Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.4.3 (phospholipase C)
 18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The Ca2+ ion exerts a profound influence on cellular processes and an understanding of control mechanisms of intracellular Ca2 homeostasis while complex is mandatory in this discussion. The identification and recognition of prolonged sustained increase in [Ca2+]i as a manifestation of neurotoxin-induced destabilization of [Ca2+]i homeostasis will be related to a variety of neurotoxicant-induced cell injuries. The sites of toxicant interaction with ATP-regulated Ca2+ pumps located in the neuronal/glial membrane and/or calciosomes; availability of Ca2+ proteins; disruption in mitochondrial mechanisms for Ca2+ storage; triggers of voltage-dependent Ca2+ channels and modulation of the Na+/Ca2+ exchanger will be identified and related to presumptive toxin action. Failure of one or more of these systems will result in continuous elevation of ionized [Ca2+]i--a reflection of Ca2+ destabilization. The targets resulting from Ca2+ destabilization will be identified, to include phospholipase C activation, PLA2 activation, protein kinase C (PKC) translocation, and activation of Ca(2+)-dependent calpain 1. The use of specific inhibitors of neurotoxicity, e.g., natural sphingolipids, sphingosine, down regulation of PKC, inhibitors and activators of adenylate cyclase, and antiprotease agents will allow for investigation of the role of these final common pathways in the evolution of neurotoxicity.
 ...
PMID:Ca(2+)-dependent processes as mediators of neurotoxicity. 150 13

Employing a cell penetrating calpain inhibitor (calpeptin), the role of calpain in platelet activation was examined. In washed platelets (WPs) both thrombin and collagen-induced platelet aggregation were dose-dependently inhibited by calpeptin. The addition of plasma to WPs interfered with the action of calpeptin, however more than 3 min preincubation of calpeptin with WPs completely abolished the influence of plasma. In thrombin-activated WPs with calcium, the increase of intracellular calcium concentration, [Ca2+]i, and the production of inositol triphosphate (IP3) were dose-dependently inhibited by calpeptin. The generation of thromboxane B2 (TxB2) was inhibited by calpeptin in collagen and thrombin-activated WPs. In [3H]-arachidonic acid (AA)-labelled platelets, calpeptin increased the amount of [3H]-AA liberated by inhibiting [3H]-AA degradation after collagen or thrombin stimulation. When [14C]-AA degradation by the platelet suspension was observed, calpeptin inhibited TxB2 and hydroxyheptadecatrienoic acid (HHT) generation but increased prostaglandin (PG) E1, E2, 12-hydroxyeicosatetraenoic acid (12HETE) and AA. Based on these findings, calpain may be involved in the activation phospholipase C and thromboxane synthetase.
 ...
PMID:Potential participation of calpain in platelet activation studied by use of a cell penetrating calpain inhibitor (calpeptin). 211 Feb 78

Trypsin causes rapid activation of intact platelets that mimics many actions of thrombin, including the stimulation of phospholipase C (PLC). We have examined the effects of thrombin and trypsin on PLC in a platelet membrane preparation using exogenous [3H]-phosphatidylinositol 4,5-bisphosphate (PIP2) as substrate. Trypsin induced PIP2 breakdown, which was maximal at 20 micrograms/ml, but was reduced at higher concentrations. alpha- and gamma-Thrombins also stimulated PLC-induced hydrolysis of PIP2 in membranes. This effect was inhibited by leupeptin. Exogenous [3H]phosphatidylinositol 4-monophosphate (PIP) was hydrolyzed in response to both thrombin and trypsin in the same ratio as PIP2. Activation of membrane-bound PLC persisted after removal of thrombin and trypsin. The hydrolysis of [3H]phosphatidylinositol was not activated by alpha-thrombin and trypsin. We examined the question of whether calpain was involved in the observed PLC activation by thrombin and trypsin. Although dibucaine activated a Ca2(+)-dependent protease as judged by the hydrolysis of actin-binding protein and by the activation of phosphoprotein phosphatases, it failed to stimulate the generation of phosphatidic acid in 32P-prelabeled platelets. Moreover, when PLC was assayed in the membranes, the addition of Ca2(+)-activated neutral proteinases did not increase the rate of hydrolysis of either PIP or PIP2. Our results show that proteases such as trypsin and thrombin are able to stimulate membrane-bound PLC, but this activation does not seem to be related to calpain.
 ...
PMID:Persistent activation of platelet membrane phospholipase C by proteolytic action of trypsin and thrombin. 229 26

