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)

Approximately 98% of turkey erythrocyte phospholipase C (PLC) is cytosolic and is released by hypotonic lysis of the cells and extensive washing of the resultant erythrocyte ghosts. Well washed turkey erythrocyte ghosts retain a fraction of tightly associated PLC, which is activated by the P2y-purinergic receptor and G-protein present in ghost membranes. The particulate PLC is sufficient to couple to all the available purinergic receptor-regulated G-protein. In contrast to ghosts, turkey erythrocyte plasma membrane preparations contain no detectable PLC. To investigate the subcellular location of the ghost-associated PLC, cytoskeletons were prepared by Triton X-100 extraction of turkey erythrocyte ghosts. The ghost-associated PLC was quantitatively recovered in cytoskeleton preparations. Cytoskeleton-associated PLC was solubilized by sodium cholate extraction, partially purified, and shown to reconstitute with PLC-free plasma membrane preparations in an agonist and guanine nucleotide-dependent fashion, indicating that the cytoskeleton-associated PLC is G-protein-regulated. Dissociation of erythrocyte ghost cytoskeletons with the actin-binding protein DNase 1 resulted in a dose-dependent inhibition of agonist and guanine nucleotide-stimulated PLC responses in ghosts and caused release of PLC from ghost or cytoskeleton preparations. These data demonstrate the specific association of a receptor and G-protein-regulated PLC with a component of the detergent-insoluble cytoskeleton and indicate that the integrity of the actin cytoskeleton is important for localization and effective coupling of PLC to the relevant G-protein.
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PMID:Association of a receptor and G-protein-regulated phospholipase C with the cytoskeleton. 142 46

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.
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PMID:Persistent activation of platelet membrane phospholipase C by proteolytic action of trypsin and thrombin. 229 26

In yeast, the cortical actin cytoskeleton seems to specify sites of growth of the cell surface. Because the actin-binding protein ABP1p is associated with the cortical cytoskeleton of Saccharomyces cerevisiae, it might be involved in the spatial organization of cell surface growth. ABP1p is localized to the cortical cytoskeleton and its overproduction causes assembly of the cortical actin cytoskeleton at inappropriate sites on the cell surface, resulting in delocalized surface growth. We have now cloned and sequenced the gene encoding ABP1p. ABP1p is a novel protein with a 50 amino-acid C-terminal domain that is very similar to the SH3 domain in the non-catalytic region of nonreceptor tyrosine kinases (including those encoded by the proto-oncogenes c-src and c-abl), in phospholipase C gamma and in alpha-spectrin. We also identified an SH3-related motif in the actin-binding tail domain of myosin-I. The identification of SH3 domains in a family of otherwise unrelated proteins that associate with the membrane cytoskeleton indicates that this domain might serve to bring together signal transduction proteins and their targets or regulators, or both, in the membrane cytoskeleton.
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PMID:Homology of a yeast actin-binding protein to signal transduction proteins and myosin-I. 240 79

A 57 kDa protein (p57) was obtained during the study on phosphatidylinositol-specific phospholipase C. Its cDNA was isolated from calf spleen and human leukemia cell line HL60 libraries and cloned. In the primary structures of p57, they have two unique amino acid sequence motifs, a WD repeat and a leucine zipper motif. Furthermore, p57 shared sequence similarity (40%) with coronin, an actin-binding protein responsible for chemotaxis, cell motility, and cytokinesis of Dictyostelium discoideum, which has only the WD repeat. p57 also showed an actin-binding activity and was mainly expressed in immune tissues. From these results, we conclude that p57 is a coronin-like novel actin-binding protein in mammalian cells but may also have a different function from coronin.
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PMID:Molecular cloning of a novel actin-binding protein, p57, with a WD repeat and a leucine zipper motif. 775 84

