Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Sorcin is a widely expressed, 22-kDa Ca2+-binding protein initially identified in multidrug-resistant cells. In the heart, sorcin localizes to the dyadic junctions of transverse tubules and sarcoplasmic reticulum and coimmunoprecipitates with the Ca2+ release channel/ryanodine receptor (RyR) (Meyers, M. B., Pickel, V. M., Sheu, S.-S., Sharma, V. K., Scotto, K. W., and Fishman, G. I. (1995) J. Biol. Chem. 270, 26411-26418). We have investigated a possible functional interaction between sorcin and cardiac RyR using purified recombinant sorcin in [3H]ryanodine binding experiments and single channel recordings of RyR. The open probability of single RyR was decreased significantly by the addition of sorcin to the cytoplasmic side of the channel (IC50 approximately 480 nM). In addition, sorcin completely inhibited [3H]ryanodine binding with an IC50 approximately 700 nM. Inhibition occurred over a wide range of [Ca2+], and sorcin-modulated RyR remained Ca2+-dependent. Furthermore, caffeine-activated RyRs were also inhibited by sorcin at low [Ca2+] (pCa 7), suggesting that Ca2+ is not an obligatory factor for sorcin inhibition of RyR. Comparisons of these inhibitory effects with those of calmodulin and calpain, proteins structurally related to sorcin, suggested that the interaction of sorcin with cardiac RyR was distinct from and independent of either of these modulatory proteins. Phosphorylation of sorcin with the catalytic subunit of protein kinase A significantly decreased the ability of sorcin to modulate RyR. These results suggest that sorcin may modulate RyR function in a normal cell environment and that the level of modulation is in turn influenced by signaling pathways that increase protein kinase A activity.
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PMID:Modulation of cardiac ryanodine receptors by sorcin. 931 52

Stimulation of beta-adrenergic receptors activates type I and II cyclic AMP-dependent protein kinase A, resulting in phosphorylation of various proteins in the heart. It has been proposed that PKA II compartmentalization by A-kinase-anchoring proteins (AKAPs) regulates cyclic AMP-dependent signaling in the cell. We investigated the expression and localization of AKAP100 in adult hearts. By immunoblotting, we identified AKAP100 in adult rat and human hearts, and showed that type I and II regulatory (RI and II) subunits of PKA are present in the rat heart. By immunofluorescence and confocal microscopy of rat cardiac myocytes and cryostat sections of rat left ventricle papillary muscles, we localized AKAP100 to the nucleus, sarcolemma, intercalated disc, and at the level of the Z-line. After double immunostaining of transverse cross-sections of the papillary muscles with AKAP100 plus alpha-actinin-specific antibodies or AKAP100 plus ryanodine receptor-specific antibodies, confocal images showed AKAP100 localization at the region of the transverse tubule/junctional sarcoplasmic reticulum. RI is distributed differently from RII in the myocytes. RII, but not RI, was colocalized with AKAP100 in the rat heart. Our studies suggest that AKAP100 tethers PKA II to multiple subcellular compartments for phosphorylation of different pools of substrate proteins in the heart.
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PMID:A-kinase anchoring protein 100 (AKAP100) is localized in multiple subcellular compartments in the adult rat heart. 967 48

A number of studies have reported that the activity of the ryanodine-sensitive calcium release channel (ryanodine receptor) in the junctional sarcoplasmic reticulum of skeletal and cardiac muscle can be modulated by protein phosphorylation-dephosphorylation through activation of endogenous protein kinases and/or by addition of exogenous protein kinases and protein phosphatases. In this study, we have investigated the possibility that protein phosphatase-1 (PP1) is targeted to the junctional sarcoplasmic reticulum by the direct isolation of PP1-binding proteins on PP1-Sepharose affinity columns. The results show that the ryanodine receptor of both skeletal and cardiac muscle bind to this affinity support, and are released at supraphysiological salt concentrations in a relatively pure state. Reciprocal experiments demonstrated that PP1 binds to the immobilized muscle ryanodine receptor. The direct binding of PP1 to the ryanodine receptor was supported by the finding that tryptic fragments of the receptor were retained on PP1-Sepharose. The ability of PP1 to dephosphorylate the ryanodine receptor that was phosphorylated by protein kinase A was also demonstrated. These studies show that PP1 is targeted to the junctional sarcoplasmic reticulum by binding to the ryanodine receptor, and provide a biochemical basis for the possibility that PP1 may play a role in the regulation of calcium flux via protein phosphorylation-dephosphorylation mechanisms.
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PMID:Binding of the catalytic subunit of protein phosphatase-1 to the ryanodine-sensitive calcium release channel protein. 992 79

