Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.6.1.3 (ATPase)
 65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cyclin-dependent kinase 9 (Cdk9) of fission yeast is an essential ortholog of metazoan positive transcription elongation factor b (P-TEFb), which is proposed to coordinate capping and elongation of RNA polymerase II (Pol II) transcripts. Here we show that Cdk9 is activated to phosphorylate Pol II and the elongation factor Spt5 by Csk1, one of two fission yeast CDK-activating kinases (CAKs). Activation depends on Cdk9 T-loop residue Thr-212. The other CAK-Mcs6, the kinase component of transcription factor IIH (TFIIH)-cannot activate Cdk9. Consistent with the specificities of the two CAKs in vitro, the kinase activity of Cdk9 is reduced approximately 10-fold by csk1 deletion, and Cdk9 complexes from csk1Delta but not csk1+ cells can be activated by Csk1 in vitro. A cdk9(T212A) mutant is viable but phenocopies conditional growth defects of csk1Delta strains, indicating a role for Csk1-dependent activation of Cdk9 in vivo. A cdk9(T212A) mcs6(S165A) strain, in which neither Cdk9 nor Mcs6 can be activated by CAK, has a synthetic growth defect, implying functional overlap between the two CDKs, which have distinct but overlapping substrate specificities. Cdk9 forms complexes in vivo with the essential cyclin Pch1 and with Pcm1, the mRNA cap methyltransferase. The carboxyl-terminal region of Cdk9, through which it interacts with another capping enzyme, the RNA triphosphatase Pct1, is essential. Together, the data support a proposed model whereby Cdk9/Pch1-the third essential CDK-cyclin complex described in fission yeast-helps to target the capping apparatus to the transcriptional elongation complex.
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PMID:Cyclin-dependent kinase 9 (Cdk9) of fission yeast is activated by the CDK-activating kinase Csk1, overlaps functionally with the TFIIH-associated kinase Mcs6, and associates with the mRNA cap methyltransferase Pcm1 in vivo. 1642 35

The coordinated action of histone acetyltransferases (HATs) and ATP-dependent chromatin remodeling enzymes in promoter-dependent transcription initiation represents a paradigm for how epigenetic information regulates gene expression. However, little is known about how such enzymes function during transcription elongation. Here, we investigated the role of RSC, a bromodomain-containing ATPase, in nucleosome transcription in vitro. Purified S. cerevisiae RNA polymerase II (Pol II) arrests at two primary locations on a positioned mononucleosome. RSC stimulates passage of Pol II through these sites. The function of RSC in elongation requires the energy of ATP hydrolysis. Moreover, the SAGA and NuA4 HATs strongly stimulated RSC's effect on elongation. The stimulation correlates closely with acetyl-CoA-dependent recruitment of RSC to nucleosomes. Thus, RSC can recognize acetylated nucleosomes and facilitate passage of Pol II through them. These data support the view that histone modifications regulate accessibility of the coding region to Pol II.
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PMID:RSC exploits histone acetylation to abrogate the nucleosomal block to RNA polymerase II elongation. 1708 96

The role of actin cytoskeleton functional state in glioma C6 cell morphology and calcium signaling was investigated through modification of myosin II activity by blocking Rho-associated kinase with the specific inhibitor Y-27632. Treatment of glioma C6 cells with ROCK inhibitor resulted in actin cytoskeleton reorganization and also in the changed shape and distribution of mitochondria. Changes in the distribution of ER, the main calcium store in glioma C6 cells, were not visible. The inhibition of myosin II activity influences the first phase of calcium signaling evoked by agonist, and both phases of thapsigargin-evoked calcium response. We suggest that the observed increase in Ca2+ release from intracellular stores induced by IP3 formation as well as inhibition of SERCA ATPase is at least in part related to severely affected mitochondria. Enhancement of capacitative calcium entry evoked by thapsigargin is probably associated with the reorganization of the acto-myosin II system. ATP-induced calcium response presents no changes in the second phase. We observed that ATP stimulation of Y-27632 pretreated cells leads to immediate morphological rearrangement of glioma C6 cells. It is a consequence of actin cytoskeleton reorganization: formation of stress fibers and relocation of phosphorylated myosin II to actin filaments. It seems that the agonist-evoked strong calcium signal may be sufficient for myosin II activation and the stress fiber organization. This is the first work showing the dependence between the functional state of the acto-myosin II system and calcium signaling stressing the reversible character of this relationship.
Acta Biochim Pol 2006
PMID:Effect of Rho-associated kinase inhibition on actin cytoskeleton structure and calcium response in glioma C6 cells. 1711 79