Inhibitors of calcium-dependent proteases (calpains) such as leupeptin and antipain have been shown to selectively inhibit platelet activation by thrombin. Based upon this observation, it has been proposed that calpains play a role in the initiation of platelet activation. In the present studies, we have examined the effect of leupeptin on the earliest known event in thrombin-induced platelet activation: the interaction between the agonist, its receptors, and the guanine nucleotide-binding proteins which stimulate phospholipase C (Gp) and inhibit adenylyl cyclase (Gi). We found that leupeptin inhibited thrombin's ability to stimulate phosphoinositide hydrolysis, suppress cAMP formation, and dissociate Gp and Gi into subunits. Leupeptin had no effect, however, on the same responses to other agonists or on thrombin binding to platelets. Although these observations might suggest, as others have concluded, that calpain is involved in the initiation of platelet activation by thrombin, we also found that: 1) substituting platelet membranes for intact platelets and decreasing the free Ca2+ concentration below the threshold required for calpain activation did not diminish the effects of leupeptin on phosphoinositide hydrolysis and cAMP formation, 2) washing the platelets after incubation with leupeptin reversed the effects of the inhibitor, 3) permeabilizing the platelets with saponin did not enhance the inhibitory effects of leupeptin, and 4) leupeptin inhibited the proteolysis of fibrinogen and the hydrolysis of S2238 by thrombin. Similar results in these assays were obtained with antipain. Therefore, our observations suggest that the inhibition of platelet activation by leupeptin is due to a direct interaction with thrombin and need not reflect a role for calpain in the initiation of platelet activation.
 ...
PMID:Inhibition of thrombin-induced platelet activation by leupeptin. Implications for the participation of calpain in the initiation of platelet activation. 283 98

A single point mutation, Glu627--> Val, equivalent to the activating mutation in the Neu oncogene, was inserted in the transmembrane domain of the human epidermal growth factor (EGF) receptor. Unlike the wild type, Glu627-EGF receptor, transfected in NIH3T3 cells, gave rise to focal transformation and growth in agar even in the absence EGF. Constitutive activity of mutant EGF receptor amounted to 20% of that of wild type receptor stimulated by EGF. In addition, the mutant receptor was more sensitive to EGF, reaching maximum transforming activity at 5 ng/ml EGF. NIH3T3 cells expressing Glu627-EGF receptor showed a transformed phenotype and were not arrested in G0 upon serum deprivation. The mutant receptor was constitutively autophosphorylated, and several other cellular proteins were phosphorylated on tyrosine in absence of the ligand. Among these, the SHC adaptor protein was phosphorylated in absence of EGF, the other adaptor, GRB-2 was constitutively associated with the Glu627-EGF receptor in vivo and in vitro, and mitogen-activated protein kinase was constitutively phosphorylated. In contrast, other EGF receptor substrates, like phospholipase C gamma, were not phosphorylated in absence of EGF. The mutant receptor showed a higher sensitivity to cleavage by calpain both in absence and presence of EGF, appeared as a 170- and 150-kDa doublet in cell extracts, and a specific calpain inhibitor blocked the appearance of the 150-kDa form. Since the calpain cleavage site is located in the receptor cytoplasmic tail, this finding suggests that the Glu627 mutation induces a slightly different conformation in the EGF receptor intracellular domain. In conclusion, our data show that a point mutation in the EGF receptor transmembrane domain was able to constitutively activate the receptor and to induce transformation via constitutive activation of the Ras pathway.
 ...
PMID:SHC and GRB-2 are constitutively by an epidermal growth factor receptor with a point mutation in the transmembrane domain. 764 41