Ca2+ released from presynaptic and postsynaptic intracellular stores plays important roles in activity-dependent synaptic plasticity, including long-term depression (LTD) of synaptic strength. At Schaffer collateral-CA1 synapses in the hippocampus, presynaptic ryanodine receptor-gated stores appear to mobilize some of the Ca2+ necessary to induce LTD. Cyclic ADP-ribose (cADPR) has recently been proposed as an endogenous activator of ryanodine receptors in sea urchin eggs and several mammalian cell types. Here, we provide evidence that cADPR-mediated signaling pathways play a key role in inducing LTD. We show that biochemical production of cGMP increases cADPR concentration in hippocampal slices in vitro, and that blockade of cGMP-dependent protein kinase, cADPR receptors, or ryanodine-sensitive Ca2+ stores each prevent the induction of LTD at Schaffer collateral-CA1 synapses. A lack of effect of postsynaptic infusion of either cADPR antagonist indicates a probable presynaptic site of action.
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PMID:Evidence of a role for cyclic ADP-ribose in long-term synaptic depression in hippocampus. 1009 63

Cardiac sarcoplasmic reticulum (SR) contains an endogenous phosphorylation system that under specific conditions phosphorylates two proteins with apparent molecular masses of 150 and 130 kDa. The conditions for their phosphorylation are as for the skeletal muscle sarcalumenin and the histidine-rich Ca2+ binding protein (HCP) with respect to: (i) Ca2+ and high concentrations of NaF are required; (ii) phosphorylation is obtained with no added Mg2+ and shows a similar time course and ATP concentration dependence; (iii) inhibition by similar concentrations of La3+; (iv) phosphorylation is obtained with [gamma-32P]GTP; (v) ryanodine binding is inhibited parallel to the phosphorylation of the two proteins. The endogenous kinase is identified as casein kinase II (CK II) based on its ability to use GTP as effectively as ATP, and its inhibition by La3+. The association of CK II with the cardiac SR, even after EGTA extraction at alkaline pH, is demonstrated using antibodies against CK II. The cardiac 130 kDa protein is identified as sarcalumenin based on its partial amino acid sequence and its blue staining with Stains-All. Cardiac sarcalumenin is different from the skeletal muscle protein based on electrophoretic mobilities, immunological analysis, peptide and phosphopeptide maps, as well as amino acid sequencing. Preincubation of SR with NaF and ATP, but not with NaF and AMP-PNP caused strong inhibition of ryanodine binding. This is due to decrease in Ca2+- and ryanodine-binding affinities of the ryanodine receptor (RyR) by about 6.6 and 18-fold, respectively. These results suggest that cardiac sarcalumenin is an isoform of the skeletal muscle protein. An endogenous CK II can phosphorylate sarcalumenin, and in parallel to its phosphorylation the properties of the ryanodine receptor are modified.
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PMID:Cardiac sarcalumenin: phosphorylation, comparison with the skeletal muscle sarcalumenin and modulation of ryanodine receptor. 1039 59

The hypothesis that cAMP-dependent protein kinase (protein kinase A; PKA) is in an active state in small arteries possessing a myogenic tone was investigated in pressurized rat tail small arteries. At a pressure of 80 mmHg, these vessels constricted to 71.6 +/- 1.0% (n = 32) of the diameter of the fully relaxed state. The PKA inhibitors Rp-8-(4-chlorophenylthio)-adenosine 3',5'-cyclic monophosphothioate (Rp-CPT-cAMPS) and N-(2-([3-(4-bromophenyl)-2-propenyl]amino)-ethyl)-5- isoquinolinesulfonamide HCl (H-89) constricted these vessels dose dependently. For example, 300 microM Rp-CPT-cAMPS and 9 microM H-89 reduced vessel diameter by 11.0 +/- 1.2% (n = 8) and 14.3 +/- 3.6% (n = 5), respectively. The cGMP-dependent protein kinase (protein kinase G; PKG) inhibitor Rp-8-bromo-beta-phenyl-1,N(2)-etheno-guanosine 3', 5'-cyclic monophosphothioate (Rp-8-Br-PET-cGMPS) did not alter vessel diameter up to a concentration of 10 microM. Neither endothelium removal nor inhibition of neural transmission affected the action of Rp-CPT-cAMPS. The effect of 300 microM Rp-CPT-cAMPS was reduced by 82% after pretreatment of the vessel with 100 nM iberiotoxin, a blocker of calcium-activated potassium (K(Ca)) channels. However, the effect of 300 microM Rp-CPT-cAMPS was not altered after pretreatment with 1 mM 4-aminopyridine, a blocker of delayed rectifier potassium channels, or 10 microM ryanodine, a blocker of ryanodine receptor-generated calcium sparks. In inside-out patch-clamp experiments on cells isolated from rat tail small arteries, 10 U/ml of the catalytic subunit of PKA together with 100 microM MgATP increased K(Ca) channel activity 30.1 +/- 9. 8-fold (n = 9). Additionally, neither inhibition of PKA or PKG nor moderate activation of PKA or PKG altered the vessel response to a pressure step from 80 to 120 mmHg. These results suggest that in rat tail small arteries possessing a myogenic tone 1) PKA is in an active state modulating the level of the myogenic tone, and 2) K(Ca) channels mediate, at least partly, this effect of PKA.
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PMID:cAMP-dependent protein kinase is in an active state in rat small arteries possessing a myogenic tone. 1048 37

