MLCK-A, an unconventional myosin light chain kinase from Dictyostelium, is activated by a cGMP-dependent pathway.
Dictyostelium myosin II is activated by phosphorylation of its regulatory light chain by myosin light chain kinase A (MLCK-A), an unconventional MLCK that is not regulated by Ca2+/calmodulin. MLCK-A is activated by autophosphorylation of threonine-289 outside of the catalytic domain and by phosphorylation of threonine-166 in the activation loop by an unidentified kinase, but the signals controlling these phosphorylations are unknown. Treatment of cells with Con A results in quantitative phosphorylation of the regulatory light chain by MLCK-A, providing an opportunity to study MLCK-A's activation mechanism. MLCK-A does not alter its cellular location upon treatment of cells with Con A, nor does it localize to the myosin-rich caps that form after treatment. However, MLCK-A activity rapidly increases 2- to 13-fold when Dictyostelium cells are exposed to Con A. This activation can occur in the absence of MLCK-A autophosphorylation. cGMP is a promising candidate for an intracellular messenger mediating Con A-triggered MLCK-A activation, as addition of cGMP to fresh Dictyostelium lysates increases MLCK-A activity 3- to 12-fold. The specific activity of MLCK-A in cGMP-treated lysates is 210-fold higher than that of recombinant MLCK-A, which is fully autophosphorylated, but lacks threonine-166 phosphorylation. Purified MLCK-A is not directly activated by cGMP, indicating that additional cellular factors, perhaps a kinase that phosphorylates threonine-166, are involved.
Silveira
L A
LA
Department of Biochemistry, Beckman Center, Stanford University Medical School, Stanford, CA 94305-5307, USA.
Smith
J L
JL
Tan
J L
JL
Spudich
J A
JA
D000205:Actomyosin / Q000378:metabolism; D000818:Animals; D003208:Concanavalin A / Q000494:pharmacology; D006152:Cyclic GMP / Q000378:metabolism*; D004023:Dictyostelium / Q000187:drug effects / Q000201:enzymology / Q000502:physiology*; D004789:Enzyme Activation; D007700:Kinetics; D018994:Myosin Light Chains / Q000302:isolation & purification / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D010766:Phosphorylation; D015800:Protozoan Proteins
D018994:Myosin Light Chains; D015800:Protozoan Proteins; D003208:Concanavalin A; D000205:Actomyosin; C115459:MLCK-A protein, Dictyostelium discoideum; D009219:Myosin-Light-Chain Kinase; D006152:Cyclic GMP