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:3.6.1.3 (ATPase)
65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two beta-actin mutants, one with proline 38 replaced with alanine (P38A) and the other with cysteine-374 replaced with serine (C374S), as well as the wild-type beta-actin, were expressed in the yeast, S. cerevisiae, purified to homogeneity, and analyzed in vitro for polymerizability and interaction with DNase I, myosin, and profilin. Both mutations interfered with the polymerization of the actin, and with its interaction with myosin. The C374S mutation had the most pronounced effect; it reduced the polymerizability of the actin, abolished its binding to profilin, and filaments containing this mutation moved at reduced rates in the in vitro 'motility assay'. The ATPase activity measured in solutions containing myosin subfragment 1 was similar for both the mutant and wild-type actins.
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PMID:Mutations in beta-actin: influence on polymer formation and on interactions with myosin and profilin. 835 91

We have studied the long-term effects of lithium on neuronal morphology and the functional expression of phospholipase C-coupled m3-muscarinic acetylcholine receptors (mAChRs) in cerebellar granule cells. There was a biphasic dose-dependent effect on cell morphology following treatment with lithium for 7 days. At low concentrations (< or = 2 mM), this drug elicited an increase in the number and thickness of connecting nerve fibers, and the size of neuronal aggregates. At high concentrations (5-10 mM), lithium induced a severe deterioration of cell morphology, which ultimately resulted in neuronal death. Carbachol-induced phosphoinositide (PI) turnover was similarly affected by lithium treatment with a significant potentiation at concentrations up to 2 mM and a marked inhibition at doses higher than 5 mM due to lithium-induced neurotoxicity. The biphasic effect on mAChR-mediated PI hydrolysis was associated with corresponding changes in the maximal extent of carbachol-induced inositol phosphate accumulation, and was accompanied by similar changes in [3H]N-methyl-scopolamine binding to mAChRs and the levels of mRNAs for m3-mAChR and c-Fos. The up-regulation of m3-mAChR mRNA induced by low concentrations of lithium was associated with a down-regulation of m2-mAChR mRNA and no change in either total RNA or beta-actin mRNA. Lithium's effects on m2- and m3-mAChR mRNAs were time-dependent, requiring a pretreatment time of > or = 3 days. The biphasic effect was also demonstrated by the binding of [3H]ouabain to Na+, K(+)-ATPase, which was shown to be a convenient method for quantifying viable neurons.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Long-term biphasic effects of lithium treatment on phospholipase C-coupled M3-muscarinic acetylcholine receptors in cultured cerebellar granule cells. 838 5

The cortical collecting duct (CCD) adapts to disturbances of acid/base balance by adjusting the direction and magnitude of its HCO3 transport. The molecular events involved in this adaptation are incompletely understood, but it seems that adaptation is accompanied by changes in the activity and intracellular distribution of the vacuolar H-ATPase. The goal of this study was to examine the effects of metabolic acidosis and alkali load on the expression of the mRNA encoding the 31 kD subunit of the vacuolar H-ATPase in rabbit CCD cells. Pairs of rabbits received either a NH4Cl load or a NaHCO3 load for 16 hours, resulting in a urinary pH of 5.53 +/- 0.38 and 8.42 +/- 0.10, respectively. CCD cells were isolated by immunodissection and mRNA levels of the H-ATPase 31 kD subunit and of beta-actin were determined from the same cDNA samples by quantitative RT-PCR. H-ATPase mRNA levels were significantly higher in CCD cells from acidotic than alkali-loaded rabbits (2.51 +/- 1.3 vs. 0.65 +/- 0.2; P < 0.05). Similar differences in the H-ATPase 31 kD subunit mRNA levels were observed by Northern blotting. beta-actin mRNA levels were comparable in CCD cells of the two groups. The distribution of the H-ATPase 31 kD subunit mRNA was determined among the three cell types of the CCD, that is in alpha- and beta-intercalated cells (alpha-ICC and beta-ICC) and principal cells (PC) isolated by fluorescence-activated cell sorting. The level of expression was comparable in alpha-ICCs and beta-ICCs, whereas PCs contained very low levels of H-ATPase mRNA. In both alpha-ICC and beta-ICC the levels of the 31 kD H-ATPase mRNA were significantly higher in acidotic than in alkali-loaded rabbits. These results indicate that in the rabbit CCD changes in acid/base balance not only regulate the subcellular distribution of the vacuolar H-ATPase but also alter its expression, at least at the mRNA level.
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PMID:Effect of acid/base balance on H-ATPase 31 kD subunit mRNA levels in collecting duct cells. 854 98

