Discrete subcellular localization of phosphoinositidase C β, γ and δ in PC12 rat pheochromocytoma cells

Phosphoinositidase C activity was revealed in nuclei isolated from PC12 rat pheochromocytoma cells incubated with tritiated phosphatidylinositol, phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate. Phosphoinositide breakdown was found to be optimal at neutral pH and Ca ++ concentrations ranging from endogenous levels to millimolar values. To characterize the enzymes involved, three monoclonal antibodies directed against the β, γ and δ phosphoinositidase C isoforms were employed. A combination of Western blot immunochemical analysis on cytoplasmic and nuclear fractions and of in situ immunocytochemistry on intact cells and isolated nuclei indicated that phosphoinositidase C γ, though predominantly cytoplasmic, was present in both cell compartments. On the contrary, phosphoinositidase C β was exclusively localized in the nucleus, whereas phosphoinositidase C δ was restricted to the cytoplasm. These data suggest that inositol lipid breakdown is controlled by different phosphoinositidase C isozymes in the various cell compartments, and support the notion that a separate phosphoinositide signalling system is located in the nucleus.