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: UNIPROT:P67775 (alpha isoform)
797 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two isoforms of the sarcoplasmic reticulum Ca2+ release channel (ryanodine receptor or RYR) are expressed together in the skeletal muscles of most vertebrates. We have studied physiological properties of the two isoforms (alpha and beta) by comparing SR preparations from specialized fish muscles that express the alpha isoform alone to preparations from muscles containing both alpha and beta. Regulation of channel activity was assessed through [3H]ryanodine binding and reconstitution into planar lipid bilayers. Distinct differences were observed in the calcium-activation and -inactivation properties of the two isoforms. The fish alpha isoform, expressed alone in extraocular muscles, closely resembled the rabbit skeletal muscle RYR. Maximum [3H]ryanodine binding and maximum open probability (Po) of the alpha RYR were achieved from 1 to 10 microM free Ca2+. Millimolar Ca2+ reduced [3H]ryanodine binding and Po close to zero. The beta isoform more closely resembled the fish cardiac RYR in Ca2+ activation of [3H]ryanodine binding. The most prominent difference of the beta and cardiac isoforms from the alpha isoform was the lack of inactivation of [3H]ryanodine binding and Po by millimolar free Ca2+. Differences in activation of [3H]ryanodine binding by adenine nucleotides and inhibition by Mg2+ suggest that the beta and cardiac RYRs are not identical, however. [3H]ryanodine binding by the alpha RYR was selectively inhibited by 100 microM tetracaine, whereas cardiac and beta RYRs were much less affected. Tetracaine can thus be used to separate the properties of the alpha and beta RYRs in preparations in which both are present. The distinct physiological properties of the alpha and beta RYRs that are present together in most vertebrate muscles support models of EC coupling incorporating both directly coupled and Ca(2+)-coupled channels within a single triad junction.
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PMID:Physiological differences between the alpha and beta ryanodine receptors of fish skeletal muscle. 769

The protein kinase C (PKC) family of enzymes is comprised of at least nine isoforms that vary with respect to co-factor dependence, cellular distribution and substrate specificity. Using specific antibodies for alpha, beta, gamma, delta, epsilon, zeta and eta PKC isoforms, and Western blot analysis, we found that alpha and zeta PKC are expressed in gastric chief cells. We then used these methods to examine the effects of carbamylcholine, a cholinergic agonist that increases cellular calcium and diacylglycerol concentrations, and PMA, a phorbol ester that activates PKC, on the subcellular distribution of these isoforms. Carbamylcholine and PMA caused an increase in membrane-associated alpha PKC, but did not alter the subcellular distribution of zeta PKC. Comparison of the dose-response curves for carbamylcholine-induced pepsinogen secretion and alpha PKC membrane-association indicates that PKC translocation is not required for carbamylcholine-induced secretion. Nevertheless, maximal carbachol-induced secretion occurs at concentrations that also cause translocation of the alpha isoform. Whereas treatment of chief cells with PMA (300 nM) for 4 h down-regulated levels of alpha PKC by 61%, there was no change in the levels of zeta PKC. Separation of the two PKC isoforms in chief cell lysates by DEAE-column chromatography revealed that kinase activity in fractions containing the alpha isoform was increased more than 3-fold by calcium and lipids. In contrast, kinase activity in fractions containing the zeta isoform was not altered. In gastric chief cells, translocation and activation of alpha PKC occurs in response to agonist-induced increases in calcium and diacylglycerol. Zeta PKC may be involved in the regulation of basal pepsinogen secretion.
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PMID:Protein kinase C expression and translocation in dispersed chief cells from guinea-pig stomach. 780 15

To investigate the role of protein kinase C (PKC) in the growth of astrocytic brain tumors, human glioblastoma cell line U-87 was stably transfected with the antisense complementary deoxyribonucleic acid encoding PKC alpha. The effect of selectively down-regulating the alpha isoform on other PKC isoforms, as well as serum-dependent proliferation and in vivo tumorigenicity, was determined. U-87 cells expressed high levels of PKC alpha and lesser amounts of the gamma, epsilon, and zeta isoforms, and a similar PKC isoform pattern was observed in two other human glioblastoma cell lines. Expression of the antisense PKC alpha complementary deoxyribonucleic acid resulted in no detectable PKC alpha by immunoblotting and a 95% reduction in total Ca2+/phospholipid-dependent PKC activity. U-87 cells expressing antisense PKC alpha exhibited an increase in doubling time in vitro, less serum-dependent growth, and reduced sensitivity to a selective PKC inhibitor, Ro 31-8220. The transplantation of U-87 cells expressing antisense PKC alpha into nude mice resulted in no tumor formation. These observations suggest that the inhibition of PKC alpha may be an important chemotherapeutic target for arresting the growth of glioblastomas.
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PMID:Antisense expression of protein kinase C alpha inhibits the growth and tumorigenicity of human glioblastoma cells. 783 40

