Govindan B, Bowser R, Novick P
Department of Cell Biology, Yale University School of Medicine, New Haven, Connecticut 06520-8002.
J Cell Biol. 1995 Mar;128(6):1055-68. doi: 10.1083/jcb.128.6.1055.
Previous studies have shown that temperature-sensitive, myo2-66 yeast arrest as large, unbudded cells that accumulate vesicles within their cytoplasm (Johnston, G. C., J. A. Prendergast, and R. A. Singer. 1991. J. Cell Biol. 113:539-551). In this study we show that myo2-66 is synthetically lethal in combination with a subset of the late-acting sec mutations. Thin section electron microscopy shows that the post-Golgi blocked secretory mutants, sec1-1 and sec6-4, rapidly accumulate vesicles in the bud, upon brief incubations at the restrictive temperature. In contrast, myo2-66 cells accumulate vesicles predominantly in the mother cell. Double mutant analysis also places Myo2 function in a post-Golgi stage of the secretory pathway. Despite the accumulation of vesicles in myo2-66 cells, pulse-chase studies show that the transit times of several secreted proteins, including invertase and alpha factor, as well as the vacuolar proteins, carboxy-peptidase Y and alkaline phosphatase, are normal. Therefore the vesicles which accumulate in this mutant may function on an exocytic pathway that transports a set of cargo proteins that is distinct from those analyzed. Our observations are consistent with a role for Myo2 in transporting a class of secretory vesicles from the mother cell along actin cables into the bud.
先前的研究表明，温度敏感型的myo2-66酵母会停滞成为大的、未出芽的细胞，这些细胞在其细胞质中积累小泡（约翰斯顿，G.C.，J.A.普伦德加斯特，和R.A.辛格。1991。《细胞生物学杂志》113:539-551）。在本研究中，我们表明myo2-66与一部分后期作用的sec突变组合时是合成致死的。超薄切片电子显微镜显示，高尔基体后阻断的分泌突变体sec1-1和sec6-4在限制温度下短暂孵育后，会在芽中迅速积累小泡。相比之下，myo2-66细胞主要在母细胞中积累小泡。双突变分析也将Myo2的功能定位在分泌途径的高尔基体后阶段。尽管myo2-66细胞中积累了小泡，但脉冲追踪研究表明，几种分泌蛋白的转运时间，包括转化酶和α因子，以及液泡蛋白羧肽酶Y和碱性磷酸酶，都是正常的。因此，在该突变体中积累的小泡可能在一条胞吐途径中发挥作用，该途径运输的一组货物蛋白与所分析的不同。我们的观察结果与Myo2在将一类分泌小泡从母细胞沿着肌动蛋白丝运输到芽中的作用一致。