The osteoclast is a cell type that is highly specialized for its bone resorption function. In order to decipher the numerous biochemical functions of osteoclasts, a description of the gene expression profile of osteoclasts would be beneficial. We have sought to identify genes that are highly expressed in osteoclasts by partially sequencing 194 randomly chosen cDNA clones from a representative rabbit osteoclast cDNA library. Comparison to nucleic acid and protein sequence databases indicates that 135 of these cDNAs are identical to or homologous to known mammalian genes. Reverse transcription-polymerase chain reaction (RT-PCR) assays with microisolated osteoclasts were used to verify the osteoclast expression of some of these genes. Fifty-nine cDNAs, including two abundantly expressed species, have no significant similarity to the sequence databases and likely represent novel genes. The most abundant of the osteoclast expressed genes encode cofilin and the vacuolar H(+)-ATPase 16 kd subunit. Each were represented at a frequency of 4.1% of the clones in the library (95% confidence interval = 2.4-6.6%). The high expression of these gene products is consistent with the high motility of osteoclasts and their very active hydrogen ion secretion. Other abundantly expressed sequences include beta-actin (95% C.I. = 2.0-6.0%), creatine kinase B (95% C.I. = 1.2-4.9%), c-fms and ribosomal protein L18 (95% C.I. = 0.8-4.3%), and cathepsin-OC2, cyclophilin, delta-aminolevulinate synthetase, 16S mitochondrial rRNA, and two novel gene sequences (95% C.I. = 0.5-3.6%).
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PMID:Osteoclast molecular phenotyping by random cDNA sequencing. 855 18

Recent studies have shown that bafilomycin A1-sensitive vacuolar-type H(+)-ATPase (V-ATPase) plays important roles in cell growth and differentiation. However, there is no published study that has focused on the expression of V-ATPase in human tumour tissues. This study was designed to examine the mRNA and protein levels for the 16 kilodalton (kDa) proteolipid of V-ATPase in human pancreatic carcinoma tissues. We first investigated the mRNA level for V-ATPase in six cases of invasive pancreatic cancers and two normal pancreases, using reverse transcription-polymerase chain reaction technique. Then, we examined immunohistochemically the level of V-ATPase protein in 49 pancreatic cancers and ten benign cystic neoplasms of the pancreas, using antisera raised against the 16 kDa proteolipid. There was a notable difference in the level of V-ATPase mRNA between normal and pancreatic carcinoma tissues, with no evident difference in the expression of the beta-actin gene. Immunohistochemically, 42 out of 46 invasive ductal cancers (92%) displayed a mild to marked immunoreactivity for V-ATPase in the cytoplasm, whereas neither non-invasive ductal cancers nor benign cystic neoplasms expressed detectable immunoreactive proteins. These findings suggest that the overexpression of V-ATPase protein is characteristic of invasive pancreatic tumours. V-ATPase may play some crucial roles in tumour progression.
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PMID:Expression of 16 kDa proteolipid of vacuolar-type H(+)-ATPase in human pancreatic cancer. 866 21