The catalytic subunit of the major protein phosphatase associated with bovine cardiac myofibrils was purified to homogeneity. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of the enzyme revealed only one band with an apparent molecular weight of 37,000. On gel filtration chromatography, the phosphatase activity and the protein co-eluted as a single peak with an apparent molecular weight of 37,000. The purified enzyme was identified as the catalytic subunit of protein phosphatase 1, as determined by sensitivity to inhibitor 1, inhibitor 2, okadaic acid and by specific immunostaining. Evidence obtained with specific antipeptide antibodies demonstrated that this myofibril protein phosphatase was predominantly the alpha isoform of protein phosphatase 1. The purified catalytic subunit was completely inactive. It was activated by pretreatment with Co2+/trypsin in the presence of high ionic strength. Treatment with trypsin alone did not activate the latent enzyme. The enzyme was also activated by Co2+ or Mn2+ alone but not by Ca2+, Mg2+, Ni2+, Cu2+ or Zn2+. Activation of the enzyme was not reversed by removal of Co2+, but Mn(2+)-activated phosphatase activity was partially reversed when Mn2+ was removed. The catalytic subunit could form a 1:1 complex with inhibitor 2 in vitro. The resulting holoenzyme was also activated by pretreatment with Co2+. Since phosphatase 1 alpha is the major phosphatase associated with cardiac myofibril, it is suggested that it is responsible for the dephosphorylation of myosin and other myofibril phosphoproteins.
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PMID:A latent form of protein phosphatase 1 alpha associated with bovine heart myofibrils. 808 38

Protein kinase C activity was detected in the cytosolic fraction of quiescent parotid acinar cells; the particulate fraction contained a much smaller proportion of the enzyme. Protein kinase C activity was increased in the membrane fraction and decreased in the cytosol after exposure of intact cells to phorbol 12-myristate 13-acetate (PMA) or the muscarinic-receptor agonist carbachol. The effect of PMA was potentiated by a subthreshold concentration of ionomycin. Immunoblot analysis with anti-protein kinase C antibodies revealed that the protein kinase C-alpha isoform is expressed in rat parotid cells. Other Ca(2+)-dependent isoforms were not detected. Further, agonist stimulation caused the redistribution of protein kinase C-alpha from cytosol to a membrane fraction. Agonists may promote parotid acinar cell activity, including amylase secretion, by increasing the affinity of protein kinase C-alpha for the membrane fraction, presumably via a rise in Ca2+ and diacylglycerol derived from polyphosphoinositide hydrolysis.
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PMID:Translocation of the alpha-isozyme of protein kinase C during stimulation of rat parotid acinar cells by phorbol ester and carbachol. 814 66

Heterotrimeric G proteins consisting of alpha, beta, and gamma subunits couple sensory, hormone, and neurotransmitter receptors to intracellular and transmembrane effectors. Several splicing variants of the GS (the G protein that stimulates adenylyl cyclase) alpha subunit (GS alpha) have been described. Some of these couple receptors to stimulation of adenylyl cyclase and Ca2+ channels, whereas others encode truncated proteins whose functions are not currently defined. We describe a 1321N1 human astrocytoma cDNA clone for a novel GS alpha isoform isolated from astrocytoma cells (G(astro)) that is identical to GS alpha-1 with the exception of a novel 5' sequence extending into the previously described exon 1 of GS alpha, a single base change, and an alternative polyadenylation site. Analysis by northern blotting and reverse transcription/PCR confirms the presence of an mRNA corresponding to this cDNA in astrocytoma cells. Additional northern analysis indicates that G(astro) recognizes two novel GS alpha mRNAs in the rat: a 2.0-kb mRNA expressed only in neural and neuroendocrine tissues and a 1.8-kb mRNA that is ubiquitously expressed. Functional analysis of G(astro) is complicated by the apparent insertion of alphoid satellite DNA into the transcription unit. The resulting cDNA encodes a truncated protein that may be translated from the methionine in exon 2 as previously described.
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PMID:Isolation and characterization of a novel cDNA which identifies both neural-specific and ubiquitously expressed GS alpha mRNAs. 833 49

Na,K-ATPase (Na,K-pump) plays an important role in the regulation of intracellular ion composition. The purpose of this study is to determine whether Na+ regulates the levels of mRNA coding for Na,K-ATPase alpha and beta subunits in cultured neonatal rat cardiocytes. We measured intracellular Na+ levels ([Na+]i) in cardiocytes using a Na(+)-sensitive fluorescence dye (SBFI). 1 mM ouabain caused a significant increase in [Na+]i in cardiocytes; from 12.8 +/- 0.3 to 28.8 +/- 1.8 mM. Exposure of cardiocytes to 1 mM ouabain resulted in a three- to fourfold increase in alpha 1, alpha 2, and alpha 3 mRNA accumulation, and an approximate two-fold increase in beta 1 mRNA accumulation. A maximum elevation was reached at 60 min in both cases. The ouabain-induced alpha 1 mRNA accumulation was still observed in the Ca(2+)-free culture medium. Exposure of cardiocytes to 10 microM monensin in the absence of extracellular Ca2+ also resulted in a threefold increase in alpha 1 mRNA accumulation. The increased alpha 1 mRNA expression by 1 mM ouabain was associated with a fourfold increase in alpha 1 subunit protein accumulation. Transfection experiments with chimeric plasmids containing 5'-flanking sequences of alpha 1, alpha 2, and alpha 3 isoform genes and a luciferase reporter gene revealed that 1 mM ouabain caused a twofold increase in luciferase activity in each alpha system. These results suggest that Na+ directly regulates Na,K-ATPase gene expression in cardiocytes. The transfection study further supports the premise that Na(+)-responsive elements are located within the 5'-flanking sequences of each alpha isoform gene.
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PMID:Regulation of Na,K-adenosine triphosphatase gene expression by sodium ions in cultured neonatal rat cardiocytes. 840 40

