Tiedje Christopher, Holland Daniel G, Just Ursula, Höfken Thomas
Institute of Biochemistry, Christian Albrecht University Kiel, Olshausenstrasse 40, 24098 Kiel, Germany.
J Cell Sci. 2007 Oct 15;120(Pt 20):3613-24. doi: 10.1242/jcs.009860. Epub 2007 Sep 25.
The Saccharomyces cerevisiae p21-activated kinase (PAK) Ste20 regulates various aspects of cell polarity during vegetative growth, mating and filamentous growth. To gain further insight into the mechanisms of Ste20 action, we screened for interactors of Ste20 using the split-ubiquitin system. Among the identified proteins were Erg4, Cbr1 and Ncp1, which are all involved in sterol biosynthesis. The interaction between Ste20 and Erg4, as well as between Ste20 and Cbr1, was confirmed by pull-down experiments. Deletion of either ERG4 or NCP1 resulted in various polarity defects, indicating a role for these proteins in bud site selection, apical bud growth, cell wall assembly, mating and invasive growth. Interestingly, Erg4 was required for the polarized localization of Ste20 during mating. Lack of CBR1 produced no detectable phenotype, whereas the deletion of CBR1 in the absence of NCP1 was lethal. Using a conditional lethal mutant we demonstrate that both proteins have overlapping functions in bud morphology.
酿酒酵母p21激活激酶（PAK）Ste20在营养生长、交配和丝状生长过程中调节细胞极性的各个方面。为了进一步深入了解Ste20的作用机制，我们使用分裂泛素系统筛选了Ste20的相互作用蛋白。在鉴定出的蛋白质中，有Erg4、Cbr1和Ncp1，它们都参与甾醇生物合成。通过下拉实验证实了Ste20与Erg4之间以及Ste20与Cbr1之间的相互作用。删除ERG4或NCP1会导致各种极性缺陷，表明这些蛋白质在芽位点选择、顶端芽生长、细胞壁组装、交配和侵袭性生长中发挥作用。有趣的是，交配过程中Ste20的极性定位需要Erg4。缺乏CBR1没有产生可检测到的表型，而在缺乏NCP1的情况下删除CBR1是致死的。使用条件致死突变体，我们证明这两种蛋白质在芽形态方面具有重叠功能。
