Protein kinase C phosphorylates G12α and inhibits its interaction with Gβγ

Abstract

Of nine G protein α subunits examined, only 12 and αz 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 αz 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 αz were unaltered by phosphorylation, as was the capacity of αz 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 αz-mediated signaling pathways by preventing their association with βγ.