De Filippi Loic, Fournier Margot, Cameroni Elisabetta, Linder Patrick, De Virgilio Claudio, Foti Michelangelo, Deloche Olivier
Département de Microbiologie et Médecine Moléculaire, Centre Médical Universitaire, Université de Genève, 1 rue Michel-Servet, 1211 Geneva 4, Switzerland.
Curr Genet. 2007 Sep;52(3-4):171-85. doi: 10.1007/s00294-007-0151-0. Epub 2007 Aug 21.
Chlorpromazine (CPZ) is a small permeable cationic amphiphilic molecule that inserts into membrane bilayers and binds to anionic lipids such as poly-phosphoinositides (PIs). Since PIs play important roles in many cellular processes, including signaling and membrane trafficking pathways, it has been proposed that CPZ affects cellular growth functions by preventing the recruitment of proteins with specific PI-binding domains. In this study, we have investigated the biological effects of CPZ in the yeast Saccharomyces cerevisiae. We screened a collection of approximately 4,800 gene knockout mutants, and found that mutants defective in membrane trafficking between the late-Golgi and endosomal compartments are highly sensitive to CPZ. Microscopy and transport analyses revealed that CPZ affects membrane structure of organelles, blocks membrane transport and activates the unfolded protein response (UPR). In addition, CPZ-treatment induces phosphorylation of the translation initiation factor (eIF2alpha), which reduces the general rate of protein synthesis and stimulates the production of Gcn4p, a major transcription factor that is activated in response to environmental stresses. Altogether, our results reveal that membrane stress within the cells rapidly activates an important gene expression program, which is followed by a general inhibition of protein synthesis. Remarkably, the increase of phosphorylated eIF2alpha and protein synthesis inhibition were also detected in CPZ-treated NIH-3T3 fibroblasts, suggesting the existence of a conserved mechanism of translational regulation that operates during a membrane stress.
氯丙嗪（CPZ）是一种可渗透的小分子阳离子两亲性分子，可插入膜双层并与阴离子脂质如多磷酸肌醇（PIs）结合。由于PIs在许多细胞过程中发挥重要作用，包括信号传导和膜运输途径，有人提出CPZ通过阻止具有特定PI结合结构域的蛋白质募集来影响细胞生长功能。在本研究中，我们研究了CPZ在酿酒酵母中的生物学效应。我们筛选了大约4800个基因敲除突变体，发现晚期高尔基体和内体区室之间膜运输缺陷的突变体对CPZ高度敏感。显微镜检查和运输分析表明，CPZ影响细胞器的膜结构，阻断膜运输并激活未折叠蛋白反应（UPR）。此外，CPZ处理诱导翻译起始因子（eIF2α）磷酸化，这降低了蛋白质合成的总体速率并刺激了Gcn4p的产生，Gcn4p是一种在应对环境压力时被激活的主要转录因子。总之，我们的结果表明细胞内的膜应激迅速激活了一个重要的基因表达程序，随后是蛋白质合成的普遍抑制。值得注意的是，在CPZ处理的NIH-3T3成纤维细胞中也检测到磷酸化eIF2α的增加和蛋白质合成抑制，这表明在膜应激期间存在一种保守的翻译调控机制。
