Heiman Maxwell G, Engel Alex, Walter Peter
Howard Hughes Medical Institute, Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158, USA.
J Cell Biol. 2007 Jan 15;176(2):209-22. doi: 10.1083/jcb.200609182. Epub 2007 Jan 8.
The molecular machines that mediate cell fusion are unknown. Previously, we identified a multispanning transmembrane protein, Prm1 (pheromone-regulated membrane protein 1), that acts during yeast mating (Heiman, M.G., and P. Walter. 2000. J. Cell Biol. 151:719-730). Without Prm1, a substantial fraction of mating pairs arrest with their plasma membranes tightly apposed yet unfused. In this study, we show that lack of the Golgi-resident protease Kex2 strongly enhances the cell fusion defect of Prm1-deficient mating pairs and causes a mild fusion defect in otherwise wild-type mating pairs. Lack of the Kex1 protease but not the Ste13 protease results in similar defects. Deltakex2 and Deltakex1 fusion defects were suppressed by osmotic support, a trait shared with mutants defective in cell wall remodeling. In contrast, other cell wall mutants do not enhance the Deltaprm1 fusion defect. Electron microscopy of Deltakex2-derived mating pairs revealed novel extracellular blebs at presumptive sites of fusion. Kex2 and Kex1 may promote cell fusion by proteolytically processing substrates that act in parallel to Prm1 as an alternative fusion machine, as cell wall components, or both.
介导细胞融合的分子机制尚不清楚。此前，我们鉴定出一种多跨膜蛋白，即信息素调节膜蛋白1（Prm1），它在酵母交配过程中发挥作用（海曼，M.G.，和P.沃尔特。2000年。《细胞生物学杂志》151:719 - 730）。没有Prm1，相当一部分交配细胞对在其质膜紧密贴靠但未融合的状态下停滞。在本研究中，我们表明缺乏高尔基体驻留蛋白酶Kex2会强烈加剧Prm1缺陷型交配细胞对的细胞融合缺陷，并在其他方面为野生型的交配细胞对中导致轻微的融合缺陷。缺乏Kex1蛋白酶而非Ste13蛋白酶会导致类似缺陷。Δkex2和Δkex1融合缺陷可被渗透支持所抑制，这是与细胞壁重塑缺陷型突变体共有的一个特征。相比之下，其他细胞壁突变体不会加剧Δprm1融合缺陷。对源自Δkex2的交配细胞对进行电子显微镜观察，发现在假定的融合位点出现了新的细胞外泡。Kex2和Kex1可能通过蛋白水解加工底物来促进细胞融合，这些底物作为替代融合机制、作为细胞壁成分或两者兼而有之，与Prm1并行发挥作用。