Chronic potassium restriction leads to active potassium reabsorption in the late distal nephron and collecting duct, segments known to express "gastric" H(+)-K(+)-adenosinetriphosphatase (H(+)-K(+)-ATPase) alpha-subunit mRNA. In this study, the cellular distribution and relative abundance of mRNA encoding this isoform were examined in kidneys of normal and potassium-deprived (2 wk) rats. In situ hybridization with isoform-specific cRNA probes demonstrated prominent expression of this gene in the connecting segment (CNT), entire collecting duct, and renal papillary surface epithelium in a comparable distribution in both groups of rats. Hypertrophy of the outer medullary collecting ducts in the inner stripe of potassium-restricted rats was observed. Competitive polymerase chain reaction analysis revealed twofold greater levels of gastric H(+)-K(+)-ATPase alpha-subunit mRNA (normalized to the level of beta-actin mRNA) in the cortex, but roughly comparable levels in the outer and inner medulla, of potassium-restricted rats compared with controls. These data suggest that chronic potassium restriction results in modestly enhanced renal cortical expression of the gastric H(+)-K(+)-ATPase alpha-subunit gene and that this isoform may participate in potassium conversation by the CNT and cortical collecting duct during potassium deprivation.
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PMID:Effects of chronic hypokalemia on renal expression of the "gastric" H(+)-K(+)-ATPase alpha-subunit gene. 896 34

Although the kidney plays the major role in the regulation of systemic K+ homeostasis, the colon also participates substantively in K+ balance. The colon is capable of both K+ absorption and secretion, the magnitude of which can be modulated in response to dietary K+ intake. The H(+)-K(+)-adenosinetriphosphatase (H(+)-K(+)-ATPase) has been proposed as a possible mediator of K+ absorption in distal colon, but inhibitor profiles obtained in recent studies suggest that two, and perhaps more, distinct H(+)-K(+)-ATPase activities may be present in mammalian distal colon. We have developed highly specific probes for the catalytic alpha-subunits of colonic and gastric H(+)-K(+)-ATPase, alpha 1-Na(+)-K(+)-ATPase, and beta-actin, which were used in Northern analysis of total RNA from whole distal colon and stomach obtained from one of three experimental groups of rats: 1) controls, 2) chronic dietary K+ depletion, and 3) chronic metabolic acidosis. The probe for the colonic but not the gastric H(+)-K(+)-ATPase alpha-isoform hybridized to distal colon total RNA in all groups. A significant increase in colonic H(+)-K(+)-ATPase mRNA abundance was observed in response to chronic dietary K+ depletion but not to chronic metabolic acidosis. The alpha 1-isoform of Na(+)-K(+)-ATPase, which is also expressed in distal colon, did not respond consistently to either chronic dietary K+ depletion or chronic metabolic acidosis. The gastric probe did not hybridize to total RNA from distal colon but, as expected, hybridized to total stomach RNA. However, the abundance of gastric H(+)-K(+)-ATPase or Na(+)-K(+)-ATPase in stomach was not altered consistently by either chronic dietary K+ depletion or metabolic acidosis. Under the conditions of this study, it appears that the mRNA encoding the colonic alpha-isoform is upregulated by chronic dietary K+ restriction, a condition shown previously to increase K+ absorption in the distal colon.
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PMID:Effect of chronic hypokalemia on H(+)-K(+)-ATPase expression in rat colon. 903 45

