Understanding how Mutations in Non-Muscle Myosin 2A Drive Human Disease

Abstract

Non-muscle myosin 2A (NM2A) is a ubiquitously expressed myosin important for organizing actin filaments in areas of cell protrusion, cell adhesion, cell migration and cytokinesis and it is essential for early embryogenesis. Dephosphorylation of its regulatory light chain promotes formation of a compact inactive, ‘shutdown’ state in which the filament-forming tail is folded onto one of the two heads (known as the “10S” molecule). In this state, molecules cannot form filaments and the motors are inactivated. Over 80 mutations in the gene that encodes the heavy chain for NM2A (MYH9) have been described, most of which are single amino acid substitutions. Collectively these cause an autosomal-dominant disorder known as MYH9 disease, which result in a combination of symptoms from clotting disorders in which platelets are enlarged and bleeding time prolonged, aggregation of NM2A in neutrophils, cataracts, deafness and glomerulosclerosis. Here, using the cryo-electron microscopy structure we recently solved of shutdown smooth muscle myosin, we map known MYH9 mutations onto a homology model of NM2A. Multiple sequence alignment analysis reveals a high degree of conservation for residues involved in 10S-stabilising ionic interactions as well as many known mutation sites. Light meromyosin constructs (LMM is the region of the coiled-coil that assembles to form filaments) have been expressed, purified, and the effects of selected mutations have been assessed via negative stain electron microscopy. We suspect that these mutations may perturb the equilibrium between filamentous and 10S inactive molecules by actively destabilising the 10S conformation or by disrupting natural filament packing. With these results, we provide new structural insight on how mutations result in NM2A dysregulation and its implication in disease.