Preceding studies have shown that the bulk of the ATP-dependent, inositol 1,4,5-trisphosphate (IP3)-sensitive Ca2+ store of hamster insulinoma (HIT) cells is located in microvilli on the cell surface. Similar results were obtained with isolated rat hepatocytes. Moreover, in vesicles of microvillar origin, passive fluxes of Ca2+, ATP, and IP3 occur through cation and anion channels, respectively, suggesting that Ca2+ storage is due to ATP-dependent Ca2+ binding to an intravesicular component. Here we demonstrate that F-actin may be a possible candidate for this function. ATP-actin monomers bind Ca2+ with high affinity (Kd = 2-8 nM) to their divalent cation binding sites. Polymerization of actin monomers decreases the rate constant for divalent cation exchange at this binding site by more than 3 orders of magnitude rendering bound cations nearly unavailable. F-actin-bound Ca2+ can be released by depolymerization and dissociation from Ca(2+)-ADP-actin monomers (Kd = 375 nM). We now provide additional evidence for the possible involvement of actin in Ca2+ storage. (1) Preincubation of surface-derived Ca(2+)-storing vesicles from HIT cells with the F-actin stabilizer, phalloidin, strongly inhibited ATP-dependent Ca2+ uptake, reducing the IP3-sensitive Ca2+ pool by 70%. Phalloidin, when added after the loading process, affected neither the amount of stored Ca2+ nor IP3 action on the store. (2) F-actin polymerized in the presence of Mg2+ in nominally Ca(2+)-free buffer still contained about half of the high affinity sites occupied with Ca2+ (Mg/Ca-F-actin). (3) Using the fura-2 technique, we found that in the presence of ATP, Mg/Ca-F-actin incorporated free Ca2+ at a relatively low rate. Short pulses of ultrasound (3-10 s) strongly accelerated Ca2+ uptake, decreasing free Ca2+ from 500 nM to below 100 nM. (4) In the presence of physiological levels of Mg2+ (0.5 mM), sonication liberated large amounts of Ca2+ from Mg/Ca-F-actin. (5) Ca-F-actin released bound Ca2+ at a very slow rate. Short ultrasonic pulses rapidly elevated free Ca2+ from about 50 nM up to 500 nM. (6) Small amounts of profilin, an actin-binding protein, released Ca2+ both from Ca- and Mg/Ca-F-actin and also inhibited uptake of Ca2+ into Mg/Ca-F-actin. (7) Phalloidin completely inhibited Ca-uptake into Mg/Ca-F-actin even during ultrasonic treatment. These findings suggest that Ca2+ storage may occur by addition of Ca-ATP-actin monomers to reactive ends of the polymer and emptying of this store by profilin-stimulated release of Ca-ADP-actin. Thus, receptor-operated Ca2+ signaling, initiated by phospholipase C activation, may proceed via the well-known phosphatidylinositol phosphate-regulated profilin/gelsolin pathway of actin reorganization/depolymerization. The importance of the proposed microvillar Ca2+ signaling system for living cells remains to be established.
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PMID:Calcium storage and release properties of F-actin: evidence for the involvement of F-actin in cellular calcium signaling. 889 81

In ileal absorptive cells, carbachol inhibits NaCl absorption and its component brush border Na+/H+ exchanger, acting via basolateral membrane receptors. This carbachol effect involves (i) activation of brush border phosphatidylinositol 4,5-bisphosphate-specific phospholipase C (PLC) activity and brush border but not basolateral membrane translocation of PLC-gamma1 (Khurana, S., Kreydiyyeh, S., Aronzon, A., Hoogerwerf, W. A., Rhee, S. G., Donowitz, M., and Cohen, M. E. (1996) Biochem. J. 313, 509-518); and (ii) brush border tyrosine kinase(s) because mucosal but not serosal addition of the tyrosine kinase inhibitor genistein prevents the carbachol-induced inhibition of NaCl absorption and brush border Na+/H+ exchange. In the present work we identify a pool of villin (a brush border actin-binding protein) in the microvillus membrane fraction of rabbit ileum; this pool of villin is tyrosine-phosphorylated and associates with brush border membrane PLC-gamma1. Villin is present both in the Triton X-100-soluble and -insoluble fractions of the brush border. The Triton X-100-soluble pool is approximately 4-fold smaller than the brush border pool of villin that is present in the Triton X-100-insoluble fraction. Only the villin present in the Triton X-100-soluble fraction of ileal villus brush border associates with PLC-gamma1 and is tyrosine-phosphorylated. Carbachol increases the tyrosine phosphorylation of villin rapidly (as early as 30 s) and transiently. Carbachol also increases the amount of tyrosine-phosphorylated villin that associates with PLC-gamma1. These studies demonstrate that carbachol effects on NaCl absorption are accompanied by an increase in brush border PLC-gamma1 association with villin and an increase in tyrosine phosphorylation of villin. To study the role of cytoskeletal rearrangement in carbachol-induced inhibition of NaCl absorption, we used the F-actin stabilizing drug jasplakinolide. Jasplakinolide prevents the carbachol inhibition of ileal NaCl absorption. This suggests that F-actin severing is necessary for carbachol to inhibit ileal villus NaCl absorption. Since villin is known to sever actin, these studies suggest a role for villin in the signaling cascade that begins at the basolateral membrane with carbachol binding to its receptor and ends at the apical membrane in inhibition of NaCl absorption.
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PMID:Ileal microvillar protein villin is tyrosine-phosphorylated and associates with PLC-gamma1. Role of cytoskeletal rearrangement in the carbachol-induced inhibition of ileal NaCl absorption. 937 90