The hypothesis that cytoplasmic proteases play a functional role in programmed cell death was tested by examining the effect of protease inhibitors on the T cell receptor-mediated death of the 2B4 murine T cell hybridoma and activated T cells. The cysteine protease inhibitors trans-epoxysuccininyl-L-leucylamido-(4-guanidino) butane (E-64) and leupeptin, the calpain selective inhibitor acetyl-leucyl-leucyl-normethional, and the serine protease inhibitors diisopropyl fluorophosphate and phenylmethylsulfonyl fluoride, all showed dose-dependent blocking of the 2B4 death response triggered by the T cell receptor complex and by anti-Thy-1. These protease inhibitors enhanced rather than inhibited IL-2 secretion triggered by T cell receptor cross-linking, showing that they did not act by preventing signal transduction. Growth inhibition induced by cross-linking the 2B4 T cell receptor, measured by inhibition of thymidine incorporation, was not generally blocked by these protease inhibitors. All five of these protease inhibitors enhanced rather than blocked 2B4 cell death triggered by dexamethasone, an agent previously shown to have a death pathway antagonistic with that of the TCR. 2B4 cytolysis by the cytotoxic agents staphylococcal alpha-toxin and dodecyl imidazole, and that caused by hypotonic conditions, was not significantly affected by the five protease inhibitors tested. The selected protease inhibitors blocked both the apoptotic nuclear morphology changes and DNA fragmentation induced by T cell receptor cross-linking, and enhanced both these properties induced by dexamethasone in 2B4 cells. The T cell receptor-induced death of activated murine lymph node T cells and human peripheral blood CD4+ T cells was blocked by both cysteine and serine protease inhibitors, showing that the protease-dependent death pathway also operates in these systems.
 ...
PMID:Protease inhibitors selectively block T cell receptor-triggered programmed cell death in a murine T cell hybridoma and activated peripheral T cells. 822 16

m-Calpain (calpain II, m-CANP), which normally requires millimolar Ca2+ for activity in vitro, was capable of proteolyzing a number of matrix proteins in isolated rat liver nuclei at Ca2+ concentrations as low as 3 microM (Mellgren, R. L. (1991) J. Biol. Chem. 266, 13920-13924). Treatment of nuclei with deoxyribonuclease I eliminated the activity of m-calpain at low Ca2+ concentrations, while ribonuclease A and phospholipase C had no effect. Addition of DNA to DNase-treated nuclei restored m-calpain activity at low Ca2+. RNA had little if any effect. Eukaryotic and prokaryotic DNA were equally effective, and synthetic polydeoxyribonucleotides were also activators. m-Calpain did not bind to a DNA-cellulose column in the presence of 200 microM Ca2+, and m-calpain preincubated in the presence of DNA and 200 microM Ca2+ was not activated at low Ca2+ concentrations following removal of the DNA. DNA did not alter the Ca2+ requirement for m-calpain-catalyzed cleavage of casein. These results demonstrate that the Ca2+ requirement for proteolysis of nuclear matrix proteins by m-calpain can be dramatically decreased in the presence of DNA. Activation did not seem to be a result of DNA binding directly to calpain but appeared to require interaction of DNA, calpain, and calpain substrates in the nuclear matrix.
 ...
PMID:m-Calpain requires DNA for activity on nuclear proteins at low calcium concentrations. 841 68

The 150-kDa phospholipase C (PLC)-beta 1 and three immunologically related proteins with molecular sizes of 140, 100, and 45 kDa were purified from bovine brain extracts. Determination of the amino-terminal amino acid sequence of the 45-kDa protein and immunoblots of the purified proteins with sequence-specific antibodies to peptides corresponding to three different regions of PLC-beta 1 suggest that a single cleavage at the linkage between amino acid residues 880 and 881 of PLC-beta 1 generates the 100- and 45-kDa proteins, which correspond to the amino-terminal and carboxyl-terminal portions, respectively, of PLC-beta 1. The Ca(2+)-dependent protease calpain appears to be responsible for the cleavage of PLC-beta 1; the PLC-beta 1 amino acid sequence contains PEST sequences which are common to proteins susceptible to calpain, and limited proteolysis of purified PLC-beta 1 by calpain generated a 100-kDa protein and a 40-kDa protein that contains the same amino-terminal sequence as the 45-kDa protein. The 140-kDa protein lacks the carboxyl-terminal-most region of PLC-beta 1, but there is no evidence it is derived from PLC-beta 1 by proteolysis. Cleavage of PLC-beta 1 by calpain had no significant effect on catalytic activity measured in the absence of the alpha subunit of the G alpha q but completely abrogated the stimulatory effect of G alpha q. On the other hand, G alpha q activated the 140-kDa enzyme. These results suggest that the region between residue 881 and the most carboxyl-terminal 10 kDa of PLC-beta 1 contains the G alpha q interaction site.
 ...
PMID:Removal of the carboxyl-terminal region of phospholipase C-beta 1 by calpain abolishes activation by G alpha q. 842 45

