Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.13.12.5 (aequorin)
 1,451 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Protein kinase C regulates the activity of a diverse group of cellular proteins including membrane ion channel proteins. Although protein kinase C and its substrate protein have been identified in both membrane and cytosolic fractions in the heart, the physiological role of this kinase in the regulation of cardiac function remains unknown. We examined the physiological role of protein kinase C by stimulating its activity with 12-deoxyphorbol 13 isobutyrate 20 acetate (DPBA) in human trabeculae carneae. This resulted in decreased peak isometric twitch force and peak intracellular sarcoplasmic reticulum calcium release as detected with aequorin. Furthermore, in the presence of DPBA, steady-state force-[Ca2+] relations were shifted to higher intracellular calcium concentrations, and the Hill coefficient was reduced, indicating a decrease in responsiveness of the myofilaments to calcium and a change in cooperativity among thin filament proteins, respectively. Thus, DPBA affects not only intracellular calcium concentration, but myofilament calcium interactions as well. The effect of DPBA on Ca2+ activation probably reflects phosphorylation of thin-filament regulatory proteins by protein kinase C.
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PMID:Effect of protein kinase C activation on sarcoplasmic reticulum function and apparent myofibrillar Ca2+ sensitivity in intact and skinned muscles from normal and diseased human myocardium. 220 99

1. Intracellular Ca2+-force relationships were investigated in ferret aortic smooth muscle by the simultaneous measurement of aequorin luminescence and isometric force. Complete calcium-force curves were constructed by plotting calibrated aequorin luminescence versus force, while intracellular [Ca2+] was made to change by increasing degrees of K+ depolarization or decreasing extracellular [Ca2+]. 2. The steady state calcium-force curve in response to K+ depolarization exhibited maximal force generation at an intracellular [Ca2+] of approximately 4 x 10(-7) M. Further increases in intracellular [Ca2+] did not yield additional increments in force. 3. Protein kinase C activation with the phorbol ester, 12-deoxyphorbol-13-isobutyrate 20 acetate (DPBA) produced contractions accompanied by no detectable increases in aequorin luminescence. DPBA significantly shifted the control [Ca2+]-force relationship leftward to lower intracellular [Ca2+] with an increase in the magnitude of maximal generated force. 4. In aorta maximally precontracted by K+ depolarization, the addition of DPBA resulted in a significant increase in force in the absence of further increases in intracellular [Ca2+]. Conversely, in muscles maximally precontracted with DPBA, responses to K+ depolarization resulted in subsequent increases in force in the presence of simultaneous sustained increases in intracellular [Ca2+]. 5. The relatively specific protein kinase C antagonist H-7 caused a significant decrease in intrinsic myogenic tone in the absence of any statistically significant decrease in intracellular [Ca2+]. 6. These results suggest that protein kinase C may be an important regulator of vascular smooth muscle contractility by: (1) providing a mechanism by which the apparent [Ca2+] sensitivity of the contractile apparatus during agonist-induced contractions is increased, and (2) maintaining intrinsic myogenic tone by a mechanism the [Ca2+] requirement of which is satisfied by the resting [Ca2+]i.
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PMID:Involvement of the protein kinase C system in calcium-force relationships in ferret aorta. 275 22

We investigated the relationship between [Ca2+]i, myosin light chain (LC20) phosphorylation and isometric force in guinea pig aortic strips during contractions activated by a thromboxane A2 analogue, (15S)-hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5Z, 13E-dienoic acid (U46619). Isometric force and [Ca2+]i were measured simultaneously using preloaded aequorin as the intracellular calcium indicator. LC20 phosphorylation levels were determined by two dimensional polyacrylamide gel electrophoresis in parallel preparations. Contractions induced by U46619 were accompanied by increases in [Ca2+]i and LC20 phosphorylation. The chelation of extracellular calcium with 2.5 mM EGTA significantly inhibited U46619-induced increases in [Ca2+]i, isometric force and LC20 phosphorylation. Steady-state force assumed a similar dependence on LC20 phosphorylation for contractions stimulated by potassium depolarization, alpha 1-adrenergic agonist phenylephrine and U46619 either in the presence or absence of extracellular calcium. On the contrary, the [Ca2+]i/force relation revealed that both U46619 and phenylephrine stimulated greater isometric force at lower [Ca2+]i than did KCl depolarization. The addition of a protein kinase C inhibitor, 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7), decreases force without significantly affecting either [Ca2+]i or LC20 phosphorylation levels. These results suggest that in guinea pig aortic smooth muscle U46619 increases the calcium sensitivity of the contractile apparatus but does not change the LC20 phosphorylation/force relation in comparison to K+ depolarization. Protein kinase C is activated during U46619-stimulated contraction and might be involved in mechanisms other than LC20 phosphorylation.
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PMID:Intracellular calcium and myosin light chain phosphorylation during U46619-activated vascular contraction. 751 55

