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

---

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

We have studied the effect of protein phosphokinase (EC 2.7.1.37; ATP:protein phosphotransferase) and phosphoprotein phosphatase (EC 3.1.3.16; phosphoprotein phosphohydrolase) on reverse transcriptase (RNA-dependent DNA nucleotidyltransferase) activity of Rous sarcoma virus. Protein kinase from Rous sarcoma virus-transformed chick embryo fibroblasts was purified by DEAE-cellulose chromatography, Sephadex gel filtration, and isoelectric focusing. Purified reverse transcriptase from Rouse sarcoma virus was preincubated with protein kinase and ATP under conditions allowing incorporation of phosphate into substrate protein. After the preincubation, reverse transcriptase activity was assayed in the presence of poly(rA).oligo(dT) as template. A 2- to 5-fold increase of reverse transcriptase activity was found after the preincubation of reverse transcriptase with protein kinase and ATP. Incubation of reverse transcriptase with heat-treated, inactive protein kinase and ATP had no effect on transcriptase activity. When the transcriptase preparation was incubated with protein kinase and [gamma-32P]ATP and subsequently purified by chromatography on phosphocellulose and Sephadex gel filtration, significant amounts of 32P-labeled proteins were found in the fractions exhibiting reverse transcriptase activity, suggesting 32P incorporation into transcriptase or transcriptase-associated proteins. A 20-60% decrease of reverse transcriptase activity was observed after incubation of reverse transcriptase with phosphatase. The results suggest that phosphorylative modification of reverse transcriptase may be critical in the regulation of reverse transcriptase-catalyzed DNA synthesis.
...
PMID:Protein kinase and its regulatory effect on reverse transcriptase activity of Rous sarcoma virus. 5 72

p34cdc2 kinase, a critical regulator of the cell cycle, has been shown to recognize the consensus sequence S/TP in proteins such as histone H1, the retinoblastoma gene product RB and the carboxyl-terminal domain of eukaryotic RNA polymerase II. Using phosphorylated synthetic peptides, representing the p34cdc2 phosphorylation sites in these proteins and histone H1 protein as substrates, we investigated the substrate specificity of the different oligomeric forms of the polycation-stimulated (PCS/type-2A) protein phosphatase and the active catalytic subunit of the ATP,Mg-dependent (AMDc/type 1) protein phosphatase. The results show that the oligomeric structure of the PCS phosphatases is an important determinant for efficient dephosphorylation. The trimeric PCSH1 and PCSM phosphatases are about 10-20-fold-better histone H1 phosphatases than the dimeric PCSH2 and PCSL phosphatases and about 100-fold better than the catalytic subunit (PCSC), suggesting a regulatory role for the 72-kDa, 65-kDa and 55-kDa subunits. The RB peptide = INGS(P)PRT(P)PRRGQNR, is preferred over phosphorylase a (8-fold) by the PCSH1 phosphatase and is about a 40-fold and 95-fold-better substrate for the PCSH1 phosphatase than for the PCSM and PCSL phosphatases, respectively. The primary structure surrounding the S/T(P)P motif, by itself a strong negative determinant for dephosphorylation, can harbour positive features which relieve the constraint imposed by the carboxyl-terminal proline. Thus, the RB peptide INGS(P)PRT(P)PRRGQNR, in which the T(P)P configuration is preferred over the S(P)P sequence, is an extremely good and specific substrate for the PCSH1 phosphatase (Km = 10 microM, Vmax = 3882 nmol.min-1.mg-1). The AMDC phosphatase is a poor phosphatase for all the phosphopeptides tested, unless Mn2+ is added. Its histone H1 phosphatase activity is much less sensitive than its phosphorylase a and phosphopeptide phosphatase activity to inhibition by the modulator or inhibitor-1. The results strongly suggest a role for the trimeric PCSH1 phosphatase in reversing the p34cdc2 phosphorylations.
...
PMID:Specificity of the polycation-stimulated (type-2A) and ATP,Mg-dependent (type-1) protein phosphatases toward substrates phosphorylated by P34cdc2 kinase. 131 64

