Xie Y, Varshavsky A
Division of Biology, 147-75, Caltech, 1200 East California Blvd., Pasadena, CA 91125, USA.
Curr Genet. 1999 Sep;36(3):113-23. doi: 10.1007/s002940050480.
The N-end rule pathway is a ubiquitin-dependent proteolytic system whose targets include proteins bearing destabilizing N-terminal residues. We carried out a synthetic lethal screen for Saccharomyces cerevisiae mutants that require the N-end rule pathway for cell viability. A mutant thus identified, termed sln2, could not grow in the absence of Ubr1p, the recognition component of the N-end rule pathway, which was not essential for viability of the parental strain under the same conditions. Further analysis showed that inviability of sln2ubr1Delta cells could be rescued either by the HIS3 gene (which was absent from the parental strain) or by a high concentration of histidine in the medium. This defect in histidine uptake, exhibited by the sln2 mutant in the absence but not in the presence of Ubr1p, was traced to the gene HIP1, which encodes the histidine transporter. HIP1 was underexpressed in sln2 ubr1Delta cells, in comparison to either sln2 UBR1 or SLN2 ubr1Delta cells. Yet another property of the sln2 mutant was its inviability at 37 degrees C, which could not be rescued by either UBR1 or HIS3. This feature of sln2 allowed the cloning of SLN2, which was found to be a gene called CIN8, encoding a kinesin-like protein. Thus, either the N-end rule pathway or Cin8p must be present for the viability-sustaining rate of histidine import in S. cerevisiae auxotrophic for histidine. We consider possible mechanisms of this previously unsuspected link between kinesins, ubiquitin-dependent proteolysis, and the import of histidine.
N端规则途径是一种依赖泛素的蛋白水解系统，其作用靶点包括带有不稳定N端残基的蛋白质。我们对酿酒酵母突变体进行了合成致死筛选，这些突变体需要N端规则途径来维持细胞活力。由此鉴定出的一个突变体，称为sln2，在缺乏Ubr1p（N端规则途径的识别成分）的情况下无法生长，而在相同条件下Ubr1p对亲本菌株的活力并非必需。进一步分析表明，sln2ubr1Δ细胞的 inviability 可以通过HIS3基因（亲本菌株中不存在）或培养基中高浓度的组氨酸来挽救。sln2突变体在缺乏Ubr1p时表现出的组氨酸摄取缺陷，而在有Ubr1p时则没有，这可追溯到编码组氨酸转运蛋白的基因HIP1。与sln2 UBR1或SLN2 ubr1Δ细胞相比，HIP1在sln2 ubr1Δ细胞中表达不足。sln2突变体的另一个特性是其在37℃时无法存活，这不能通过UBR1或HIS3来挽救。sln2的这一特性使得能够克隆出SLN2，发现它是一个名为CIN8的基因，编码一种类驱动蛋白。因此，对于组氨酸营养缺陷型的酿酒酵母，要维持组氨酸导入的活力维持率，必须存在N端规则途径或Cin8p。我们考虑了驱动蛋白、泛素依赖性蛋白水解和组氨酸导入之间这种先前未被怀疑的联系的可能机制。
