Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The discovery of a calcium receptor has stimulated interest in the signaling events underlying extracellular calcium ([Ca2+]o)-induced cell-specific responses. In osteoblasts, elevated levels of extracellular calcium mediate both mitogenesis and chemotaxis. Here we provide evidence that [Ca2+]o-stimulated chemotaxis of MC3T3-E1 osteoblast-like cells involves a G-protein-linked calcium-sensing receptor. [Ca2+]o promotes chemotaxis in a concentration-dependent manner. Pertussis toxin blocked almost all of [Ca2+]o-stimulated chemotaxis but had only a small effect on platelet-derived growth factor (PDGF)-stimulated chemotaxis. Consistent with the signaling model for PDGF-mediated chemotaxis, activation of phospholipase C played a critical role in [Ca2+]o-initiated chemotaxis: U-73122, an inhibitor of the activation of phospholipase C, blocked approximately 50% of PDGF-stimulated chemotaxis but blocked nearly all of the [Ca2+]o-stimulated chemotaxis. Down-regulation of protein kinase C also blocked about 50% of PDGF-stimulated chemotaxis but did not block [Ca2+]o-stimulated chemotaxis. Thus, unlike PDGF-mediated chemotaxis, chemotaxis stimulated by [Ca2+]o does not appear to require protein kinase C activation. This finding suggests events downstream of inositol 1,4,5-trisphosphate production rather than diacylglycerol production are critical to [Ca2+]o-promoted chemotaxis of MC3T3-E1 cells. The signal transduction mechanism underlying PDGF-induced chemotaxis involves the activation of phosphoinositide 3-kinase, as judged by the in vivo production of phosphatidylinositol 3,4-diphosphate and 3,4,5-trisphosphate and the partial sensitivity of chemotaxis to wortmannin, an inhibitor of phosphoinositide 3-kinase. In contrast, [Ca2+]o-stimulated chemotaxis was not blocked by wortmannin and elevations in [Ca2+]o did not increase the production of lipid products of phosphoinositide 3-kinase. Overall, [Ca2+]o-promoted chemotaxis of osteoblasts appears to utilize a unique signaling mechanism via a calcium-sensing receptor.
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PMID:Extracellular calcium and platelet-derived growth factor promote receptor-mediated chemotaxis in osteoblasts through different signaling pathways. 911 Oct 36

We examined whether calcium receptor (CaR) signaling is affected by protein kinase C (PKC) activation by assessing the effects of phorbol-12-myristate-13-acetate (PMA) on 45Ca2+ efflux from Xenopus laevis oocytes expressing wild-type (WT) and mutant bovine parathyroid CaRs. Raising extracellular [Ca2+] ([Ca2+]0) from 0.5 to 5.5 mM increased 45Ca efflux (26 +/- 3-fold) in oocytes expressing full-length and C-terminally truncated receptor (amino acid 1-895). These increases in 45Ca efflux were blocked by 88 +/- 3% after PMA treatment for 20 min. Three consensus PKC phosphorylation sites (Thr-647, Ser-795, and Thr-889) were mutated in the context of the full-length and truncated CaR. PMA treatment inhibited high [Ca2+]0-induced responses in oocytes expressing the Ser795Ala CaR (1-895), Thr889Ala CaR (1-895), and Ser795Ala/Thr889Ala CaR (1-895) by 30-40% compared with untreated controls (P < 0.05). A triple mutant of the full-length CaR demonstrated similarly reduced susceptibility to inhibition of 45Ca efflux by PMA. Thus, these sites are important in mediating the effects of PKC activation on CaRs, but other residues and effector molecules are likely to participate in the effects of PKC on CaR-induced signal transduction in target cells.
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PMID:Protein kinase C activation blocks calcium receptor signaling in Xenopus laevis oocytes. 1063 Apr 1

