Regulation of myosin self-assembly: phosphorylation of Dictyostelium heavy chain inhibits formation of thick filaments.

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Abstract
Dictyostelium myosin is composed of 2 H chains and 2 pairs of L chains in a 1:1:1 stoichiometry. Myosin purified from amoebae grown in medium containing [32P]phosphate had 2 subunits labeled (0.2-0.3 mol phosphate/mol of 210,000-dalton H chain and .apprxeq. 0.1 mol phosphate/mol of 18,000-dalton L chain). Kinase activities specific for the 210,000 dalton and for the 18,000 dalton subunits were identified in extracts of Dictyostelium amoebae and the H chain kinase were purified 50-fold. This kinase phosphorylated Dictyostelium myosin to a maximum of 0.5-1.0 mol phosphate/mol of H chain. H chain phosphate but not L chain phosphate can be removed with bacterial alkaline phosphatase. Actin-activated myosin ATPase increased 80% when phosphorylated myosin was dephosphorylated to a level of .apprxeq. 0.06 mol phosphate/mol of H chain. This effect was reversed by rephosphorylating the myosin. The ability of myosin to self-assemble into thick filaments was inhibited by H chain phosphorylation, e.g., in 80-100 mM KCl, only 10-20% of the myosin was assembled into thick filaments when the H chains were fully phosphorylated. Removal of the H chain phosphate resulted in 70-90% thick filament formation. This effect on self-assembly was reversed by rephosphorylating the dephosphorylated myosin. H chain phosphorylation may regulate cell contractile events by altering the state of myosin assembly.