Ren Yixin, West-Foyle Hoku, Surcel Alexandra, Miller Christopher, Robinson Douglas N
Department of Cell Biology, Johns Hopkins University School of Medicine, Baltimore, MD 21205.
Department of Cell Biology, Johns Hopkins University School of Medicine, Baltimore, MD 21205 Summer Academic Research Experience, Johns Hopkins University School of Medicine, Baltimore, MD 21205.
Mol Biol Cell. 2014 Dec 15;25(25):4150-65. doi: 10.1091/mbc.E14-08-1322. Epub 2014 Oct 15.
How myosin II localizes to the cleavage furrow in Dictyostelium and metazoan cells remains largely unknown despite significant advances in understanding its regulation. We designed a genetic selection using cDNA library suppression of 3xAsp myosin II to identify factors involved in myosin cleavage furrow accumulation. The 3xAsp mutant is deficient in bipolar thick filament assembly, fails to accumulate at the cleavage furrow, cannot rescue myoII-null cytokinesis, and has impaired mechanosensitive accumulation. Eleven genes suppressed this dominant cytokinesis deficiency when 3xAsp was expressed in wild-type cells. 3xAsp myosin II's localization to the cleavage furrow was rescued by constructs encoding rcdBB, mmsdh, RMD1, actin, one novel protein, and a 14-3-3 hairpin. Further characterization showed that RMD1 is required for myosin II cleavage furrow accumulation, acting in parallel with mechanical stress. 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. Overall, myosin II accumulation is the result of multiple parallel and partially redundant pathways that comprise a cellular contractility control system.
尽管在理解肌球蛋白II的调控方面取得了重大进展，但肌球蛋白II如何定位于盘基网柄菌和后生动物细胞的分裂沟在很大程度上仍不清楚。我们设计了一种基因筛选方法，利用对3xAsp肌球蛋白II的cDNA文库抑制来鉴定参与肌球蛋白在分裂沟积累的因子。3xAsp突变体在双极粗丝组装方面存在缺陷，无法在分裂沟积累，不能挽救肌球蛋白II基因缺失的胞质分裂，并且机械敏感积累受损。当在野生型细胞中表达3xAsp时，11个基因抑制了这种显性胞质分裂缺陷。编码rcdBB、mmsdh、RMD1、肌动蛋白、一种新蛋白和一个14-3-3发夹的构建体挽救了3xAsp肌球蛋白II在分裂沟的定位。进一步的表征表明，RMD1是肌球蛋白II在分裂沟积累所必需的，它与机械应力并行起作用。对几种突变菌株的分析表明，子细胞对称所需的肌球蛋白II活性阈值与沟侵入动力学所需的阈值不同。最后，一种具有更长杠杆臂（2xELC）的工程化肌球蛋白II，产生了一种高度机械敏感的马达，也可以部分抑制基因内的3xAsp。总的来说，肌球蛋白II的积累是多个平行且部分冗余途径的结果，这些途径构成了一个细胞收缩控制系统。