Recent studies have demonstrated that Ca(2+)/calmodulin-dependent protein kinase phosphorylates the Ca(2+)-pumping ATPase of cardiac sarcoplasmic reticulum (SR) in vitro. Also, evidence from in vitro studies suggested that this phosphorylation, occurring at Ser(38), results in stimulation of Ca(2+) transport. In the present study, we investigated whether serine phosphorylation of the SR Ca(2+)-ATPase occurs in the intact functioning heart. Hearts removed from anesthetized rabbits were subjected to retrograde aortic perfusion of the coronary arteries with oxygenated mammalian Ringer solution containing (32)P(i) and contractions were monitored by recording systolic left ventricular pressure development. Following 45-50 min of (32)P perfusion, the hearts were freeze-clamped, SR isolated, and analyzed for protein phosphorylation. SDS-polyacrylamide gel electrophoresis and autoradiography showed phosphorylation of several peptides including the Ca(2+)-ATPase and Ca(2+) release channel (ryanodine receptor). The identity of Ca(2+)-ATPase as a phosphorylated substrate was confirmed by Western immunoblotting as well as immunoprecipitation using a cardiac SR Ca(2+)-ATPase-specific monoclonal antibody. The Ca(2+)-ATPase showed immunoreactivity with a phosphoserine monoclonal antibody indicating that the in situ phosphorylation occurred at the serine residue. Quantification of Ca(2+)-ATPase phosphorylation in situ yielded a value of 208 +/- 12 pmol (32)P/mg SR protein which corresponded to the phosphorylation of approximately 20% of the Ca(2+) pump units in the SR membrane. Since this phosphorylation occurred under basal conditions (i.e., in the absence of any inotropic intervention), a considerable steady-state pool of serine-phosphorylated Ca(2+)-ATPase likely exists in the normally beating heart. These findings demonstrate that serine phosphorylation of the Ca(2+)-ATPase is a physiological event which may be important in the regulation of SR function.
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PMID:Serine phosphorylation of the sarcoplasmic reticulum Ca(2+)-ATPase in the intact beating rabbit heart. 1052 72

Calmodulin (CaM) and Ca(2+)/CaM-dependent protein kinase II (CaM kinase) are tightly associated with cardiac sarcoplasmic reticulum (SR) and are implicated in the regulation of transmembrane Ca(2+) cycling. In order to assess the importance of membrane-associated CaM in modulating the Ca(2+) pump (Ca(2+)-ATPase) function of SR, the present study investigated the effects of a synthetic, high affinity CaM-binding peptide (CaM BP; amino acid sequence, LKWKKLLKLLKKLLKLG) on the ATP-energized Ca(2+) uptake, Ca(2+)-stimulated ATP hydrolysis, and CaM kinase-mediated protein phosphorylation in rabbit cardiac SR vesicles. The results revealed a strong concentration-dependent inhibitory action of CaM BP on Ca(2+) uptake and Ca(2+)-ATPase activities of SR (50% inhibition at approximately 2-3 microM CaM BP). The inhibition, which followed the association of CaM BP with its SR target(s), was of rapid onset (manifested within 30 s) and was accompanied by a decrease in V(max) of Ca(2+) uptake, unaltered K(0.5) for Ca(2+) activation of Ca(2+) transport, and a 10-fold decrease in the apparent affinity of the Ca(2+)-ATPase for its substrate, ATP. Thus, the mechanism of inhibition involved alterations at the catalytic site but not the Ca(2+)-binding sites of the Ca(2+)-ATPase. Endogenous CaM kinase-mediated phosphorylation of Ca(2+)-ATPase, phospholamban, and ryanodine receptor-Ca(2+) release channel was also strongly inhibited by CaM BP. The inhibitory action of CaM BP on SR Ca(2+) pump function and protein phosphorylation was fully reversed by exogenous CaM (1-3 microM). A peptide inhibitor of CaM kinase markedly attenuated the ability of CaM to reverse CaM BP-mediated inhibition of Ca(2+) transport. These findings suggest a critical role for membrane-bound CaM in controlling the velocity of Ca(2+) pumping in native cardiac SR. Consistent with its ability to inhibit SR Ca(2+) pump function, CaM BP (1-2.5 microM) caused marked depression of contractility and diastolic dysfunction in isolated perfused, spontaneously beating rabbit heart preparations. Full or partial recovery of contractile function occurred gradually following withdrawal of CaM BP from the perfusate, presumably due to slow dissociation of CaM BP from its target sites promoted by endogenous cytosolic CaM.
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PMID:Reversible inhibition of the calcium-pumping ATPase in native cardiac sarcoplasmic reticulum by a calmodulin-binding peptide. Evidence for calmodulin-dependent regulation of the V(max) of calcium transport. 1066 Jun 12