CCAAT/enhancer binding proteindelta (C/EBPdelta) gene transcription is highly induced in G(0) growth arrested mammary epithelial cells and "loss of function" alterations in C/EBPdelta have been reported in human breast cancer. To gain a better understanding of the positive and negative factors that control C/EBPdelta gene expression we investigated the role of transcriptional activators, coactivators, repressors, histone modifications, chromatin remodeling and basal transcriptional machinery components in growing and growth arrested HC11 mouse mammary epithelial cells. Growth arrest treatments result in increased STAT3 activation (pSTAT3) and increased C/EBPdelta expression. Co-immunoprecipitation and chromatin immunoprecipitation (ChIP) assays demonstrated that pSTAT3 and Sp1 interact and bind to the transcriptionally active C/EBPdelta promoter. ChIP assays performed under exponentially growing (C/EBPdelta non-expressing) conditions demonstrated that the C/EBPdelta promoter is preloaded with transcriptional activators (Sp1 and CREB) and transcriptional machinery components (TBP and RNA Pol II). In contrast, under G(0) growth arrest (C/EBPdelta expressing) conditions ChIP analysis detected pSTAT3, Sp1, NCoA/SRC1, CBP/p300, pCREB, TBP, and serine 2 phosphorylated Pol II (pPol II) in association with the C/EBPdelta proximal promoter. C/EBPdelta promoter-associated histone post-translational modification analysis revealed histone H3 and H4 acetylation and methylation patterns consistent with a constitutively "open" chromatin conformation. Chromatin remodeling experiments demonstrated that BRG1, the ATPase component of the SWI/SNF chromatin remodeling complex, is required for C/EBPdelta transcription. Finally, C/EBPdelta expression is repressed in proliferating mammary epithelial cells by c-Myc via a mechanism that involves the binding of c-Myc:Max dimers to C/EBPdelta promoter-bound Miz-1. These results provide a molecular model of C/EBPdelta transcriptional regulation under G(0) growth arrest conditions.
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PMID:The mouse C/EBPdelta gene promoter is regulated by STAT3 and Sp1 transcriptional activators, chromatin remodeling and c-Myc repression. 1747 7

The well known and accepted mode of action of cardiac glycosides is inhibition of the ubiquitous plasma membrane Na+, K+-ATPase that leads to increased intracellular Ca2+ ion concentrations. Ca2+ ions play pivotal role in many signaling pathways including those regulating apoptosis. It has been suggested that some forms of cardiac glycosides inhibit proliferation and induce apoptosis in prostate cancer cells in clinically relevant concentrations. It was also found out that the degree to which cardiac glycosides inhibited cancer cell growth was correlated to topoisomerase II-inhibiting activity. Digitoxin at concentrations found in cardiac patients induced levels of DNA-topoisomerase II cleavable complexes similar to etoposide, a topoisomerase II poison widely used in cancer chemotherapy. Cardiac glycosides can also regulate one of the most potent angiogenesis promoting substances, fibroblast growth factor-2 (FGF-2), and may inhibit activation of the transcription factor NF-kappaB. FGF-2 and NF-kappaB are relevant targets for anticancer drugs. There is growing interest in evaluating the oleander products and possibly other cardiac glycosides as antineoplastic agents. The first of these therapies to be developed in the United States is a patented, water-soluble oleander extract called Anvirzel.
Acta Pol Pharm
PMID:Cardiac glycosides in cancer research and cancer therapy. 1751 73