We examined the role of protein kinase C (PKC) in the mechanism and regulation of intracellular Ca(2+) concentration ([Ca(2+)](i)) oscillations elicited by an increase in the extracellular concentration of Ca(2+) ([Ca(2+)](e)) in human embryonic kidney 293 cells expressing the Ca(2+)-sensing receptor (CaR). Exposure to the PKC inhibitors bisindolylmaleimide I (GF I) or Ro-31-8220 converted oscillatory responses to transient, non-oscillatory responses, significantly reducing the percentage of cells that showed [Ca(2+)](i) oscillations but without decreasing the overall response to increase in [Ca(2+)](e). Exposure to 100 nm phorbol 12,13-dibutyrate, a direct activator of PKC, eliminated [Ca(2+)](i) oscillations. Addition of phorbol 12,13-dibutyrate at lower concentrations (3 and 10 nm) did not eliminate the oscillations but greatly reduced their frequency in a dose-dependent manner. Co-expression of CaR with constitutively active mutants of PKC (either epsilon or beta(1) isoforms) also reduced [Ca(2+)](i) oscillation frequency. Expression of a mutant CaR in which the major PKC phosphorylation site is altered by substitution of alanine for threonine (T888A) eliminated oscillatory behavior, producing [Ca(2+)](i) responses almost identical to those produced by the wild type CaR exposed to PKC inhibitors. These results support a model in which phosphorylation of the CaR at the inhibitory threonine 888 by PKC provides the negative feedback needed to cause [Ca(2+)](i) oscillations mediated by this receptor.
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PMID:Ca2+-stimulated Ca2+ oscillations produced by the Ca2+-sensing receptor require negative feedback by protein kinase C. 1235 61

Medullary thick ascending limb (mTAL) cells in primary culture express the Ca(2+)-sensing receptor (CaR), a G protein-coupled receptor that senses changes in extracellular Ca(2+) (Ca(o)(2+)) concentration, resulting in increases of intracellular Ca(2+) concentration and PKC activity. Exposure of mTAL cells to either Ca(o)(2+) or the CaR-selective agonist poly-L-arginine increased TNF-alpha synthesis. Moreover, the response to Ca(o)(2+) was enhanced in mTAL cells transfected with a CaR overexpression vector. Transfection of mTAL cells with a TNF promoter construct revealed an increase in reporter gene activity after exposure of the cells to Ca(o)(2+), suggesting that intracellular signaling pathways initiated by means of activation of a CaR contribute to TNF synthesis by a mechanism that involves transcription of the TNF gene. Neutralization of TNF activity with an anti-TNF antibody attenuated Ca(2+)-mediated increases in cyclooxygenase-2 (COX-2) protein expression and PGE(2) synthesis, suggesting that TNF exerts an autocrine effect in the mTAL, which contributes to COX-2-mediated PGE(2) production. Preincubation with the PKC inhibitor bisindolylmaleimide I inhibited Ca(2+)-mediated TNF production. Significant inhibition of COX-2 protein expression and PGE(2) synthesis also was observed when cells were challenged with Ca(o)(2+) in the presence of bisindolylmaleimide I. The data suggest that increases in TNF production subsequent to activation of the CaR may be the basis of an important renal mechanism that regulates salt and water excretion.
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PMID:Calcium-sensing receptor-mediated TNF production in medullary thick ascending limb cells. 1237 72