In this study, we highlight a role for the nitric oxide-cGMP-dependent protein kinase (NO-G-kinase) signaling pathway in glial intercellular Ca(2+) wave initiation and propagation. Addition of the NO donor molsidomine (100-500 microM) or puffing aqueous NO onto primary glial cell cultures evoked an increase in [Ca(2+)](i) in individual cells and also local intercellular Ca(2+) waves, which persisted after removal of extracellular Ca(2+). High concentrations of ryanodine (100-200 microM) and antagonists of the NO-G-kinase signaling pathway essentially abrogated the NO-induced increase in [Ca(2+)](i), indicating that NO mobilizes Ca(2+) from a ryanodine receptor-linked store, via the NO-G-kinase signaling pathway. Addition of 10 microM nicardipine to cells resulted in a slowing of the molsidomine-induced rise in [Ca(2+)](i), and inhibition of Mn(2+) quench of cytosolic fura-2 fluorescence mediated by a bolus application of 2 microM aqueous NO to cells, indicating that NO also induces Ca(2+) influx in glia. Mechanical stress of individual glial cells resulted in an increase in intracellular NO in target and neighboring cells and intercellular Ca(2+) waves, which were NO, cGMP, and G-kinase dependent, because incubating cells with nitric oxide synthase, guanylate cyclase, and G-kinase inhibitors, or NO scavengers, reduced Delta[Ca(2+)](i) and the rate of Ca(2+) wave propagation in these cultures. Results from this study suggest that NO-G-kinase signaling is coupled to Ca(2+) mobilization and influx in glial cells and that this pathway plays a fundamental role in the generation and propagation of intercellular Ca(2+) waves in glia.
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PMID:A fundamental role for the nitric oxide-G-kinase signaling pathway in mediating intercellular Ca(2+) waves in glia. 1068 78

Although Ca(2+)/calmodulin-dependent protein kinase-II (CaMK) is known to phosphorylate different Ca(2+) cycling proteins in the cardiac sarcoplasmic reticulum (SR) and regulate its function, the status of CaMK in heart failure has not been investigated previously. In this study, we examined the hypothesis that changes in the CaMK-mediated phosphorylation of the SR Ca(2+) cycling proteins are associated with heart failure. For this purpose, heart failure in rats was induced by occluding the coronary artery for 8 weeks, and animals with >30% infarct of the left ventricle wall plus septum mass were used. Noninfarcted left ventricle was used for biochemical assessment; sham-operated animals served as control. A significant depression in SR Ca(2+) uptake and release activities was associated with a decrease in SR CaMK phosphorylation of the SR proteins, ryanodine receptor (RyR), Ca(2+) pump ATPase (SR/endoplasmic reticulum Ca(2+) ATPase [SERCA2a]), and phospholamban (PLB) in the failing heart. The SR protein contents for RyR, SERCA2a, and PLB were decreased in the failing hearts. Although the SR Ca(2+)/calmodulin-dependent CaMK activity, CaMK content, and CaMK autophosphorylation were depressed, the SR phosphatase activity was enhanced in the failing heart. On the other hand, the cAMP-dependent protein kinase-mediated phosphorylation of RyR and PLB was not affected in the failing heart. On the basis of these results, we conclude that alterations in SR CaMK-mediated phosphorylation may be partly responsible for impaired SR function in heart failure.
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PMID:Sarcoplasmic reticulum Ca(2+)/Calmodulin-dependent protein kinase is altered in heart failure. 1072 Apr 22


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