The effects of replacing Sst2 with the heterologous RGS4 on polarization and mating in yeast.
The effects of replacing Sst2 with the heterologous RGS4 on polarization and mating in yeast.
复制标题
用异源 RGS4 替换 Sst2 对酵母极化和交配的影响。
DOI：
10.1016/j.bpj.2010.04.078
复制
发表时间：
2010
影响因子：
3.4
通讯作者：
Yi,Tau-Mu
中科院分区：
文献类型：
--
作者：
Tanaka,Hiromasa;Yi,Tau-Mu
RGS蛋白刺激异源三聚体G蛋白的失活。酵母RGS蛋白Sst 2在转录和转录后水平上都受到调控。我们将SST 2基因替换为人类RGS 4基因，该基因由催化结构域和N端膜附着肽组成，并将天然启动子（PSST 2）替换为异源四环素抑制性启动子（PTET）。然后，我们测量的替代信息素的敏感性，交配和极化的效果。虽然信息素敏感性基本正常，但在交配和极化方面存在差异。特别是，RGS 4取代菌株在高水平的α-因子下没有形成多个交配突起，而是形成了一个畸形突起，这通常会产生芽。我们提供的证据表明，这种表型的出现，因为不像Sst 2，RGS 4没有本地化的投影。我们使用数学建模认为，本地化的Sst 2的投影防止过多的G-蛋白激活信息素反应过程中。此外，建模和实验表明，Sst 2的剂量影响交配突起形成的频率。
RGS proteins stimulate the deactivation of heterotrimeric G-proteins. The yeast RGS protein Sst2 is regulated at both the transcriptional and posttranscriptional levels. We replaced theSST2gene with the distantly related humanRGS4gene, which consists of the catalytic domain and an N-terminal membrane attachment peptide, and replaced the native promoter (PSST2) with the heterologous tetracycline-repressible promoter (PTET). We then measured the effect of the substitutions on pheromone sensitivity, mating, and polarization. Although the pheromone sensitivity was essentially normal, there were differences in mating and polarization. In particular, theRGS4-substituted strains did not form multiple mating projections at high levels ofα-factor, but instead formed a single malformed projection, which frequently gave rise to a bud. We provide evidence that this phenotype arose because unlike Sst2, RGS4 did not localize to the projection. We use mathematical modeling to argue that localization of Sst2 to the projection prevents excess G-protein activation during the pheromone response. In addition, modeling and experiments demonstrate that the dose of Sst2 influences the frequency of mating projection formation.