Pryciak P M, Hartwell L H
Department of Genetics, University of Washington, Seattle 98195-7360, USA.
Mol Cell Biol. 1996 Jun;16(6):2614-26. doi: 10.1128/MCB.16.6.2614.
Mating pheromones of Saccharomyces cerevisiae control both signal transduction events and changes in cell shape. The G beta gamma complex of the pheromone receptor-coupled G protein activates the signal transduction pathway, leading to transcriptional induction and cell cycle arrest, but how pheromone-dependent signalling leads to cell shape changes is unclear. We used a two-hybrid system to search for proteins that interact with the G beta gamma complex and that might be involved in cell shape changes. We identified the ankyrin repeat-containing protein Akr1p and show here that it interacts with the free G beta gamma complex. This interaction may be regulated by pheromone, since Akr1p is excluded from the G alpha beta gamma heterotrimer. Both haploid and diploid cells lacking Akr1p grow slowly and develop deformed buds or projections, suggesting that this protein participates in the control of cell shape. In addition, Akr1p has a negative influence on the pheromone response pathway. Epistasis analysis demonstrates that this negative effect does not act on the G beta gamma complex but instead affects the kinase cascade downstream of G beta gamma, so that the kinase Ste20p and components downstream of Ste20p (e.g., Ste11p and Ste7p) are partially activated in cells lacking Akr1p. Although the elevated signalling is eliminated by deletion of Ste20p (or components downstream of Ste20p), the growth and morphological abnormalities of cells lacking Akr1p are not rescued by deletion of any of the known pheromone response pathway components. We therefore propose that Akr1p negatively affects the activity of a protein that both controls cell shape and contributes to the pheromone response pathway upstream of Ste20p but downstream of G beta gamma. Specifically, because recent evidence suggests that Bem1p, Cdc24p, and Cdc42p can act in the pheromone response pathway, we suggest that Akr1p affects the functions of these proteins, by preventing them from activating mating-specific targets including the pheromone-responsive kinase cascade, until G beta gamma is activated by pheromone.
酿酒酵母的交配信息素可控制信号转导事件和细胞形态变化。信息素受体偶联G蛋白的Gβγ复合物激活信号转导途径，导致转录诱导和细胞周期停滞，但信息素依赖性信号传导如何导致细胞形态变化尚不清楚。我们使用双杂交系统寻找与Gβγ复合物相互作用且可能参与细胞形态变化的蛋白质。我们鉴定出含锚蛋白重复序列的蛋白质Akr1p，并在此表明它与游离的Gβγ复合物相互作用。这种相互作用可能受信息素调节，因为Akr1p被排除在Gαβγ异源三聚体之外。缺乏Akr1p的单倍体和二倍体细胞生长缓慢，并形成变形的芽或突起，表明该蛋白参与细胞形态的控制。此外，Akr1p对信息素反应途径有负面影响。上位性分析表明，这种负面影响不是作用于Gβγ复合物，而是影响Gβγ下游的激酶级联反应，因此在缺乏Akr1p的细胞中，激酶Ste20p和Ste20p下游的成分（如Ste11p和Ste7p）被部分激活。尽管通过缺失Ste20p（或Ste20p下游的成分）可消除升高的信号传导，但缺失任何已知的信息素反应途径成分都无法挽救缺乏Akr1p的细胞的生长和形态异常。因此，我们提出Akr1p对一种蛋白质的活性产生负面影响，该蛋白质既控制细胞形态，又在Ste20p上游但在Gβγ下游对信息素反应途径起作用。具体而言，因为最近的证据表明Bem1p、Cdc24p和Cdc42p可在信息素反应途径中起作用，我们认为Akr1p通过阻止它们激活包括信息素反应性激酶级联反应在内的交配特异性靶标来影响这些蛋白质的功能，直到Gβγ被信息素激活。