We examined the effects of okadaic acid (OA), a potent and specific inhibitor of serine phosphatases 2A and 1, on the transient expression of an hsp 70 promoter-reporter gene construct in IMR-90 human diploid lung fibroblasts. We showed that OA markedly potentiated the heat-induced but not the basal expression of pHBCAT, a full-length human hsp-70-promoter-driven CAT gene construct. This effect of OA was dose and time dependent and promoter specific. Importantly, the potentiating effects of OA appeared to be independent of the binding of the activated heat shock transcription factor (HSTF) to its consensus DNA sequence, the heat shock element (HSE). Thus, OA had no effect on the HSTF DNA-binding activity as measured by mobility shift assay, and mutation of the HSE sequence did not obliterate the stimulatory effects of OA on reporter gene expression under a heat shock condition, although heat shock by itself was without effect. Analysis of the status of phosphorylation of the largest subunit of RNA polymerase II provided evidence that this effect of OA is attributable, at least in part, to the increased phosphorylation of RNA polymerase II. These results provided evidence that the heat-induced hsp 70 promoter activity is negatively regulated by serine phosphatases. We propose that the heat-induced transcriptional activation of hsps is associated with phosphorylation of component(s) of the transcription complex; one of the likely candidates being the transcriptionally engaged RNA polymerase II. OA, by inhibiting phosphatase 2A and 1 activity, enhanced this phosphorylation and potentiated the transcriptional activation of hsps.
...
PMID:The heat-induced hsp 70 promoter activity is negatively regulated by serine phosphatases: evidence from the effects of okadaic acid. 133 79

We have previously described the trypanosomal gene encoding the largest subunit of RNA polymerase II (RNAP II) and found that two almost identical genes are encoded within the Trypanosoma brucei genome. Here we show by Southern analyses that the 5' breakpoint between both loci is located approximately 7.5 kb upstream of the RNAP II genes. Northern analyses revealed that the 5' duplicated segment contains at least four other genes, which are transcribed in both bloodstream and procyclic trypanosomes. The gene located immediately upstream of the RNAP II gene in both loci was characterized by sequence analyses. The deduced amino acid sequences show a high degree of similarity to the catalytic subunit of protein phosphatase class 1 (PP1) genes. S1 mapping provided strong evidence in support of the fact that the PP1 and RNAP II genes belong to a single transcription unit.
...
PMID:The Trypanosoma brucei protein phosphatase gene: polycistronic transcription with the RNA polymerase II largest subunit gene. 216 4

Reversion analysis has identified four suppressor genes that permit transcription of the Saccharomyces cerevisiae HIS4 gene in the absence of GCN4, BAS1, and BAS2, trans-acting proteins normally required for activation of HIS4 transcription. These suppressor genes encode factors that affect the transcription of many diverse genes. Two of these suppressors, SIT1 and SIT2, are encoded by RPB1 and RPB2, the genes for the two largest subunits of RNA polymerase II. All strains containing suppressor mutations in RPB1 and RPB2 have reduced transcription of the INO1 gene and an inositol requirement. Mutations in SIT3 or high copy number SIT3 increase HIS4 transcription in the absence of GCN4, BAS1, and BAS2. This increase in HIS4 transcription by high copy number SIT3 or by sit3 alleles is largely independent of the HIS4 TATA sequence. The SIT4 protein is over 50% identical to the catalytic subunit of bovine type 2A protein phosphatase. sit4 mutations in combination with suppressor mutations in RPB1 or RPB2 (sit1, sit4 or sit2, sit4) are lethal, suggesting an interaction between SIT4 and RNA polymerase II.
...
PMID:A suppressor of a HIS4 transcriptional defect encodes a protein with homology to the catalytic subunit of protein phosphatases. 253 49

The repetitive C-terminal domain (CTD) of RNA polymerase (RNAP) II is extensively phosphorylated concomitant with the initiation of transcription and must be dephosphorylated before RNAP II can begin another round of transcription. A CTD phosphatase was purified more than 7,500-fold from a HeLa cell extract. SDS-polyacrylamide gel electrophoresis shows a predominant protein of 205 kDa and a less abundant protein of 150 kDa co-eluting with the CTD phosphatase activity. Sedimentation and gel filtration analysis suggest that CTD phosphatase has an elongated structure with a M(r) of 200,000. This enzyme is a type 2C phosphatase in that it requires Mg2+ for activity and is resistant to okadaic acid. CTD phosphatase appears to processively dephosphorylate the CTD and is specific in that it does not dephosphorylate phosphorylase a, the alpha or beta subunits of phosphorylase kinase or RNAP II phosphorylated with casein kinase II. CTD phosphatase dephosphorylates RNAP IIO purified from calf thymus or generated in vitro by two previously described CTD kinases. These results suggest that CTD phosphatase has the properties expected for a protein phosphatase that catalyzes the conversion of RNAP IIO to RNAP IIA and may play a key role in the transcription cycle of RNAP II.
...
PMID:Purification and characterization of a phosphatase from HeLa cells which dephosphorylates the C-terminal domain of RNA polymerase II. 792 41

Protein kinase(s) and protein phosphatase(s) present in a Xenopus S-100 transcription extract strongly influence promoter-dependent transcription by RNA polymerase I. The protein kinase inhibitor 6-dimethyl-aminopurine causes transcription to increase, while the protein phosphatase inhibitor okadaic acid causes transcription to decrease. Repression is also observed with inhibitor 2, and the addition of extra protein phosphatase 1 stimulates transcription, indicating that the endogenous phosphatase is a type 1 enzyme. Partial fractionation of the system, single-round transcription reactions, and kinetic experiments show that two different steps during ribosomal gene transcription are sensitive to protein phosphorylation: okadaic acid affects a step before or during transcription initiation, while 6-dimethylaminopurine stimulates a process "late" in the reaction, possibly reinitiation. The present results are a clear demonstration that transcription by RNA polymerase I can be regulated by protein phosphorylation.
...
PMID:Identification of two steps during Xenopus ribosomal gene transcription that are sensitive to protein phosphorylation. 811 32