Gelsolin, an actin-binding protein, shows a strong ability to bind to phosphatidylinositol 4,5-bisphosphate (PIP(2)). Here we showed in in vitro experiments that gelsolin inhibited recombinant phospholipase D1 (PLD1) and PLD2 activities but not the oleate-dependent PLD and that this inhibition was not reversed by increasing PIP(2) concentration. To investigate the role of gelsolin in agonist-mediated PLD activation, we used NIH 3T3 fibroblasts stably transfected with the cDNA for human cytosolic gelsolin. Gelsolin overexpression suppressed bradykinin-induced activation of phospholipase C (PLC) and PLD. On the other hand, sphingosine 1-phosphate (S1P)-induced PLD activation could not be modified by gelsolin overexpression, whereas PLC activation was suppressed. PLD activation by phorbol myristate acetate or Ca(2+) ionophore A23187 was not affected by gelsolin overexpression. Stimulation of control cells with either bradykinin or S1P caused translocation of protein kinase C (PKC) to the membranes. Translocation of PKC-alpha and PKC-beta1 but not PKC-epsilon was reduced in gelsolin-overexpressed cells, whereas phosphorylation of mitogen-activated protein kinase was not changed. S1P-induced PLC activation and mitogen-activated protein kinase phosphorylation were sensitive to pertussis toxin, but PLD response was insensitive to such treatment, suggesting that S1P induced PLD activation via certain G protein distinct from G(i) for PLC and mitogen-activated protein kinase pathway. Our results suggest that gelsolin modulates bradykinin-mediated PLD activation via suppression of PLC and PKC activities but did not affect S1P-mediated PLD activation.
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PMID:Differential phospholipase D activation by bradykinin and sphingosine 1-phosphate in NIH 3T3 fibroblasts overexpressing gelsolin. 1048 69

Cofilin, an actin-binding protein, plays an important role in the migration, phagocytosis, and superoxide production of activated phagocytes through cytoskeletal reorganization. In unstimulated phagocytes, cofilin is a major phosphoprotein. However, upon activation, the phosphoprotein is dephosphorylated and translocated from cytosol to plasma membranes. Only the unphosphorylated form of cofilin is an active form that binds actin, whereas the regulatory mechanisms of cofilin have not been elucidated. We found that 1-[6-[[17beta-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-dione (U73122), an inhibitor of phospholipase C (PLC), suppressed both opsonized zymosan (OZ)-induced dephosphorylation and translocation of cofilin in macrophage-like U937 cells at 4 microM concentration. OZ triggered an increase in inositol 1,4,5-trisphosphate (IP3), and U73122 inhibited it. 1-[6-[[17beta-3-Methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-pyrrodione-dione (U73343), which was employed as an inactive analogue, had no such inhibitory activities as did U73122. Furthermore, herbimycin A, an inhibitor of src-type tyrosine kinase, also inhibited OZ-triggered IP3 formation. These results suggest that the activity and localization of cofilin are regulated by PLC at the downstream of src-family tyrosine kinase.
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PMID:U73122 inhibits the dephosphorylation and translocation of cofilin in activated macrophage-like U937 cells. 1125 43

While there is circumstantial evidence to suggest a requirement for phospholipase C-gamma(1) (PLC-gamma(1)) in actin reorganization and cell migration, few studies have examined the direct mechanisms that link regulators of the actin cytoskeleton with this crucial signaling molecule. This study was aimed to examine the role that villin, an epithelial cell-specific actin-binding protein, and its ligand PLC-gamma(1) play in migration in intestinal and renal epithelial cell lines that endogenously or ectopically express human villin. Basal as well as epidermal growth factor (EGF)-stimulated cell migration was accompanied by tyrosine phosphorylation of villin and its association with PLC-gamma(1). Inhibition of villin phosphorylation prevented villin-PLC-gamma(1) complex formation as well as villin-induced cell migration. The absolute requirement for PLC-gamma(1) in villin-induced cell migration was demonstrated by measuring cell motility in PLC-gamma(1)(-/-) cells and by downregulation of endogenous PLC-gamma(1). EGF-stimulated direct interaction of villin with the Src homology domain 2 domain of PLC-gamma(1) at the plasma membrane was demonstrated in living cells by using fluorescence resonance energy transfer. These results demonstrate that villin provides an important link between the activation of phosphoinositide signal transduction pathway and epithelial cell migration.
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PMID:Obligatory role for phospholipase C-gamma(1) in villin-induced epithelial cell migration. 1722 14