Moores S L, Sabry J H, Spudich J A
Department of Biochemistry, Stanford University School of Medicine, CA 94305, USA.
Proc Natl Acad Sci U S A. 1996 Jan 9;93(1):443-6. doi: 10.1073/pnas.93.1.443.
Conventional myosin plays a key role in the cytoskeletal reorganization necessary for cytokinesis, migration, and morphological changes associated with development in nonmuscle cells. We have made a fusion between the green fluorescent protein (GFP) and the Dictyostelium discoideum myosin heavy chain (GFP-myosin). The unique Dictyostelium system allows us to test the GFP-tagged myosin for activity both in vivo and in vitro. Expression of GFP-myosin rescues all myosin null cell defects. Additionally, GFP-myosin purified from these cells exhibits the same ATPase activities and in vitro motility as wild-type myosin. GFP-myosin is concentrated in the cleavage furrow during cytokinesis and in the posterior cortex of migrating cells. Surprisingly, GFP-myosin concentration increases transiently in the tips of retracting pseudopods. Contrary to previous thinking, this suggests that conventional myosin may play an important role in the dynamics of pseudopods as well as filopodia, lamellipodia, and other cellular protrusions.
传统肌球蛋白在胞质分裂、迁移以及非肌肉细胞发育相关的形态变化所必需的细胞骨架重组中起关键作用。我们已将绿色荧光蛋白（GFP）与盘基网柄菌肌球蛋白重链（GFP-肌球蛋白）进行了融合。独特的盘基网柄菌系统使我们能够在体内和体外测试带有GFP标签的肌球蛋白的活性。GFP-肌球蛋白的表达挽救了所有肌球蛋白缺失细胞的缺陷。此外，从这些细胞中纯化的GFP-肌球蛋白表现出与野生型肌球蛋白相同的ATP酶活性和体外运动性。在胞质分裂期间，GFP-肌球蛋白集中在分裂沟中，而在迁移细胞的后皮层中也有集中。令人惊讶的是，在缩回的伪足尖端，GFP-肌球蛋白的浓度会短暂增加。与之前的想法相反，这表明传统肌球蛋白可能在伪足以及丝状伪足、片状伪足和其他细胞突起的动态变化中起重要作用。
