Molecular Mechanism of the Inhibition of Phospholipase C β3 by Protein Kinase C

• Published in: The Journal of biological chemistry 2000-09, Vol.275 (39), p.30220
• Main Authors: Caiping Yue, Chun-Ying Ku, Mingyao Liu, Melvin I. Simon, Barbara M. Sanborn
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M004276200

Activation of protein kinase C (PKC) can result from stimulation of the receptor-G protein-phospholipase C (PLCβ) pathway. In turn, phosphorylation of PLCβ by PKC may play a role in the regulation of receptor-mediated phosphatidylinositide (PI) turnover and intracellular Ca 2+ release. Activation of endogenous PKC by phorbol 12-myristate 13-acetate inhibited both Gα q -coupled (oxytocin and M1 muscarinic) and Gα i -coupled (formyl-Met-Leu-Phe) receptor-stimulated PI turnover by 50–100% in PHM1, HeLa, COSM6, and RBL-2H3 cells expressing PLCβ 3 . Activation of conventional PKCs with thymeleatoxin similarly inhibited oxytocin or formyl-Met-Leu-Phe receptor-stimulated PI turnover. The PKC inhibitory effect was also observed when PLCβ 3 was stimulated directly by Gα q or Gβγ in overexpression assays. PKC phosphorylated PLCβ 3 at the same predominant site in vivo and in vitro . Peptide sequencing of in vitro phosphorylated recombinant PLCβ 3 and site-directed mutagenesis identified Ser 1105 as the predominant phosphorylation site. Ser 1105 is also phosphorylated by protein kinase A (PKA; Yue, C., Dodge, K. L., Weber, G., and Sanborn, B. M. (1998) J. Biol. Chem. 273, 18023–18027). Similar to PKA, the inhibition by PKC of Gα q -stimulated PLCβ 3 activity was completely abolished by mutation of Ser 1105 to Ala. In contrast, mutation of Ser 1105 or Ser 26 , another putative phosphorylation target, to Ala had no effect on inhibition of Gβγ-stimulated PLCβ 3 activity by PKC or PKA. These data indicate that PKC and PKA act similarly in that they inhibit Gα q -stimulated PLCβ 3 as a result of phosphorylation of Ser 1105 . Moreover, PKC and PKA both inhibit Gβγ-stimulated activity by mechanisms that do not involve Ser 1105 .