Characterization of a G protein-activated phosphoinositide 3-kinase in vascular smooth muscle cell nuclei

Recent studies highlight the existence of an autonomous nuclear polyphospho inositide metabolism related to cellular proliferation and differentiation. However, only few data document the nuclear production of the putative sec ond messengers, the 3-phosphorylated phosphoinositides, by the phosphoinosi tide 3-kinase (PI3K). In the present paper, we examine whether GTP-binding proteins can directly modulate 3-phosphorylated phosphoinositide metabolism in membrane-free nuclei isolated from pig aorta smooth muscle cells (VSMCs ). In vitro PI3K assays performed without the addition of any exogenous sub strates revealed that guanosine 5 '-(gamma -thio)triphosphate (GTP gammaS) specifically stimulated the nuclear synthesis of phosphatidylinositol 3,4,5 -trisphosphate (PtdIns(3,4,5)P-3), whereas guanosine 5 '-(beta -thio)diphos phate was ineffective. PI3K inhibitors wortmannin and LY294002 prevented GT P gammaS-induced PtdIns(3,4,5)P-3 synthesis. Moreover, pertussis toxin inhi bited partially PtdIns(3,4,5)P-3 accumulation, suggesting that nuclear G(i) /G(0) proteins are involved in the activation of PI3K. Immunoblot experimen ts showed the presence of G alpha (0), proteins in VSMC nuclei. In contrast with previous reports, immunoblots and indirect immunofluorescence failed to detect the p85 alpha subunit of the heterodimeric PI3K within VSMC nucle i. By contrast, we have detected the presence of a 117-kDa protein immunolo gically related to the PI3K gamma. These results indicate the existence of a G protein-activated PI3K inside VSMC nucleus that might be involved in th e contol of VSMC proliferation and in the pathogenesis of vascular prolifer ative disorders.