ABLATION OF G(O) ALPHA-SUBUNIT RESULTS IN A TRANSFORMED PHENOTYPE ANDCONSTITUTIVELY ACTIVE PHOSPHATIDYLCHOLINE-SPECIFIC PHOSPHOLIPASE-C

Modulation of the components involved in mitogenic signaling cascades is critical to the regulation of cell growth. GTP-binding proteins and the stimulation of phosphatidylcholine (PC) hydrolysis have been show n to play major roles in these cascades, One of the enzymes involved i n PC hydrolysis, a PC-specific phospholipase C (PC-PLC) has received r elatively little attention, In this paper we examined the role of a pa rticular heterotrimeric GTP-binding protein, G(o), in the regulation o f cell growth and PC-PLC-mediated hydrolysis of PC in IIC9 fibroblasts . The G(o) alpha-subunit was ablated in IIC9 cells by stable expressio n of antisense RNA, These stably transfected cells acquired a transfor med phenotype as indicated by: (a) the formation of multiple foci in m onolayer cultures, (b) the acquisition of anchorage-independent growth in soft agar; and (c) an increased level of thymidine incorporation i n the absence of added mitogens. These data implicate G(o) alpha as a novel tumor suppressor, Interestingly, PC-PLC activity was constitutiv ely active in the G(o) alpha-ablated cells as evidenced by the chronic ally elevated levels of diacylglycerol and phosphorylcholine in the ab sence of growth factors, In contrast, basal activities of PG-phospholi pase D, phospholipase A(2), or phosphoinositol-PLC were not affected, These data demonstrate, for the first time, a role for G(o) in regulat ing cell growth and provide definitive evidence for the existence of a PC-PLC in eukaryotic cells, The data further indicate that a subunit of G(o), is involved is regulating this enzyme.