Although extracellular calcium (Ca(2+)(o)) has been suggested to modulate bone remodeling, the exact mechanism is unclear. This study was performed to explore the signaling pathways of high Ca(2+)(o) that are responsible for controlling the expression of receptor activator of NF-kappaB ligand (RANKL) in mouse osteoblastic cells. As previously reported, high Ca(2+)(o) increased RANKL expression. However, the G protein-coupled Ca(2+)(o)-sensing receptor (CaSR) was not detected in the primary cultured mouse osteoblastic cell. The inhibition of the pertussis-sensitive G protein, phospholipase C, protein kinase C, intracellular calcium mobilization, p38 MAPK, or phosphoinositide 3-kinase did not block RANKL induction caused by high Ca(2+)(o). In contrast, the inhibition of p44/42 MAPK pathway reduced the RANKL expression induced by high Ca(2+)(o). Moreover, high Ca(2+)(o) activated p44/42 MAPK and MEK1/2. These results suggest that RANKL induction by high Ca(2+)(o) might not be mediated by CaSR and its putative downstream signaling pathways, but the pathway employing p44/42 MAPK is involved in the high Ca(2+)(o)-induced RANKL expression in mouse osteoblastic cells.
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PMID:p44/42 MAPK activation is necessary for receptor activator of nuclear factor-kappaB ligand induction by high extracellular calcium. 1272 16

Current therapy for secondary hyperparathyroidism in uremia has relatively poor success in achieving the target levels of parathyroid hormone (PTH), calcium and phosphate established by the NKF-K/DOQI guidelines. The discovery and characterization of a new membrane receptor able to sense minimal Ca changes (CaSR) started intensive research in the attempt to characterize better its functions and its finding compounds, which could modulate its activity. CaSR is expressed not only in the cells that secrete calcium-regulating hormones (parathyroid cells and thyroid C-cells) and in cells involved in calcium transport mechanisms (ie intestinal cells, bone-forming osteoblasts, and cells of different nephron segments), but also in other tissues with, as yet, a not completely defined role. CaSR stimulation by the agonists is followed by the activation of a great number of G-proteins mediated intracellular signalling pathways (PLC, PLA, PLD, PKC, PKA, etc). At the level of parathyroid cells, the main effect is the increase in IP3, followed by a mobilization of intracellular Ca stores, which inhibit PTH secretion in a few seconds or minutes. Long-term CaSR stimulation is also able to induce a reduction in both PTH synthesis and parathyroid cell proliferation. More than 100 mutations of the gene coding for CaSR have been described. Some of these mutations are matched by a gain or reduction/loss of function. Notwithstanding, CaSR is widely represented on different tissue cells, the main clinical manifestations of the above genetic changes mainly involve PTH and calcium metabolism. A great number of inorganic and organic cations can interact with the Ca-sensitive N-terminus domain of CaSR, mimicking Ca effects (type I calcimimetics), but these substances have substantial limitations for use in clinical practice. A second class of compounds was produced (NPS R-467, S-467, R-568, S-568, AMG 073), for use in the clinical setting, type II calcimimetics. These compounds, after having interacted with the membrane-spanning domains of the CaSR, induce conformational changes in the N-terminus domain, increasing its affinity for Ca. The preclinical experiences with calcimimetics demonstrated that they were effective in reducing circulating PTH, preventing the progression of secondary hyperparathyroidism, suppressing parathyroid cell proliferation, and reversing osteitis fibrosa at least in animal models. Clinical studies were performed mainly using AMG 073, due to its greater bioavailability and more consistent pharmacokinetic profile. Clinical studies performed in primary hyperparathyroidism proved AMG 073 to be effective in reducing both PTH and Ca serum levels, with a good safety profile. Further studies, mainly focused on the efficacy of AMG 073 in the control of secondary hyperparathyroidism in uremia, confirmed the efficacy of this compound in reducing PTH levels >30% in about 50% of patients. Furthermore, the fall in PTH was matched by a reduction in both calcium and phosphate serum levels of about 5-7%, with a significant reduction in calcium x phosphate product (about 15%). The latter aspect represents a unique pharmacological profile, as compared to all the other available therapeutic means to control secondary hyperparathyroidism in uremia. In addition to their effectiveness, calcimimetics present a relatively safe profile, the only adverse events referred to consist of transient and easily remediable hypocalcemic episodes and some gastrointestinal discomfort symptoms. However, although calcimimetics represent a real advancement in the field of treating secondary hyperparathyroidism in uremic patients, their use should be matched by the awareness that previously the success of a high number of new drugs proposed have been flawed by negative consequences in the long term. Therefore, strict clinical control is necessary in the next few years when the use of these new compounds will widen.
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PMID:[Calcimimetics]. 1652 Oct 71