Hsp70s are chaperone proteins that are conserved in evolution and present in all prokaryotic and eukaryotic organisms. In the archaea, which form a distinct kingdom, the Hsp70 chaperones have been found in some species only, including Methanosarcina mazei. Both the bacterial and archaeal Hsp70(DnaK) chaperones cooperate with a GrpE co-chaperone which stimulates the ATPase activity of the DnaK protein. It is currently believed that the archaeal Hsp70 system was obtained by the lateral transfer of chaperone genes from bacteria. Our previous finding that the DnaK and GrpE proteins of M. mazei can functionally cooperate with the Escherichia coli GrpE and DnaK supported this hypothesis. However, the cooperation was surprising, considering the very low identity of the GrpE proteins (26%) and the relatively low identity of the DnaK proteins (56%). The aim of this work was to investigate the molecular basis of the observed interspecies chaperone interaction. Infrared resolution-enhanced spectra of the M. mazei and E. coli DnaK proteins were almost identical, indicating high similarity of their secondary structures, however, some small differences in band position and in the intensity of amide I' band components were observed and discussed. Profiles of thermal denaturation of both proteins were similar, although they indicated a higher thermostability of the M. mazei DnaK compared to the E. coli DnaK. Electrophoresis under non-denaturing conditions demonstrated that purified DnaK and GrpE of E. coli and M. mazei formed mixed complexes. Protein modeling revealed high similarity of the 3-dimensional structures of the archaeal and bacterial DnaK and GrpE proteins.
Acta Biochim Pol 2007
PMID:Structural basis of the interspecies interaction between the chaperone DnaK(Hsp70) and the co-chaperone GrpE of archaea and bacteria. 1756 88

Trichophyton rubrum is a cosmopolitan and anthropophilic fungus able to invade keratinized tissue, causing infection in human skin and nails. This work evaluated the changes in the extracellular pH during its growth in keratin (after 6, 12, 24, 48, 72h and 7 days) at initial pH 5.0. We observed a gradual increase of basal pH under keratin exposure when compared to glucose condition. Also, we identified 576T. rubrum transcripts differentially expressed by subtractive suppression hybridization (SSH) using conidia cultivated for 72h in keratin as tester, and cultivated in glucose as driver. The over-expression of 238 transcripts obtained under keratin condition was confirmed by macro-array dot-blot, revealing 28 unigenes. Putative proteins encoded by these genes showed similarity to fungi proteins involved in basic metabolism, growth and virulence, i.e., transporters ABC-MDR, MFS and ATPase of copper, NIMA interactive protein, Gag-Pol polyprotein, virulence factors serine-protease subtilisin and metalloprotease, cytochrome P450, GlcN-6-phosphate deaminase and Hsp30. The upregulation of T. rubrum genes encoding subtilisin, metalloprotease and Gag-Pol polyprotein was also validated by northern blot. The results of this study provide the first insight into genes differentially expressed during T. rubrum grown in keratin that may be involved in fungal pathogenesis.
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PMID:Isolation of transcripts over-expressed in human pathogen Trichophyton rubrum during growth in keratin. 1759 Mar 7

