MYH9 mutation N93K modestly reduces NM2-A motor activity but significantly enhances filament stability and interaction with UNC45a, causing distinct cytoplasmic aggregation with co-localization of wild-type NM2-A and NM2-B.
Distinct molecular defects caused by different MYH9 mutations affect the distribution and function of the wild type allele differently, potentially explaining the varying severity of MYH9-related diseases.
Mutations in the MYH9 gene, which encodes the heavy chain of the actin-based molecular motor non-muscle myosin II-A (NM2-A), cause a spectrum of rare blood disorders collectively termed MYH9-related diseases (MYH9-RD). Previous data indicate that mutations in the motor domain result in more severe phenotypes than those in the dimerization/filamentation domain. Here, we show that N93K mutation, previously described as motor-impairing, has only a minor effect on myosin motor function in vitro, but it significantly enhances NM2-A filament stability and interaction with the chaperone UNC45a. When expressed in stress fiber-competent cells, e.g., COS7 or U2OS, and stress fiber-incompetent cells, e.g., MEG-01, NM2-A N93K formed amorphous aggregates that colocalized with wild-type NM2-A, NM2-B and UNC45a, but not F-actin, tropomyosin-4 or phosphoSer19, active, RLC. Another motor mutant, NM2-A R705H, caused milder defects in filament stability, adhesion maturation and aggregation in both types of cells. Conversely, the tail domain mutant NM2-A E1841K enhanced filament stability in COS7 cells without forming aggregates, while it promoted the formation of small, elongated aggregates in MEG-01 cells that did not co-localize with wild type NM2-A. These data indicate that the molecular defect caused by NM2A-N93K affects the distribution and function of the wild type allele, whereas NM2-A E1841K aggregates do not affect the wild type allele, which could constitute the molecular basis of the differences in severity in mutant MYH9 allele carriers.
Llorente-González et al. (Sat,) conducted a other in Human cell lines (COS7, U2OS, MEG-01) expressing MYH9 mutations related to MYH9-related diseases, a spectrum of rare blood disorders with macrothrombocytopenia and various extra-hematological complications. Expression of MYH9 mutant NM2-A variants N93K, R705H, and E1841K vs. Wild type NM2-A expression was evaluated on NM2-A filament stability, cellular localization, aggregation phenotypes and interaction with chaperone UNC45a. MYH9 mutation N93K modestly reduces NM2-A motor activity but significantly enhances filament stability and interaction with UNC45a, causing distinct cytoplasmic aggregation with co-localization of wild-type NM2-A and NM2-B.