REGULATION OF PURIFIED SUBTYPES OF PHOSPHATIDYLINOSITOL-SPECIFIC PHOSPHOLIPASE-C BETA BY G-PROTEIN ALPHA-SUBUNIT AND BETA-GAMMA-SUBUNITS

Specific antisera were produced to peptides representing the carboxyl termini of three subtypes of phosphatidylinositol-specific phospholipase C (PIPLC) beta which have been identified by isolation of cDNAs (Kriz, R., Lin, L., Sultzman, L., Ellis, C., Heldin, C., Pawson, T., and Knopf, J. (1990) Ciba Found. Symp. 150, 112-127). Screening with the antisera indicates that PIPLC beta3 is present in a variety of cell lines and rat tissues, whereas the distribution of PIPLC beta1 and beta2 is more restricted. A combination of conventional and immunoaffinity chromatographic techniques was used to purify PIPLC beta1 and beta3 from rat brain membranes. PIPLC beta2 was purified from cytosol of HL60 cells. All three subtypes were activated by purified G protein alpha(q/11) subunits with the following relative efficacies: PIPLC beta3 greater-than-or-equal-to PIPLC beta1 >> PIPLC beta2. All three PIPLC subtypes were also activated by G protein betagamma subunits with varying efficacies. The presence of betagamma subunits depressed the ability of alpha(q/11) to activate PIPLC beta1 and beta3 at low Mg2+ concentrations (1 mM). At higher concentrations of Mg2+ (2 mM or greater), activation of PIPLC beta3, but not PIPLC beta1, by betagamma and alpha(q/11) became additive. PIPLC beta3 was activated by alpha(q/11) even in the presence of a saturating concentration of betagamma subunits. This indicates that there are separate sites for interaction of PIPLCs with G protein subunits and that this interaction differs depending on the enzyme subtype and the concentration of Mg2+.