The rat dorsal root ganglion (DRG) Ca(2+)-sensing receptor (CaR) was stably expressed in-frame as an enhanced green fluorescent protein (EGFP) fusion protein in human embryonic kidney (HEK)293 cells, and is functionally linked to changes in intracellular Ca(2+) concentration ([Ca(2+)](i)). RT-PCR analysis indicated the presence of the message for the DRG CaR cDNA. Western blot analysis of membrane proteins showed a doublet of 168-175 and 185 kDa, consistent with immature and mature forms of the CaR.EGFP fusion protein, respectively. Increasing extracellular [Ca(2+)] ([Ca(2+)](e)) from 0.5 to 1 mM resulted in increases in [Ca(2+)](i) levels, which were blocked by 30 microM 2-aminoethyldiphenyl borate. [Ca(2+)](e)-response studies indicate a Ca(2+) sensitivity with an EC(50) of 1.75 +/- 0.10 mM. NPS R-467 and Gd(3+) activated the CaR. When [Ca(2+)](e) was successively raised from 0.25 to 4 mM, peak [Ca(2+)](i), attained with 0.5 mM, was reduced by approximately 50%. Similar reductions were observed with repeated applications of 10 mM Ca(2+), 1 and 10 microM NPS R-467, or 50 and 100 microM Gd(3+), indicating desensitization of the response. Furthermore, Ca(2+) mobilization increased phosphorylated protein kinase C (PKC)alpha levels in the cells. However, the PKC activator, phorbol myristate acetate did not inhibit CaR-mediated Ca(2+) signaling. Rather, a spectrum of PKC inhibitors partially reduced peak responses to Ca(e)(2+). Treatment of cells with 100 nM PMA for 24 h, to downregulate PKC, reduced [Ca(2+)](i) transients by 49.9 +/- 5.2% (at 1 mM Ca(2+)) and 40.5 +/- 6.5% (at 2 mM Ca(2+)), compared with controls. The findings suggest involvement of PKC in the pathway for Ca(2+) mobilization following CaR activation.
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PMID:Ca(2+) mobilization through dorsal root ganglion Ca(2+)-sensing receptor stably expressed in HEK293 cells. 1726 50

Gastric acid secretion is not only stimulated via the classical known neuronal and hormonal pathways but also by the Ca(2+)-Sensing Receptor (CaSR) located at the basolateral membrane of the acid-secretory gastric parietal cell. Stimulation of CaSR with divalent cations or the potent agonist Gd(3+) leads to activation of the H(+)/K(+)-ATPase and subsequently to gastric acid secretion. Here we investigated the intracellular mechanism(s) mediating the effects of the CaSR on H(+)/K(+)-ATPase activity in freshly isolated human gastric glands. Inhibition of heterotrimeric G-proteins (G(i) and G(o)) with pertussis toxin during stimulation of the CaSR with Gd(3+) only partly reduced the observed stimulatory effect. A similar effect was observed with the PLC inhibitor U73122. The reduction of the H(+)/K(+)-ATPase activity measured after incubation of gastric glands with BAPTA-AM, a chelator of intracellular Ca(2+), showed that intracellular Ca(2+) plays an important role in the signalling cascade. TMB-8, a ER Ca(2+)store release inhibitor, prevented the stimulation of H(+)/K(+)-ATPase activity. Also verapamil, an inhibitor of L-type Ca(2+)-channels reduced stimulation suggesting that both the release of intracellular Ca(2+) from the ER as well as Ca(2+) influx into the cell are involved in CaSR-mediated H(+)/K(+)-ATPase activation. Chelerythrine, a general inhibitor of protein kinase C, and Go 6976 which selectively inhibits Ca(2+)-dependent PKC(alpha) and PKC(betaI)-isozymes completely abolished the stimulatory effect of Gd(3+). In contrast, Ro 31-8220, a selective inhibitor of the Ca(2+)-independent PKCepsilon and PKC-delta isoforms reduced the stimulatory effect of Gd(3+) only about 60 %. On the other hand, activation of PKC with DOG led to an activation of H(+)/K(+)-ATPase activity which was only about 60 % of the effect observed with Gd(3+). Incubation of the parietal cells with PD 098059 to inhibit ERK1/2 MAP-kinases showed a significant reduction of the Gd(3+) effect. Thus, in the human gastric parietal cell the CaSR is coupled to pertussis toxin sensitive heterotrimeric G-Proteins and requires calcium to enhance the activity of the proton-pump. PLC, ERK 1/2 MAP-kinases as well as Ca(2+) dependent and Ca(2+)-independent PKC isoforms are part of the down-stream signalling cascade.
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PMID:Stimulatory pathways of the Calcium-sensing receptor on acid secretion in freshly isolated human gastric glands. 1731 98

