Effects of FSGS-associated mutations on the stability and function of myosin-1 in fission yeast

Abstract

Point mutations in the human MYO1E gene, encoding class I myosin Myo1e, are associated with focal segmental glomerulosclerosis (FSGS), a primary kidney disorder that leads to end-stage kidney disease. In this study, we used a simple model organism, fission yeast Schizosaccharomyces pombe, to test the effects of FSGS-associated mutations on myosin activity. Fission yeast has only one class I myosin, Myo1, which is involved in actin patch assembly at the sites of endocytosis. The amino acid residues mutated in individuals with FSGS are conserved between human Myo1e and yeast Myo1, which allowed us to introduce equivalent mutations into yeast myosin and use the resulting mutant strains for functional analysis. Yeast strains expressing mutant Myo1 exhibited defects in growth and endocytosis similar to those observed in the myo1 deletion strain. These mutations also disrupted Myo1 localization to endocytic actin patches and resulted in mis-localization of Myo1 to eisosomes, linear membrane microdomains found in yeast cells. Although both mutants examined in this study exhibited loss of function, one of these mutants was also characterized by the decreased protein stability. Thus, using the yeast model system, we were able to determine that the kidney-disease-associated mutations impair myosin functional activity and have differential effects on protein stability.

Membrane-cytoskeletal crosstalk mediated by myosin-I regulates adhesion turnover during phagocytosis

“…6a-d ). In addition, a point mutation that disrupts motor domain function also prevented myosin localization 45 (Supplementary Fig. 6b , c).…”

Human Myosin 1e tail but not motor domain replaces fission yeast Myo1 domains to support myosin-I function during endocytosis

“…However, we found that the Myo1e tail can successfully substitute for the Myo1 tail to support localization and function of myosin-I at endocytic sites, although contributions of individual tail domains differ between these two myosins. Altogether, this work provides novel insights into the roles of individual protein domains and demonstrates that fission yeast can be used as a model system to study the effects of disease-associated Myo1e mutations located in the tail (21).…”

“…On the other hand, performing human-yeast domain switching studies using a model system with a straightforward readout, such as yeast endocytosis, provides a solid foundation for future work aiming to characterize the effects of disease-associated mutations in specific myosin domains. In our previous work, the use of the yeast system for functional testing of the effects of kidney disease-associated HsMyo1e mutations was limited to conserved residues in the motor domain of SpMyo1 (21). By demonstrating that chimeras consisting of SpMyo1 motor and HsMyo1e tail functionally complement myo1Δ while retaining localization requirements similar to those observed for HsMyo1e in mammalian cells, this study expands the utility of S. pombe as a simple model to effectively study disease-associated mutations residing in the HsMyo1e tail.…”

“…The conservation of the overall body plan, binding partners, and functional activities between yeast and vertebrate long-tailed class I myosins suggests that the functions of individual protein domains may also be conserved. Given these similarities, we have previously used fission yeast Schizosaccharomyces pombe, which has only a single class I myosin Myo1 (7,8), to test the effects of point mutations in the Myo1 motor domain that are homologous to mutations found in the human Myo1e (21). Patients homozygous for these MYO1E mutations develop kidney disease (22,23).…”

Membrane-cytoskeleton mechanical feedback mediated by myosin-I controls phagocytic efficiency

“…In addition, a point mutation that disrupts motor domain function also prevented myosin localization ( Supplementary Fig. 4g-h; 45 ). Thus, long-tailed myosins (myo1e/f) appear to have a distinctive role at FcR adhesions.…”