Isozyme-specific stimulation of phospholipase C-gamma2 by Rac GTPases.
The regulation of the two isoforms of phospholipase C-gamma, PLCgamma(1) and PLCgamma(2), by cell surface receptors involves protein tyrosine phosphorylation as well as interaction with adapter proteins and phosphatidylinositol 3,4,5-trisphosphate (PtdInsP(3)) generated by inositol phospholipid 3-kinases (PI3Ks). All three processes may lead to recruitment of the PLCgamma isozymes to the plasma membrane and/or stimulation of their catalytic activity. Recent evidence suggests that PLCgamma may also be regulated by Rho GTPases. In this study, PLCgamma(1) and PLCgamma(2) were reconstituted in intact cells and in a cell-free system with Rho GTPases to examine their influence on PLCgamma activity. PLCgamma(2), but not PLCgamma(1), was markedly activated in intact cells by constitutively active Rac1(G12V), Rac2(G12V), and Rac3(G12V) but not by Cdc42(G12V) and RhoA(G14V). The mechanism of PLCgamma(2) activation was apparently independent of phosphorylation of tyrosine residues known to be modified by PLCgamma(2)-activating protein-tyrosine kinases. Activation of PLCgamma(2) by Rac2(G12V) in intact cells coincided with a translocation of PLCgamma(2) from the soluble to the particulate fraction. PLCgamma isozyme-specific activation of PLCgamma(2) by Rac GTPases (Rac1 approximately Rac2 > Rac3), but not by Cdc42 or RhoA, was also observed in a cell-free system. Herein, activation of wild-type Rac GTPases with guanosine 5'-(3-O-thio)triphosphate caused a marked stimulation of PLCgamma(2) but had no effect on the activity of PLCgamma(1). PLCgamma(1) and PLCgamma(2) have previously been shown to be indiscriminately activated by PtdInsP(3) in vitro. Thus, the results suggest a novel mechanism of PLCgamma(2) activation by Rac GTPases involving neither protein tyrosine phosphorylation nor PI3K-mediated generation of PtdInsP(3).
Piechulek
Thomas
T
Department of Pharmacology and Toxicology, University of Ulm, Albert-Einstein-Allee 11, 89081 Ulm, Germany.
Rehlen
Tobias
T
Walliser
Claudia
C
Vatter
Petra
P
Moepps
Barbara
B
Gierschik
Peter
P
D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002474:Cell-Free System; D002522:Chlorocebus aethiops; D004789:Enzyme Activation; D006801:Humans; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D009154:Mutation; D051966:Phospholipase C gamma / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D020744:rac GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism; D000097766:RAC2 GTP-Binding Protein
D007527:Isoenzymes; D051966:Phospholipase C gamma; D011994:Recombinant Proteins; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein; D000097766:RAC2 GTP-Binding Protein; C411563:RAC3 protein, human