Blocked replication forks often need to be processed by recombination proteins prior to replication restart. In Escherichia coli, the UvrD repair helicase was recently shown to act at inactivated replication forks, where it counteracts a deleterious action of RecA. Using two mutants affected for different subunits of the polymerase III holoenzyme (Pol IIIh), we show here that the anti-RecA action of UvrD at blocked forks reflects two different activities of this enzyme. A defective UvrD mutant is able to antagonize RecA in cells affected for the Pol IIIh catalytic subunit DnaE. In this mutant, RecA action at blocked forks specifically requires the protein RarA (MgsA). We propose that UvrD prevents RecA binding, possibly by counteracting RarA. In contrast, at forks affected for the Pol IIIh clamp (DnaN), RarA is not required for RecA binding and the ATPase function of UvrD is essential to counteract RecA, supporting the idea that UvrD removes RecA from DNA. UvrD action on RecA is conserved in evolution as it can be performed in E. coli by the UvrD homologue from Bacillus subtilis, PcrA.
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PMID:UvrD controls the access of recombination proteins to blocked replication forks. 1764 84

In circulation, platelets may come into contact with both exogenous (cardiac glycoside treatment) and endogenously produced inhibitors of Na+/K(+)-ATPase. We examined whether blocking of platelet Na+/K(+)-ATPase by ouabain results in generation of procoagulant activity. It was shown that an in vitro treatment of platelets with ouabain (20-200 microM for 20 to 60 min) is associated with an intracellular accumulation of sodium ([Na+](i)), generation of a weak calcium signal, and expression of procoagulant activity. The ouabain-induced procoagulant response was dose- and time-related, less pronounced than that evoked by collagen and similar to that produced by gramicidin, not affected by EDTA or aspirin, and strongly reduced in the absence of extracellular Na+ or by hyperosmolality. Flow cytometry studies revealed that ouabain treatment results in a unimodal left shift in the forward and side scatter of the entire platelet population indicating morphological changes of the plasma membrane. The shift was dose related, weaker than that evoked by collagen and similar to that produced by gramicidin. Ouabain-treated platelets express phosphatidylserine (PS). The ouabain-evoked PS expression was dose- and time-dependent, weaker than that produced by collagen and similar to that evoked by gramicidin. Electronic cell sizing measurements showed a dose-dependent increase in mean platelet volume upon treatment with ouabain. Hypoosmotically-evoked platelet swelling resulted in the appearance of procoagulant activity. Thromboelastography measurements indicate that, in whole blood, nanomolar (50-1000 nM, 15 min) concentrations of ouabain significantly accelerate the rate of clot formation initiated by contact and high extracellular concentration of calcium. We conclude that inefficiently operating platelet Na+/K(+)-ATPase results in a rise in [Na+](i). An increase in [Na+](i) and the swelling associated with it may produce PS exposure and a rise in membrane curvature leading to the generation of a procoagulant activity.
Acta Biochim Pol 2007
PMID:The involvement of Na+/K(+)-ATPase in the development of platelet procoagulant response. 1765 2

Myosin can be precipitated from soluble fraction under different assay conditions. This paper describes a new method for precipitating myosin V from rat brain soluble fraction. Brains were homogenized in 50 mM imidazole/HCl buffer, pH 8.0, containing 10 mM EDTA/EGTA, 250 mM sucrose, 1 mM DTT and 1 mM benzamidine, centrifuged at 45000 x g for 40 min and the supernatant was frozen at -20 degrees C. Forty-eight hours later, the supernatant was thawed, centrifuged at 45000 x g for 40 min and the precipitate was washed in 20 mM imidazole buffer pH 8.0. SDS/PAGE analysis showed four polypeptides in the precipitate: 205, 150, 57 and 43 kDa. The precipitate presented high Mg(2+)-ATPase activity, which co-purifies with p205. This polypeptide was recognized by a specific myosin V antibody and was proteolised by calpain, generating two stable polypeptides: p130 and p90. The Mg(2+)-ATPase activity was not stimulated by calcium in both the absence and presence of exogenous calmodulin and the K+/EDTA-ATPase activity represented 25% of the Mg(2+)-ATPase activity. In this work, myosin V from rat brain was precipitated by freezing the soluble fraction and was co-purificated with a 45 kDa polypeptide.
Acta Biochim Pol 2007
PMID:A new method to precipitate myosin V from rat brain soluble fraction. 1788 23


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