Genetic suppression of a phosphomimic myosin II identifies system-level factors that promote myosin II cleavage furrow accumulation.
Bottom Line: How myosin II localizes to the cleavage furrow in Dictyostelium and metazoan cells remains largely unknown despite significant advances in understanding its regulation.Analysis of several mutant strains revealed that different thresholds of myosin II activity are required for daughter cell symmetry than for furrow ingression dynamics.Finally, an engineered myosin II with a longer lever arm (2xELC), producing a highly mechanosensitive motor, could also partially suppress the intragenic 3xAsp.
Affiliation: Department of Cell Biology, Johns Hopkins University School of Medicine, Baltimore, MD 21205.Show MeSH
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Mentions: To ascertain the functional state of myosin II in the rmd1hp cells as well as in myoII::3xAsp cells rescued by RMD1, we measured the furrow ingression dynamics and symmetry of daughter cell sizes for these cells. None of the mutants was able to recover the near-exponential WT furrow ingression dynamics (Figure 7A). The products of cell division are two daughter cells, which for WT cells are highly symmetrical in size. To quantify this, we measured the two-dimensional cross-sectional area of the daughter cells and calculated the ratio of the larger cell to the smaller cell (Figure 7B). For WT, this ratio was 1.10 ± 0.021 (mean ± SEM), whereas for myoII- cells, the ratio increased to 1.34 ± 0.084 and was more broadly distributed. As compared with WT cells, depletion of rmd1 did not alter the daughter cell symmetry despite altering furrow ingression kinetics (Figures 5D and 7, A and B). Of interest, RMD1 did not improve the furrow ingression kinetics of myoII::3xAsp, which was identical to myoII- kinetics (Figure 7A), but it did increase the symmetry of the resulting daughter cells (Figure 7B). Thus daughter cell symmetry and furrow ingression kinetics appear to be established by different thresholds of myosin II activity: depletion of rmd1 from WT cells or expression of RMD1 in myoII::3xAsp cells causes the phenotype to converge to an intermediate level of myosin II function, where daughter cell symmetry is normal but furrow ingression kinetics is not (Figure 7E).
Affiliation: Department of Cell Biology, Johns Hopkins University School of Medicine, Baltimore, MD 21205.