Heavy chain phosphorylation of Dictyostelium myosin inhibited its ability to self-assemble into thick filaments (10-20% assembly vs 70-90% when dephosphorylated) and decreased actin-activated ATPase.
Heavy chain phosphorylation inhibits the self-assembly of Dictyostelium myosin into thick filaments, suggesting a regulatory mechanism for cell contractile events.
Dictyostelium myosin is composed of two heavy chains and two pairs of light chains in a 1:1:1 stoichiometry. Myosin purified from amoebae grown in medium containing 32Pphosphate had two of the subunits labeled (0.2-0.3 mol of phosphate per mol of 210,000-dalton heavy chains and approximately 0.1 mol of phosphate per mol of 18,000-dalton light chain). Kinase activities specific for the 210,000-dalton and for the 18,000-dalton subunits have been identified in extracts of Dictyostelium amoebae, and the heavy chain kinase has been purified 50-fold. This kinase phosphorylated Dictyostelium myosin to a maximum of 0.5-1.0 mol of phosphate per mol of heavy chain. Heavy chain phosphate, but not light chain phosphate, can be removed with bacterial alkaline phosphatase. Actin-activated myosin ATPase increased 80% when phosphorylated myosin was dephosphorylated to a level of approximately 0.06 mol of phosphate per mol of heavy chain. This effect could be reversed by rephosphorylating the myosin. The ability of myosin to self-assemble into thick filaments was inhibited by heavy chain phosphorylation. For example, in 80-100 mM KCl, only 10-20% of the myosin was assembled into thick filaments when the heavy chains were fully phosphorylated. Removal of the heavy chain phosphate resulted in 70-90% thick filament formation. This effect on self-assembly could be reversed by rephosphorylating the dephosphorylated myosin. These findings suggest that heavy chain phosphorylation may regulate cell contractile events by altering the state of myosin assembly.
Kuczmarski et al. (Mon,) reported a other. Heavy chain phosphorylation vs. Dephosphorylation was evaluated on Myosin self-assembly into thick filaments and actin-activated myosin ATPase activity. Heavy chain phosphorylation of Dictyostelium myosin inhibited its ability to self-assemble into thick filaments (10-20% assembly vs 70-90% when dephosphorylated) and decreased actin-activated ATPase.