Premium
Phospholipase C, Protein Kinase C, Ca 2+ /Calmodulin‐Dependent Protein Kinase II, and Tyrosine Phosphorylation Are Involved in Carbachol‐Induced Phospholipase D Activation in Chinese Hamster Ovary Cells Expressing Muscarinic Acetylcholine Receptor of Caenorhabditis elegans
Author(s) -
Min Do Sik,
Cho Nam Jeong,
Yoon Shin Hee,
Lee Young Han,
Hahn SangJune,
Lee KweonHaeng,
Kim MyungSuk,
Jo YangHyeok
Publication year - 2000
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.2000.0750274.x
Subject(s) - protein kinase c , phospholipase c , tyrosine phosphorylation , calphostin c , phospholipase d , wortmannin , tyrosine kinase , microbiology and biotechnology , biology , chemistry , map kinase kinase kinase , signal transduction , protein kinase b
: Recently, we have isolated a cDNA encoding a muscarinic acetylcholine receptor (mAChR) from Caenorhabditis elegans . To investigate the regulation of phospholipase D (PLD) signaling via a muscarinic receptor, we generated stable transfected Chinese hamster ovary (CHO) cells that overexpress the mAChR of C. elegans (CHO‐GAR‐3). Carbachol (CCh) induced inositol phosphate formation and a significantly higher Ca 2+ elevation and stimulated PLD activity through the mAChR ; this was insensitive to pertussis toxin, but its activity was abolished by the phospholipase C (PLC) inhibitor U73122. Western blot analysis revealed several apparent tyrosine‐phosphorylated protein bands after CCh treatment. The CCh‐induced PLD activation and tyrosine phosphorylation were significantly reduced by the protein kinase C (PKC) inhibitor calphostin C and down‐regulation of PKC and the tyrosine kinase inhibitor genistein. Moreover, the Ca 2+ ‐calmodulin‐dependent protein kinase II (CaM kinase II) inhibitor KN62, in addition to chelation of extracellular or intracellular Ca 2+ by EGTA and BAPTA/AM, abolished CCh‐induced PLD activation and protein tyrosine phosphorylation. Taken together, these results suggest that the PLC/PKC‐PLD pathway and the CaM kinase II/tyrosine kinase‐PLD pathway are involved in the activation of PLD through mAChRs of C. elegans .