Oki M, Noguchi E, Hayashi N, Nishimoto T
Department of Molecular Biology, Graduate School of Medical Science, Kyushu University, Fukuoka, Japan.
Mol Gen Genet. 1998 Apr;257(6):624-34. doi: 10.1007/s004380050690.
A series of ts mutations in the GSP1 gene of Saccharomyces cerevisiae was isolated by error-prone PCR. A total of 25 ts gsp1 strains was obtained. Each of these mutants showed between one and seven different amino acid alterations. In several of these ts gsp1 strains, the same amino acid residues in Gsp1p were repeatedly mutated, indicating that our screen for ts gsp1 mutations was saturating. All of the ts gsp1 strains isolated had a defect in nuclear protein import, but only 16 of the 25 ts gsp1 strains had a defect in mRNA export. Thus, Gsp1p is suggested to be directly involved in nuclear protein import, but not in mRNA export. Following release from alpha-factor arrest, 11 of the ts gsp1 mutants arrested in G1; the remainder did not show any specific cell-cycle arrest, at 37 degrees C, the nonpermissive temperature. While the mutants that are defective in both mRNA export and protein import have a tendency to arrest in G1, there was no clear correlation between the cell cycle phenotype and the defects in mRNA export and nuclear protein import. Based on this, we assume that Ran/Gsp1p GTPase regulates the cell cycle and the nucleus/cytosol exchange of macromolecules through interactions with effectors that were independent of each other, and are differentially affected by mutation.
通过易错PCR分离出酿酒酵母GSP1基因中的一系列温度敏感（ts）突变。共获得25个ts gsp1菌株。这些突变体中的每一个都显示出一到七个不同的氨基酸改变。在其中几个ts gsp1菌株中，Gsp1p中的相同氨基酸残基反复发生突变，这表明我们对ts gsp1突变的筛选已趋于饱和。所有分离出的ts gsp1菌株在核蛋白输入方面都存在缺陷，但25个ts gsp1菌株中只有16个在mRNA输出方面存在缺陷。因此，推测Gsp1p直接参与核蛋白输入，而不参与mRNA输出。从α因子阻滞中释放后，11个ts gsp1突变体在G1期阻滞；其余的在37℃（非允许温度）下未表现出任何特定的细胞周期阻滞。虽然在mRNA输出和蛋白输入方面都有缺陷的突变体有在G1期阻滞的倾向，但细胞周期表型与mRNA输出和核蛋白输入缺陷之间没有明显的相关性。基于此，我们假设Ran/Gsp1p GTP酶通过与彼此独立且受突变影响不同的效应器相互作用来调节细胞周期和大分子的核/质交换。
