Deshaies R J, Schekman R
J Cell Biol. 1987 Aug;105(2):633-45. doi: 10.1083/jcb.105.2.633.
We have devised a genetic selection for mutant yeast cells that fail to translocate secretory protein precursors into the lumen of the endoplasmic reticulum (ER). Mutant cells are selected by a procedure that requires a signal peptide-containing cytoplasmic enzyme chimera to remain in contact with the cytosol. This approach has uncovered a new secretory mutant, sec61, that is thermosensitive for growth and that accumulates multiple secretory and vacuolar precursor proteins that have not acquired any detectable posttranslational modifications associated with translocation into the ER. Preproteins that accumulate at the sec61 block sediment with the particulate fraction, but are exposed to the cytosol as judged by sensitivity to proteinase K. Thus, the sec61 mutation defines a gene that is required for an early cytoplasmic or ER membrane-associated step in protein translocation.
我们设计了一种针对突变酵母细胞的遗传筛选方法，这些细胞无法将分泌蛋白前体转运到内质网（ER）腔中。通过一种程序筛选突变细胞，该程序要求含有信号肽的胞质酶嵌合体保持与胞质溶胶接触。这种方法发现了一种新的分泌突变体sec61，它对生长具有温度敏感性，并且积累了多种未获得与转运到ER相关的任何可检测翻译后修饰的分泌和液泡前体蛋白。在sec61阻滞点积累的前体蛋白与颗粒部分一起沉降，但根据对蛋白酶K的敏感性判断，它们暴露于胞质溶胶中。因此，sec61突变定义了一个基因，该基因是蛋白质转运过程中早期胞质或ER膜相关步骤所必需的。
