Cytokinesis in animal cells requires the assembly and constriction of a contractile actomyosin ring. Non-muscle myosin II is essential for cytokinesis, but the role of its motor activity remains unclear. Here, we examine cytokinesis in C. elegans embryos expressing non-muscle myosin motor mutants generated by genome editing. Two non-muscle motor-dead myosins capable of binding F-actin do not support cytokinesis in the one-cell embryo, and two partially motor-impaired myosins delay cytokinesis and render rings more sensitive to reduced myosin levels. Further analysis of myosin mutants suggests that it is myosin motor activity, and not myosin9s ability to crosslink F-actin, that drives the alignment and compaction of F-actin bundles during contractile ring assembly, and that myosin motor activity sets the pace of contractile ring constriction. We conclude that myosin motor activity is required at all stages of cytokinesis. Finally, characterization of the corresponding motor mutations in C. elegans major muscle myosin shows that motor activity is required for muscle contraction but dispensable for F-actin organization in adult muscles.

Cytokinesis in animal cells requires the assembly of a contractile actomyosin ring, whose subsequent constriction physically separates the two daughter cells. Non-muscle myosin II (myosin) is essential for cytokinesis, but the role of its motor activity remains poorly defined. Here, we examine cytokinesis in C. elegans one-cell embryos expressing myosin motor mutants generated by genome editing. Motor-dead myosin, which is capable of binding F-actin, does not support cytokinesis, and embryos co-expressing motor-dead and wild-type myosin are delayed in cytokinesis. Partially motor-impaired myosin also delays cytokinesis and renders contractile rings more sensitive to reduced myosin levels. Thus, myosin motor activity, rather than its ability to cross-link actin filaments, drives contractile ring assembly and constriction. We further demonstrate that myosin motor activity is required for long-range cortical actin flows, but that flows per se play a minor role in contractile ring assembly. Our results suggest that flow-independent recruitment of motor-competent myosin to the cell equator is both essential and rate-limiting for cytokinesis.

Primary ciliary dyskinesia (PCD) is a rare genetic disorder mainly characterized by impaired mucociliary clearance and chronic respiratory symptoms. From a consanguineous family, a male patient, although with respiratory complaints since birth, was diagnosed with PCD only in adulthood. Whole-exome sequencing disclosed a novel homozygous intronic single-nucleotide deletion, NM_001369.3(DNAH5):c.13723+4del, initially classified as of uncertain clinical significance. Digital highspeed videomicroscopy (HSVM) evidenced a null ciliary beating frequency; transmission electron microscopy showed absence of outer dynein arms (class-1); and immunofluorescence (IF) demonstrated markedly absent DNAH5 protein level in the apical cilia region with delocalization to the transition and basal-body regions. Bioinformatic analysis predicted altered splicing at the donor splice site of exon 78, whereas mRNA sequencing revealed two splicing defects: the mainly expressed transcript corresponding to exon 78 skipping and a minor transcript originated from a cryptic splice site in exon 78. The patient was infertile and showed severe oligoteratozoospermia. Sperm IF analysis revealed absence of DNAH5 from the flagellum with accumulation at the neck region. The family study confirmed homozygosity. The present results support a pathogenic role for the c.13723+4del variant and underscore the importance of integrating clinical, ultrastructural, DNA, mRNA and protein analyses to clarify and contribute to PCD diagnosis.