A multicopy genomic library of Saccharomyces cerevisiae (strain FL100) was screened for its ability to suppress conditionally defective mutations altering the 31 kDa subunit (rpc31-236) or the 53 kDa subunit (rpc53-254/424) of RNA polymerase III. In addition to allele-specific suppressors, we identified seven suppressor clones that acted on both mutations and also suppressed several other conditional mutations defective in RNA polymerases I or II. All these clones harbored a complete copy of the SSD1 gene. The same pleiotropic suppression pattern was found with the dominant SSD1-v allele present in some laboratory strains of S. cerevisiae. SSD1-v was previously shown to suppress mutations defective in the SIT4 gene product (a predicted protein phosphatase subunit) or in the regulatory subunit of the cyclic AMP-dependent protein kinase. We propose that the SSD1 gene product modulates the activity (or the level) of the three nuclear RNA polymerases, possibly by altering their degree of phosphorylation.
...
PMID:A general suppressor of RNA polymerase I, II and III mutations in Saccharomyces cerevisiae. 851 Jun 44

HeLa cell nuclei were permeabilized and reconstituted with nuclear extract to identify soluble nuclear factors which play a role in the organization of pre-mRNA splicing factors in the mammalian cell nucleus. Permeabilized nuclei reconstituted with nuclear extract were active in transcription and DNA replication and nuclear speckles containing pre-mRNA splicing factors were maintained over several hours independent of soluble nuclear components. The characteristic rounding up of nuclear speckles in response to inhibition of RNA polymerase II seen in vivo was reproduced in permeabilized cells and was strictly dependent on a catalytic activity present in the nuclear extract. By inhibitor titration experiments and sensitivity to inhibitor 2, this activity was identified as a member of the serine/threonine protein phosphatase 1 family (PP1). Interference with PP1 activity affected the distribution of pre-mRNA splicing factors in transcriptionally active, permeabilized cells, and excess PP1 activity caused increased dephosphorylation of SR proteins in nuclear speckles. These data show that the dynamic reorganization of the mammalian cell nucleus can be studied in permeabilized cells and that PP1 is involved in the rounding up of speckles as well as the overall organization of pre-mRNA splicing factors in the mammalian cell nucleus.
...
PMID:Serine/threonine phosphatase 1 modulates the subnuclear distribution of pre-mRNA splicing factors. 889 62

1. We have used the patch-clamp technique to study modulation of the inwardly rectifying K+ current (IK(IR)) in cultured bovine pulmonary artery endothelial cells (CPAE cells). In whole-cell mode, IK(IR) was defined as the Ba(2+)-sensitive current. In single channel recordings, we observed a strongly inwardly rectifying and K(+)-selective channel with a conductance of 31 +/- 3 pS. 2. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis and functional data suggest that the endothelial IRK is most probably Kir2.1. 3. Intracellular ATP is required to prevent run-down of IRK in whole-cell mode. Single channel activity disappeared in inside-out patches exposed to ATP-free solution and in cell-attached patches on cells exposed to metabolic inhibition (KCN, 2-deoxyglucose). 4. The non-hydrolysable ATP analogues, ATP gamma S and adenylyl imidodiphosphate (AMP-PNP), did not prevent run-down. Run-down did not occur in the presence of okadaic acid, a phosphatase inhibitor, but was enhanced in the presence of protamine, an activator of phosphatase 2A (PP2A). 5. GTP gamma S and AlF4- inhibited IRK, also in the presence of ATP. GTP beta S antagonized the GTP gamma S effect. Pretreatment of the cells with PTX did not affect the GTP gamma S-induced inhibition. Okadaic acid, however, slowed this inhibition. 6. Neither activation of protein kinase A (PKA) nor activation of protein kinase C (PKC) affected IRK. Additionally, neither cytochalasin B nor a high concentration of intracellular Ca2+ affected the time course of IRK run-down. 7. We conclude that run-down of IRK is probably due to dephosphorylation by PP2A. Activation of a PTX-insensitive G protein inhibits this current by a mechanism that is neither mediated via the PKA and PKC pathways nor by intracellular Ca2+, but supposedly by a G protein-dependent activation of a phosphatase.
...
PMID:Modulation of inwardly rectifying potassium channels in cultured bovine pulmonary artery endothelial cells. 940 63


1 2 3 4 5 6 7 Next >>

---