RNA polymerase II) are abrogated in T714A and T734A mutants. Our findings suggest that phosphorylation of BARD1 is critical for the DNA damage functions of the BRCA1/BARD1 complex.
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PMID:DNA damage-induced BARD1 phosphorylation is critical for the inhibition of messenger RNA processing by BRCA1/BARD1 complex. 1665 5


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