The Enzymatic Motor Activity of Nonmuscle Myosin II-B is not Critical for Cardiac Myocyte Cytokinesis

Abstract

Nonmuscle myosin (NM) II is believed to drive contractile ring constriction during cytokinesis and NM II-B is the major nonmuscle myosin II isoform expressed in cardiac myocytes. Ablation of NM II-B in mice resulted in premature binucleation in cardiac myocytes consistent with a role for NM II-B in cytokinesis. Surprisingly binucleation can be rescued by expressing a motor-impaired mutant form of NM II-B (R709C) in the mouse heart, suggesting that NM II-B enzymatic motor activity is not essential for cytokinesis. We tested this hypothesis directly by using a COS-7 cell line which resembles cardiac myocytes in containing only NM II-B and small quantities of NM II-C (<7%) but no NM II-A. Previous work had shown that using siRNA to deplete NMHC II-B in this cell line resulted in multinucleation due to a failure in cytokinesis. We now show that COS-7 cell multinucleation can be rescued by transfecting with mutant forms of NM II-B (R709C) or NM II-A (N93K). These two mutant NM IIs have previously been shown to have marked reductions in their actin-activated MgATPase activity (70% in the case of NM II-B and 96% in the case of NM II-A) and neither could propel actin filaments in an in vitro motility assay. Importantly, cytokinesis is inhibited by the myosin II inhibitor blebbistatin in COS-7 cells expressing either wild-type NM II-B or the mutant form of NM II-A (or II-B). Whereas blebbistatin blocks myosin in an actin-detached state, the mutant NM IIs (R709C II-B and N93K II-A) have no defect in their actin-binding activity. Our results therefore argue that the role of NM II in cytokinesis depends more on its actin binding (cross-linking) property than its enzymatic Mg2+-ATPase activity.