Under normoxia, FIH-1 (factor inhibiting HIF-1) inhibits the transcriptional activity of hypoxia-inducible factor (HIF); however, under such conditions, we observed a significant level of HIF activity in renal cell carcinoma (RCC). This phenomenon could be attributed to a decrease in the level of functional FIH that has been identified in our previous work. Nonetheless, the molecular mechanism of FIH regulation in cancer, in particular RCC, was unclear until now. In this communication, we have demonstrated that in RCC, the Cut-like homeodomain protein (CDP/Cut) is involved in FIH transcriptional regulation and is controlled by a specific signaling event involving protein kinase C (PKC) zeta. Furthermore, we have defined a unique CDP/Cut binding site on the FIH promoter. With chromatin immunoprecipitation assays, we show that CDP binds to the FIH-1 promoter in vivo and that this binding is PKC zeta dependent. Moreover, we have also defined a potential phosphorylation site in CDP (serine 987) that modulates FIH expression. CDP/Cut is a transcriptional repressor that decreases FIH-1 expression and subsequently leads to a decrease in the repressor activity of FIH-1. Without this repression, HIF activity increases, allowing for the increased transcription of the genes it regulates, such as the vascular endothelial growth factor and GLUT-1 genes. Both CDP and HIF levels are increased in several cancers and are responsible for the metastatic progression of the tumors. Taken together, our results suggest for the first time a potential connection between CDP and FIH that could lead to the development of future therapeutic interventions.
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PMID:Protein kinase C-mediated modulation of FIH-1 expression by the homeodomain protein CDP/Cut/Cux. 1768 59

Calcium is a second messenger for many signaling pathways in B cells, but its role as a receptor ligand has not been well characterized. However, pulses of free calcium were found to cause the rapid release of internal calcium stores in normal human B cells. This response appeared to be mediated by a cell surface protein with receptor properties as it could be blocked by pretreatment with trypsin and with kinase and phospholipase Cgamma inhibitors. The calcium receptor on B cells was not the conventional calcium-sensing receptor (CaSR) since B cells did not express CaSR and calcium-induced responses could not be blocked by specific CaSR inhibitors. B-cell responses to extracellular calcium activated phosphoinositide-3 kinase/AKT, calcineurin, extracellular signal regulated kinase, p38 mitogen-activated protein kinase, protein kinase C, Ca(2+)/calmodulin kinase II, and nuclear factor-kappaB signaling pathways, and resulted in transcription of the early response gene, CD83. This extracellular calcium sensor enhanced B-cell responses to Toll-like receptor, B-cell receptor, and cytokine receptor agonists. These findings suggest a means by which B cells prepare to engage in immune responses by responding to calcium fluctuations in their environment.
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PMID:Extracellular calcium sensing promotes human B-cell activation and function. 1772 42


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