Protein kinase C phosphorylates G_12α and inhibits its interaction with G_βγ

Of nine G protein α subunits examined, only ₁₂ and α_z served as substrates for phosphorylation by various isoforms of protein kinase C in vitro. A close homolog of α₁₂, α₁₃, was not phosphorylated. Exposure of NTH 3T3 cells that stably express α₁₂ to phorbol 12-myristate 13-acetate also resulted in phosphorylation of the protein. Phosphorylation in vitro occurred near the amino terminus (probably Ser³⁸), and approximately 1 mol of phosphate was incorporated per mol of α₁₂. Although G protein heterotrimers containing either α₁₂ or α_z were poor substrates for phosphorylation, the isolated α subunits were phosphorylated equally well in their GDP- or GTPγS-bound forms. The guanine nucleotide binding properties of purified α₁₂ and α_z were unaltered by phosphorylation, as was the capacity of α_z to inhibit type V adenylyl cyclase. However, phosphorylation of either protein greatly reduced its affinity for G protein βγ subunits, consistent with the newly determined crystal structure of a G protein heterotrimer. We suggest that protein kinase C regulates α₁₂ and α_z-mediated signaling pathways by preventing their association with βγ.