Evidence that a steric clash in the upper 50KDa domain of the motor Myo2p leads to cytokinesis defects in fission yeast

Abstract

Cytokinesis in many eukaryotes requires a contractile actomyosin ring that is placed at the division site. In fission yeast, an attractive organism for the study of cytokinesis, actomyosin ring assembly and contraction requires the myosin II heavy chain, Myo2p. Although myo2-E1, a temperature-sensitive mutant defective in the upper 50KDa domain of Myo2p has been studied extensively, the molecular basis of cytokinesis defect is not understood. Here we isolate myo2-E1-Sup2, an intragenic suppressor that contains the original mutation in myo2-E1 (G345R) and a second mutation in the upper 50KDa domain (Y297C). Unlike myo2-E1-Sup1, a previously characterized myo2-E1 suppressor, myo2-E1-Sup2 reverses actomyosin ring contraction defects in vitro and in vivo. Structural analysis of available myosin motor domain conformations suggests that a steric clash in myo2-E1, due to the replacement of a glycine with a bulky arginine, is relieved in myo2-E1-Sup2, by mutation of a tyrosine with the smaller cysteine. Our work provides insight into the function of the upper 50KDa domain of Myo2p, informs a molecular basis for the cytokinesis defect in myo2-E1, and may be relevant to understand certain cardiomyopathies.