Calcium ionophore-induced secretion from mast cells correlates with myosin light chain phosphorylation by protein kinase C.
Rat basophilic leukemia mast cells (RBL-2H3) secrete histamine when activated by Ag. This secretion correlates with increased phosphorylation of myosin light chain by protein kinase C (PKC). Calcium ionophores (A23187) also elicit secretion, which is enhanced by PMA. To analyze the roles of Ca2+ and PKC in the secretory process, A23187-induced myosin light chain phosphorylation was examined in the presence and absence of PMA. A23187-induced secretion correlated best with myosin light chain phosphorylation by PKC, not with phosphorylation by myosin light chain kinase (MLCK). A23187 induced the translocation to membranes of the alpha, beta, delta, and epsilon isozymes of PKC. PMA not only increased the phosphorylation of myosin light chains at PKC-specific sites (Ser1 and Ser2) but also at sites attributed to MLCK (Ser19 and Thr18-Ser19). A23187 plus PMA induced higher levels of secretion concomitantly with increased myosin light chain phosphorylation at the PKC-specific sites. However, there was little correlation between the translocation of specific PKC isozymes and the phosphorylation of myosin light chains by PKC. Activation induced a novel triphosphorylated form of myosin light chain with a higher level of phosphorylation at the diphosphorylated MLCK sites. Quantitation of A23187 plus PMA-induced myosin light chain phosphorylation revealed that phosphorylation at PKC sites increased from zero to 0.35 mol/mol, was little changed at the monophosphorylated MLCK site (0.30 mol/mol), and increased from zero to 0.06 mol/mol at the diphosphorylated MLCK sites. Therefore, Ca2+-induced secretion correlates best with myosin light chain phosphorylation by PKC, and diphosphorylation by MLCK is unlikely to contribute to secretion.
Ludowyke
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RI
Centre for Immunology, St. Vincent's Hospital, Sydney, New South Wales, Australia.
Scurr
L L
LL
McNally
C M
CM
D000818:Animals; D001665:Binding Sites; D000001:Calcimycin / Q000008:administration & dosage / Q000494:pharmacology*; D002460:Cell Line; D004347:Drug Interactions; D006636:Histamine Release / Q000187:drug effects* / Q000502:physiology; D007476:Ionophores / Q000008:administration & dosage / Q000494:pharmacology*; D007527:Isoenzymes / Q000378:metabolism; D008407:Mast Cells / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology*; D018994:Myosin Light Chains / Q000737:chemistry / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000008:administration & dosage / Q000494:pharmacology
D007476:Ionophores; D007527:Isoenzymes; D018994:Myosin Light Chains; D000001:Calcimycin; D011493:Protein Kinase C; D009219:Myosin-Light-Chain Kinase; D013755:Tetradecanoylphorbol Acetate