Abeliovich H, Dunn W A, Kim J, Klionsky D J
University of Michigan, Department of Biology, Ann Arbor, Michigan 48109, USA.
J Cell Biol. 2000 Nov 27;151(5):1025-34. doi: 10.1083/jcb.151.5.1025.
Rapamycin, an antifungal macrolide antibiotic, mimics starvation conditions in Saccharomyces cerevisiae through activation of a general G(0) program that includes widespread effects on translation and transcription. Macroautophagy, a catabolic membrane trafficking phenomenon, is a prominent part of this response. Two views of the induction of autophagy may be considered. In one, up-regulation of proteins involved in autophagy causes its induction, implying that autophagy is the result of a signal transduction mechanism leading from Tor to the transcriptional and translational machinery. An alternative hypothesis postulates the existence of a dedicated signal transduction mechanism that induces autophagy directly. We tested these possibilities by assaying the effects of cycloheximide and specific mutations on the induction of autophagy. We find that induction of autophagy takes place in the absence of de novo protein synthesis, including that of specific autophagy-related proteins that are up-regulated in response to rapamycin. We also find that dephosphorylation of Apg13p, a signal transduction event that correlates with the onset of autophagy, is also independent of new protein synthesis. Finally, our data indicate that autophagosomes that form in the absence of protein synthesis are significantly smaller than normal, indicating a role for de novo protein synthesis in the regulation of autophagosome expansion. Our results define the existence of a signal transduction-dependent nucleation step and a separate autophagosome expansion step that together coordinate autophagosome biogenesis.
雷帕霉素是一种抗真菌大环内酯类抗生素，它通过激活一种通用的G(0)程序在酿酒酵母中模拟饥饿状态，该程序对翻译和转录具有广泛影响。巨自噬是一种分解代谢的膜运输现象，是这种反应的一个突出部分。关于自噬的诱导可以考虑两种观点。一种观点认为，参与自噬的蛋白质上调导致其诱导，这意味着自噬是从Tor到转录和翻译机制的信号转导机制的结果。另一种假设假定存在一种直接诱导自噬的专用信号转导机制。我们通过检测环己酰亚胺和特定突变对自噬诱导的影响来测试这些可能性。我们发现，在没有从头合成蛋白质的情况下，包括响应雷帕霉素而上调的特定自噬相关蛋白质的从头合成，自噬仍会发生。我们还发现，Apg13p的去磷酸化是一个与自噬开始相关的信号转导事件，它也独立于新的蛋白质合成。最后，我们的数据表明，在没有蛋白质合成的情况下形成的自噬体明显小于正常自噬体，这表明从头合成蛋白质在自噬体扩张的调节中起作用。我们的结果确定了存在一个信号转导依赖性的成核步骤和一个单独的自噬体扩张步骤，它们共同协调自噬体的生物发生。
