Phosphoinositide metabolism in transmembrane signalling

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It is well established that many extracellular signals elicit the receptor-linked phosphoinositide turnover and thereby induce both Ca^<2+> mobilization and protein kinase C activation. The mechanisms of signalling from the cell surface receptors to the phospholipase C have not been clarified. In this research project, we have investigated the roles and actions of GTP-binding proteins (G proteins) in these transmembrane signalling mechanisms. First, we have purified and characterized the synaptic membrane phospholipase C. We have also separated multiple G proteins with Mr values of about 20.000 (small Mr G proteins) from several tissues including cerebrum, purified and characterized two novel G proteins designated as smg p25A and smg p21, in addition to c-Ki-ras and rho proteins. It has been suggested that the ras protein modifies the receptor-linked phosphoinositide turnover, we are currently attempting to reconstitute these ras and ras-related small Mr G proteins with the membrane phospholipase C and cell surface receptors in a cell-free system. We have also characterized the growth factor receptor-linked phosphoinositide turnover using several cell lines. In NIH/3T3 cells and cultured smooth muscle cells, the bombesin-and angiotensin II-induced phosphoinositide hydrolysis is inhibited by protein kinase C by uncoupling the G protein to the phospholipase C. In contrast, the platelet derived-growth factor-induced reaction is insensitive to protein kinase C in both cell types. These results together with our earlier observations suggest that there are various signalling pathways from the receptors to the phospholipase C.