d'Enfert C, Barlowe C, Nishikawa S, Nakano A, Schekman R
Department of Molecular and Cellular Biology, Howard Hughes Medical Institute, University of California, Berkeley 94720.
Mol Cell Biol. 1991 Nov;11(11):5727-34. doi: 10.1128/mcb.11.11.5727-5734.1991.
Sec12p is a membrane glycoprotein required for the formation of a vesicular intermediate in protein transport from the endoplasmic reticulum to the Golgi apparatus in Saccharomyces cerevisiae. Comparison of the N-linked glycosylation of Sec12p, a Sec12p-invertase hybrid protein, and a derivative of Sec12p lacking 71 carboxy-terminal amino acids showed that Sec12p is a type II membrane protein. Analysis of two truncated forms of Sec12p and of a temperature-sensitive mutant indicated that the C-terminal domain of Sec12p is not essential for protein transport, whereas the integrity and membrane attachment of the cytoplasmic N-terminal domain are essential. Expression of a soluble cytoplasmic domain dramatically inhibited the growth of a sec12 temperature-sensitive strain by increasing the transport defect at a normally permissive temperature. This growth inhibition as well as the sec12 temperature-sensitive defect were suppressed by the overproduction of Sar1p, a small GTP-binding protein that participates in protein transport. Sar1p membrane association was enhanced by elevated levels of Sec12p. These results suggest that the cytoplasmic domain of Sec12p interacts with Sar1p and that the complex may function to promote vesicle formation.
Sec12p是一种膜糖蛋白，在酿酒酵母中，它是蛋白质从内质网运输到高尔基体过程中形成囊泡中间体所必需的。对Sec12p、Sec12p-转化酶杂合蛋白以及缺失71个羧基末端氨基酸的Sec12p衍生物的N-连接糖基化进行比较，结果表明Sec12p是一种II型膜蛋白。对Sec12p的两种截短形式和一个温度敏感突变体的分析表明，Sec12p的C末端结构域对于蛋白质运输并非必不可少，而细胞质N末端结构域的完整性和膜附着则至关重要。可溶性细胞质结构域的表达通过在正常允许温度下增加运输缺陷，显著抑制了sec12温度敏感菌株的生长。Sar1p（一种参与蛋白质运输的小GTP结合蛋白）的过量表达抑制了这种生长抑制以及sec12温度敏感缺陷。Sec12p水平的升高增强了Sar1p与膜的结合。这些结果表明，Sec12p的细胞质结构域与Sar1p相互作用，并且该复合物可能起到促进囊泡形成的作用。