The gene for the alpha isoform of Ca2+/calmodulin-dependent kinase II (alpha CaMKII) codes for a multifunctional protein kinase that is found exclusively in the brain. Here we show that in skeletal muscle, an alternative nonkinase product, hereafter referred to as alpha KAP (alpha CaMKII association protein), is expressed from the same gene. alpha KAP consists of a C-terminal region that is identical to the association domain of alpha CaMKII, with the exception of 11 amino acids inserted in the variable region. The N-terminal sequence of alpha KAP is highly hydrophobic and not present in any known CaMKII protein. The catalytic and regulatory domains of alpha CaMKII are missing in alpha KAP. Analysis of the exon-intron structure revealed that the alpha KAP transcript is derived from the alpha CaMKII gene by alternative promoter usage and RNA splicing. The transcriptional start site of alpha KAP mRNA is located within an intron of the alpha CaMKII gene. Therefore, the relationship between alpha KAP and alpha CaMKII is that of a gene within a gene. Immunostaining using anti-alpha KAP antibodies suggests that alpha KAP is associated with sarcomeres of skeletal muscle fibers. On the basis of its primary structure and specific location, the possible function of alpha KAP as an anchoring protein for CaMKII is discussed.
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PMID:An alternative, nonkinase product of the brain-specifically expressed Ca2+/calmodulin-dependent kinase II alpha isoform gene in skeletal muscle. 852 7

Cyclic ADP-ribose (cADPR) is generated in pancreatic islets by glucose stimulation, serving as a second messenger for Ca2+ mobilization from the endoplasmic reticulum for insulin secretion (Takasawa, S., Nata, K., Yonekura, H., and Okamoto, H. (1993) Science 259, 370-373). In the present study, we observed that the addition of calmodulin (CaM) to rat islet microsomes sensitized and activated the cADPR-mediated Ca2+ release. Inhibitors for CaM-dependent protein kinase II (CaM kinase II) completely abolished the glucose-induced insulin secretion as well as the cADPR-mediated and CaM-activated Ca2+ mobilization. Western blot analysis revealed that the microsomes contain the alpha isoform of CaM kinase II but do not contain CaM. When the active 30-kDa chymotryptic fragment of CaM kinase II was added to the microsomes, fully activated cADPR-mediated Ca2+ release was observed in the absence of CaM. These results along with available evidence strongly suggest that CaM kinase II is required to phosphorylate and activate the ryanodine-like receptor, a Ca2+ channel for cADPR as an endogenous activator, for the cADPR-mediated Ca2+ release.
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PMID:Requirement of calmodulin-dependent protein kinase II in cyclic ADP-ribose-mediated intracellular Ca2+ mobilization. 853 Apr 41

Phospholipase D (PLD) which was partially purified from membranes of porcine brain could be stimulated by multiple cytosolic components; these included ADP-ribosylation factor (Arf) and RhoA, which required guanine nucleotides for activity, and an unidentified factor which activated the enzyme in a nucleotide-independent manner (Singer, W. D., Brown, H. A., Bokoch, G. M., and Sternweis, P. C. (1995) J. Biol. Chem. 270, 14944-14950). Here, we report purification of the latter factor, its identification as the alpha isoform of protein kinase C (PKCalpha), and characterization of its regulation of PLD activity. Stimulation of PLD by purified PKCalpha or recombinant PKCalpha (rPKCalpha) occurred in the absence of any nucleotide and required activators such as Ca2+ or phorbol ester. This action was synergistic with stimulation of PLD evoked by either Arf or RhoA. Dephosphorylation of rPKC alpha with protein phosphatase 1 or 2A resulted in a loss of its kinase activity, but had little effect on its ability to stimulate PLD either alone or in conjunction with Arf. Staurosporine inhibited the kinase activity of PKCalpha without affecting activation of PLD. Finally, gel filtration of PKCalpha that had been cleaved with trypsin demonstrated that stimulatory activity for PLD coeluted with the regulatory domain of the enzyme. These data indicate that PKC may regulate signaling events through direct molecular interaction with downstream effectors as well as through its well characterized catalytic modification of proteins by phosphorylation.
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PMID:Regulation of phospholipase D by protein kinase C is synergistic with ADP-ribosylation factor and independent of protein kinase activity. 862 5


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