Schmitt H D, Puzicha M, Gallwitz D
Max Planck Institute for Biophysical Chemistry, Department of Molecular Genetics, Göttingen, Federal Republic of Germany.
Cell. 1988 May 20;53(4):635-47. doi: 10.1016/0092-8674(88)90579-x.
Intragenic mutations were isolated that suppressed the dominant-lethal phenotype of the YPT1ile121 mutant gene in a temperature-dependent fashion. Among different amino acid substitutions resulting from single point mutations, two, Ala161----Val (A161V) and Met165----Ile (M165I), restored the function of the YPT1ile121 mutant protein. Mutants expressing the YPT1ile121/val161 allele (ypt1ts) only, grew normally at temperatures up to 30 degrees C but were arrested at 37 degrees C. At the restrictive temperature, ypt1ts mutants accumulated ER membranes, small vesicles, and unprocessed invertase, and they exhibited cytoskeletal defects and an enhanced 45Ca2+ uptake. Similar alterations were seen in YPT1-depleted cells. The ypt1ts mutant cells could be rescued from growth arrest by increasing extracellular Ca2+, and, even at the permissive temperature, they displayed increased trifluoperazine sensitivity.
分离出了以温度依赖方式抑制YPT1ile121突变基因显性致死表型的基因内突变。在单点突变导致的不同氨基酸替换中，有两个，即丙氨酸161（Ala161）替换为缬氨酸（Val）（A161V）和甲硫氨酸165（Met165）替换为异亮氨酸（Ile）（M165I），恢复了YPT1ile121突变蛋白的功能。仅表达YPT1ile121/val161等位基因（ypt1ts）的突变体在高达30摄氏度的温度下正常生长，但在37摄氏度时停滞生长。在限制温度下，ypt1ts突变体积累内质网、小泡和未加工的转化酶，并且它们表现出细胞骨架缺陷和增强的45Ca2+摄取。在YPT1缺失的细胞中也观察到类似的变化。通过增加细胞外Ca2+，ypt1ts突变体细胞可以从生长停滞中挽救出来，并且即使在允许温度下，它们对三氟拉嗪的敏感性也增加。