We have previously proposed that fluctuations in Ca(2+) levels should play an important role in bacteria as in eukaryotes in regulating cell cycle events (Norris et al., J. Theor. Biol. 134 (1998) 341-350). This proposal implied the presence of Ca(2+) uptake systems in bacteria, cell cycle mutants simultaneously defective in Ca(2+)-homeostasis, and perturbation of cell cycle processes when cellular Ca(2+) levels are depleted. We review the properties of new cell cycle mutants in E. coli and B. subtilis resistant to inhibitors of calmodulin, PKC or Ca(2+)-channels; the evidence for Ca(2+)-binding proteins including Acp and FtsZ; and Ca(2+)-transporters. In addition, the effects of EGTA and verapamil (a Ca(2+) channel inhibitor) on growth, protein synthesis and cell cycle events in E. coli are described. We also describe new measurements of free Ca(2+)-levels, using aequorin, in E. coli. Several new cell cycle mutants were obtained using this approach, affecting either initiation of DNA replication or in particular cell division at non-permissive temperature. Several of the mutants were also hypersensitive to EGTA and or Ca(2+). However, none of the mutants apparently involved direct alteration of a drug target and surprisingly in some cases involved specific tRNAs or a tRNA synthetase. The results also indicate that the expression of several genes in E. coli may be regulated by Ca(2+). Cell division in particular appears very sensitive to the level of cell Ca(2+), with the frequency of division clearly reduced by EGTA and by verapamil. However, whilst the effect of EGTA was clearly correlated with depletion of cellular Ca(2+) including free Ca(2+), this was not the case with verapamil which appears to change membrane fluidity and the consequent activity of membrane proteins. Measurement of free Ca(2+) in living cells indicated levels of 200-300 nM, tightly regulated in wild type cells in exponential phase, somewhat less so in stationary phase, with apparently La(2+)-sensitive PHB-polyphosphate complexes involved in Ca(2+) influx. The evidence reviewed increasingly supports a role for Ca(2+) in cellular processes in bacteria, however, any direct link to the control of cell cycle events remains to be established.
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PMID:An assessment of the role of intracellular free Ca2+ in E. coli. 1057 4

We analyzed the role of inositol 1,4,5-trisphosphate-induced Ca(2+) release from the endoplasmic reticulum (ER) (i) in powering mitochondrial Ca(2+) uptake and (ii) in maintaining a sustained elevation of cytosolic Ca(2+) concentration ([Ca(2+)](c)). For this purpose, we expressed in HeLa cells aequorin-based Ca(2+)-sensitive probes targeted to different intracellular compartments and studied the effect of two agonists: histamine, acting on endogenous H(1) receptors, and glutamate, acting on co-transfected metabotropic glutamate receptor (mGluR1a), which rapidly inactivates through protein kinase C-dependent phosphorylation and thus causes transient inositol 1,4,5-trisphosphate production. Glutamate induced a transient [Ca(2+)](c) rise and drop in ER luminal [Ca(2+)] ([Ca(2+)](er)), and then the ER refilled with [Ca(2+)](c) at resting values. With histamine, [Ca(2+)](c) after the initial peak stabilized at a sustained plateau, and [Ca(2+)](er) decreased to a low steady-state value. In mitochondria, histamine evoked a much larger mitochondrial Ca(2+) response than glutamate ( approximately 15 versus approximately 65 microm). Protein kinase C inhibition, partly relieving mGluR1a desensitization, reestablished both the [Ca(2+)](c) plateau and the sustained ER Ca(2+) release and markedly increased the mitochondrial Ca(2+) response. Conversely, mitochondrial Ca(2+) uptake evoked by histamine was drastically reduced by very transient ( approximately 2-s) agonist applications. These data indicate that efficient mitochondrial Ca(2+) uptake depends on the preservation of high Ca(2+) microdomains at the mouth of ER Ca(2+) release sites close to mitochondria. This in turn depends on continuous Ca(2+) release balanced by Ca(2+) reuptake into the ER and maintained by Ca(2+) influx from the extracellular space.
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PMID:Mitochondrial Ca2+ uptake requires sustained Ca2+ release from the endoplasmic reticulum. 1258 23

Alpha(1)-adrenoceptor stimulation (alpha(1)ARS) shows a positive inotropic effect in most mammalian myocardium. In mouse myocardium, however, alpha(1)ARS showed the negative inotropic effect, of which intracellular mechanisms are not fully clarified. The purpose of this study is to investigate the intracellular mechanism of the negative inotropic effect by alpha(1)ARS in C57BL/6 mouse myocardium. We used isolated ventricular papillary muscles of C57BL/6 strain mouse which is widely used for genetic manipulation. We simultaneously measured isometric tension and intracellular Ca(2+) concentration ([Ca(2+)](i)) using the aequorin method. In twitch contraction, phenylephrine concentration-dependently (1-100 microM) decreased tension without significant changes in the Ca(2+) transient, and these effects were completely blocked by prazosin (3 microM) or calphostin C (a PKC inhibitor, 1 microM). Phorbol 12-myristate 13-acetate (PMA) (a PKC activator, 1 microM) decreased tension as observed in phenylephrine. After PMA application, the negative inotropic effect of phenylephrine disappeared. To estimate the Ca(2+) sensitivity, tetanic contraction was produced, and the relation between [Ca(2+)](i) and tension at a steady state was measured. Phenylephrine (10 microM) decreased the Ca(2+) sensitivity, and PMA showed a similar Ca(2+) desensitizing effect. These results suggest that the negative inotropic effect of phenylephrine in mouse myocardium can be explained by the decrease in the Ca(2+) sensitivity through the activation of PKC. The present result indicates that the effect of alpha(1)ARS differs among species and strains of experiment animals. Thus, we should be careful about using the results of mouse myocardium to understand the functions of the human heart.
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PMID:Intracellular mechanism of the negative inotropic effect induced by alpha1-adrenoceptor stimulation in mouse myocardium. 1688 59