Murine peritoneal exudate macrophages (PEM) co-express granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage CSF (M-CSF) receptors, among others. Treatment of PEM with lipopolysaccharide (LPS) or tumor-promoting phorbol ester (12-O-tetradecanoylphorbol-13-acetate [TPA]) induces a rapid but transient loss of M-CSF receptors in PEM. GM-CSF receptors are not affected by this treatment. The loss of M-CSF receptors induced by LPS can be inhibited by neomycin and compound 48/80, two potent phospholipase C (PLC) inhibitors, but not by phospholipase A2, calpain, protein kinase C (PKC) or protease inhibitors. On the other hand, the loss of M-CSF receptors induced by TPA has been prevented by PKC inhibitors but not by PLC inhibitors. PLC inhibitors also prevent LPS-suppressed receptor-mediated internalization of radiolabeled recombinant human (rh) M-CSF by macrophages. Similar prevention of LPS-induced M-CSF receptor downregulation was observed in human monocytes that had been pretreated with PLC inhibitors. Our results show that 1) TPA-induced M-CSF receptor loss is strictly dependent on PKC activation; 2) PLC activation alone also leads to downregulation of M-CSF receptors; and 3) LPS-induced M-CSF receptor downregulation in PEM is mediated primarily through a PLC-dependent pathway. Our data also imply that the expression of M-CSF but not GM-CSF receptors is linked to an important, yet unknown, PLC-sensitive component(s) whose hydrolysis may lead to downregulation of M-CSF receptors.
 ...
PMID:Downregulation of M-CSF receptors by lipopolysaccharide in murine peritoneal exudate macrophages is mediated through a phospholipase C dependent pathway. 851 62

Thrombopoietin has an essential role in megakaryopoiesis and thrombopoiesis. To investigate the signaling processes induced by thrombopoietin, we have employed human platelets and recently demonstrated that thrombopoietin induces rapid tyrosine phosphorylation of Jak-2, Tyk2, Shc, Stat3, Stat5, p120(c-cbl) and other proteins in human platelets. Because the apparent molecular weight of a major tyrosine phosphorylated protein in platelets stimulated by thrombopoietin is approximately 85 to 95 kD, we examined the possibility that this could be Vav, a 95-kD proto-oncogene product. Specific antisera against Vav recognized the same 95 kD protein in lysates of Jurkat cells, which are known to express Vav, and platelets, indicating that platelets have Vav. Thrombopoietin induced rapid tyrosine phosphorylation of Vav in platelets without an elevation in cytosolic free calcium concentration or activation of protein kinase C. Vav was also tyrosine phosphorylated upon treatment of platelets with thrombin, collagen, or U46619, which activate phospholipase C, leading to an increased ionized calcium concentration and activation of protein kinase C. Ionomycin or phorbol 12-myristate 13-acetate (PMA) also induces tyrosine phosphorylation of Vav, suggesting that an increase in ionized calcium concentration or activation of protein kinase C may lead to phosphorylation of Vav. Thrombopoietin also induced tyrosine phosphorylation of Vav in FDCP-2 cells, genetically engineered to express human c-Mpl (FDCP-hMpl5). However, neither ionomycin nor PMA induced an increase in tyrosine phosphorylation of Vav in FDCP-hMpl5 cells, suggesting that the calcium and protein kinase C pathways of Vav phosphorylation may be unique to platelets. Further, Vav became incorporated into the Triton X-100 insoluble 10,000 g sedimentable residue in an aggregation-dependent manner, suggesting that it may have a regulatory role in platelet cytoskeletal processes. Vav was constitutively associated with a 28-kD adapter protein, Grb2, which is also incorporated into the cytoskeleton in an aggregation-dependent fashion. Lastly, we found that Vav is cleaved when there is activation of calpain, a protease that may have a role in postaggregation signaling processes. Our data suggest that thrombopoietin and other agonists may induce tyrosine phosphorylation of Vav by different mechanisms and Vav may also be involved in signaling during platelet aggregation by its redistribution to the cytoskeleton.
 ...
PMID:Thrombopoietin and thrombin induce tyrosine phosphorylation of Vav in human blood platelets. 910 97


1 2 3 4 Next >>