DIFFERENTIAL REGULATION OF FORMYL PEPTIDE AND PLATELET-ACTIVATING-FACTOR RECEPTORS - ROLE OF PHOSPHOLIPASE C-BETA(3) PHOSPHORYLATION BY PROTEIN-KINASE-A

Formylated peptides (e.g. n-formyl-Met-Leu-Phe (fMLP)) and platelet-ac tivating factor (PAF) mediate chemotactic and cytotoxic responses in l eukocytes through receptors coupled to G proteins that activate phosph olipase C (PLC), In RBL-2H3 cells, fMLP utilizes a pertussis toxin (pt s)-sensitive G protein to activate PLC, whereas PAF utilizes a pts-ins ensitive G protein. Here we demonstrate that fMLP, but not PAF, enhanc ed intracellular cAMP levels via a ptx-sensitive mechanism. Protein ki nase A (PRA) inhibition by H-89 enhanced inositol phosphate formation stimulated by fMLP but not PAF, Furthermore, a membrane-permeable cAMP analog 8-(4-chlorophenylthio)-cAMP (cpt-cAMP) inhibited phosphoinosit ide hydrolysis and secretion stimulated by fMLP but not PAF. Both cpt- cAMP and fMLP stimulated PLC beta(3) phosphorylation in intact RBL cel ls. The purified catalytic subunit of PKA phosphorylated PLC beta(3) i mmunoprecipitated from RBL cell lysate, Pretreatment of intact cells w ith cpt-cAMP and fMLP, but not PAF, resulted in an inhibition of subse quent PLC beta(3) phosphorylation by PRA in vitro. These data demonstr ate that fMLP receptor, which couples to a ptx-sensitive G protein, ac tivates both PLC and cAMP production. The resulting PKA activation pho sphorylates PLC beta(3) and appears to block the ability of G(beta gam ma) to activate PLC, Thus, both fMLP and PAF generate stimulatory sign als for PLC beta(3), but only fMLP produces a PKA-dependent inhibitory signal. This suggests a novel mechanism for the bidirectional regulat ion of receptors which activate PLC by ptx-sensitive G proteins.