Leavitt L M, Macaluso C R, Kim K S, Martin N P, Dumont M E
Department of Biochemistry and Biophysics, University of Rochester Medical Center, NY 14642, USA.
Mol Gen Genet. 1999 Jul;261(6):917-32. doi: 10.1007/s004380051039.
The alpha-mating pheromone receptor encoded by the STE2 gene of the yeast Saccharomyces cerevisiae is a G protein-coupled receptor (GPCR) that is homologous to the large family of GPCRs that mediate multiple types of signal transduction in mammals. We have screened libraries of mutant receptors to identify dominant negative alleles that are capable of interfering with the function of a co-expressed normal receptor. Two dominant negative alleles have been recovered in this manner. In addition, we find that previously isolated loss-of-function mutations in the alpha-factor receptor exhibit dominant negative effects. Detection of the dominant effects requires high-level expression of the mutant receptors but does not require a high ratio of mutant to normal receptors. Cellular levels of the normal receptors are not affected by co-expression of the dominant negative alleles. Expression of the mutant receptors does not interfere with constitutive signaling in a strain that lacks the G protein alpha subunit encoded by GPA1, indicating that interference with signaling occurs at the level of the receptor or the interacting G protein. Expression of increased levels of G protein subunits partially reverses the dominant negative effects. The dominant negative behavior of the mutant receptors is diminished by deletion of the SST2 gene, which encodes an RGS (Regulator of G protein Signaling) protein involved in desensitization of pheromone signaling. The most likely explanation for the dominant negative effects of the mutations appears to be the existence of an interaction between unactivated receptors and the trimeric G protein that titrates the G protein away from the normal receptors or renders the G protein insensitive to receptor activation. This interaction appears to be mediated by the SST2 gene product.
酿酒酵母STE2基因编码的α交配信息素受体是一种G蛋白偶联受体（GPCR），与在哺乳动物中介导多种信号转导的GPCR大家族同源。我们筛选了突变受体文库，以鉴定能够干扰共表达的正常受体功能的显性负等位基因。通过这种方式已获得两个显性负等位基因。此外，我们发现先前分离出的α因子受体功能丧失突变表现出显性负效应。显性效应的检测需要突变受体的高水平表达，但不需要突变受体与正常受体的高比例。正常受体的细胞水平不受显性负等位基因共表达的影响。突变受体的表达不会干扰缺乏GPA1编码的G蛋白α亚基的菌株中的组成型信号传导，这表明对信号传导的干扰发生在受体或相互作用的G蛋白水平。增加G蛋白亚基的表达水平可部分逆转显性负效应。通过缺失SST2基因可减少突变受体的显性负行为，SST2基因编码一种参与信息素信号脱敏的RGS（G蛋白信号调节剂）蛋白。突变的显性负效应最可能的解释似乎是未活化的受体与三聚体G蛋白之间存在相互作用，这种相互作用使G蛋白从正常受体上脱离或使G蛋白对受体激活不敏感。这种相互作用似乎是由SST2基因产物介导的。
