Phosphorylation of brush border myosin I by protein kinase C is regulated by Ca(2+)-stimulated binding of myosin I to phosphatidylserine concerted with calmodulin dissociation.
Brush border myosin I from chicken intestine is phosphorylated in vitro by chicken intestinal epithelial cell protein kinase C. Phosphorylation on serine and threonine to a maximum of 0.93 mol of P/mol of myosin I occurs within an approximately 20 kDa region at the end of the COOH-terminal tail of the 119-kDa heavy chain. The effects of Ca2+ on myosin I phosphorylation by protein kinase C are complex, with up to 4-fold stimulation occurring at 0.5-3 microM Ca2+, and up to 80% inhibition occurring at 3-320 microM Ca2+. Phosphorylation required that brush border myosin I be in its phosphatidylserine vesicle-bound state. Previously unknown Ca2+ stimulation of brush border myosin I binding to phosphatidylserine vesicles was found to coincide with Ca2+ stimulation of phosphorylation. A myosin I proteolytic fragment lacking approximately 20 kDa of its tail retained Ca(2+)-stimulated binding, but showed reduced Ca(2+)-independent binding. Ca(2+)-dependent phosphatidylserine binding is apparently due to the concomitant phosphatidylserine-promoted, Ca(2+)-induced dissociation of up to three of the four calmodulin light chains from myosin I. Four highly basic putative calmodulin-binding sites in the Ca(2+)-dependent phosphatidylserine binding region of the heavy chain were identified based on the similarity in their sequence to the calmodulin- and phosphatidylserine-binding site of neuromodulin. Calmodulin dissociation is now shown to occur in the low micromolar Ca2+ concentration range and may regulate the association of brush border myosin I with membranes and its phosphorylation by protein kinase C.
Swanljung-Collins
H
H
Department of Biochemistry, Eastern Virginia Medical School, Norfolk 23501.
Collins
J H
JH
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000378:metabolism*; D002645:Chickens; D002918:Chymotrypsin / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D004848:Epithelium / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism; D007700:Kinetics; D008871:Microvilli / Q000378:metabolism*; D008969:Molecular Sequence Data; D009218:Myosins / Q000302:isolation & purification / Q000378:metabolism*; D010449:Peptide Mapping; D010718:Phosphatidylserines / Q000378:metabolism*; D010748:Phosphopeptides / Q000302:isolation & purification; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D012689:Sequence Homology, Nucleic Acid
D002147:Calmodulin; D010718:Phosphatidylserines; D010748:Phosphopeptides; D011493:Protein Kinase C; D002918:Chymotrypsin; D009218:Myosins; D002118:Calcium