On the basis of paradigms in development wherein discrete transcriptional events are pivotal regulatory steps, we tested the hypothesis that transcriptional sodium (Na+)-response mechanisms are involved in in vivo Na+-induced responses relevant to normal (homeostatic) and pathophysiological (salt-sensitive hypertension) conditions. We used Na,K-ATPase alpha-subunit genes as molecular probes and the Na+ ionophore monensin to induce a dose-specific incremental increase in [Na+]i in rat A10 embryonic aortic smooth muscle cells. RNA blot analysis of rat A10 cells revealed a dose-specific (0.022 to 30 micromol/L monensin) upregulation of alpha1-, alpha2-, and beta1-subunit Na,K-ATPase RNA levels. Control beta-actin and alpha-tropomyosin RNA levels did not change. With the use of chloramphenicol acetyltransferase (CAT) as reporter gene, CAT assays of rat alpha1[-1288]CAT and human alpha2[-798]CAT promoter constructs exhibited induction of CAT activity in monensin (10 micromol/L)-treated A10 cells compared with untreated A10 cells. Promoter deletion constructs for rat alpha1[-1288]CAT defined a positive Na+-response regulatory region within -358 to -169 that is distinct from the basal transcriptional activation region of -155 to -49 previously defined. Similarly, a positive Na+-response regulatory region is delimited to within -301 in the human alpha2 Na,K-ATPase 5' flanking region. Analysis of transgenic TgH alpha2[-798]CAT rats demonstrated sodium activation of human alpha2[-798]CAT transgene expression in aorta parallel to observations made in rat A10 aortic tissue culture cells. Southwestern blot analysis of nuclear extracts from monensin (10 micromol/L)-treated and control untreated A10 cells revealed a nuclear DNA binding protein (approximately 95 kD) that is upregulated by increased [Na+]i. These data provide initial characterization of a transcriptional Na+-response mechanism delimiting a positive Na+-response regulatory region in two target genes (alpha1 and alpha2 Na,K-ATPase) as well as detection of a Na+-response nuclear DNA binding protein. The in vitro data are corroborated by in vivo experimental and transgenic promoter expression studies, thus validating the biological relevance of the observations.
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PMID:Characterization of a sodium-response transcriptional mechanism. 926 Sep 79

To assess whether metabolic acidosis per se regulates rBSC-1, the rat medullary thick ascending limb (MTAL) apical Na+-K+(NH4+)-2Cl- cotransporter, rat MTALs were incubated for 16 h in an acid 1:1 mixture of Ham's nutrient mixture F-12 and Dulbecco's modified Eagle's medium. Cotransport activity was estimated in intact cells and membrane vesicles by intracellular pH and 22Na+ uptake measurements, respectively; rBSC-1 protein was quantified by immunoblotting analysis and mRNA by quantitative reverse transcription-polymerase chain reaction. As compared with incubation at pH approximately 7.35, acid incubation (pH approximately 7.10) up-regulated by 35-100% rBSC-1 transport activity in cells and membrane vesicles, and rBSC-1 protein and mRNA abundance. In contrast, acid incubation did not alter alkaline phosphatase and Na+/K+-ATPase enzyme activities or beta-actin protein abundance. After 3 h of in vivo chronic metabolic acidosis (CMA) rBSC-1 mRNA abundance increased in freshly harvested MTALs, which was accompanied after 1-6 days of CMA with enhanced rBSC-1 protein abundance. These results demonstrate that both in vivo and in vitro CMA stimulate rBSC-1 expression, which would contribute to the adaptive increase in MTAL absorption and urinary excretion of NH4+ in response to CMA.
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PMID:Stimulation by in vivo and in vitro metabolic acidosis of expression of rBSC-1, the Na+-K+(NH4+)-2Cl- cotransporter of the rat medullary thick ascending limb. 983 54

Lymphocyte activation is accompanied by visible changes in chromatin structure. We find that antigen receptor signaling induces the rapid association of the BAF complex with chromatin. PIP2, which is regulated by activation stimuli, is sufficient in vitro to target the BAF complex to chromatin, but it has no effect on related chromatin remodeling complexes containing SNF2L or hISWI. Purification and peptide sequencing of the subunits of the complex revealed beta-actin as well as a novel actin-related protein, BAF53. beta-actin and BAF53 are required for maximal ATPase activity of BRG1 and are also required with BRG1 for association of the complex with chromatin/matrix. This work indicates that membrane signals control the activity of the mammalian SWI/SNF or BAF complex and demonstrates a direct interface between signaling and chromatin regulation.
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PMID:Rapid and phosphoinositol-dependent binding of the SWI/SNF-like BAF complex to chromatin after T lymphocyte receptor signaling. 984 65


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