Nadanaka Satomi, Yoshida Hiderou, Sato Ryuichiro, Mori Kazutoshi
Department of Biophysics, Graduate School of Science, Kyoto University, Kitashirakawa-Oiwake, Sakyo-ku, Kyoto 606-8502, Japan.
Cell Struct Funct. 2006;31(2):109-16. doi: 10.1247/csf.06015. Epub 2006 Nov 17.
Mammalian transcription factor ATF6 is constitutively synthesized as a type II transmembrane protein embedded in the endoplasmic reticulum (ER). It is activated when unfolded proteins are accumulated in the ER under ER stress through a process called regulated intramembrane proteolysis (Rip), in which ATF6 is transported from the ER to the Golgi apparatus where it undergoes sequential cleavage by Site-1 and Site-2 proteases. The cytosolic transcription factor domain of ATF6 liberated from the Golgi membrane enters the nucleus where it activates transcription of ER-localized molecular chaperones and folding enzymes, leading to the maintenance of the homeostasis of the ER. Here, we analyzed M19 cells, a mutant of Chinese hamster ovary cells deficient in Site-2 protease. It was previously shown that M19 cells are defective in the induction of mRNA encoding the major ER chaperone BiP. In M19 cells, ATF6 was not converted from the membrane-bound precursor form to the cleaved and nuclear form as expected. Moreover, some of the ATF6 was constitutively relocated to the Golgi apparatus, where it was cleaved by Site-1 protease, and remained associated with the Golgi apparatus, indicating that the ER of M19 cells was constitutively stressed. Consistent with this notion, the two other ER stress response mediators, IRE1 and PERK, were also constitutively activated in M19 cells. M19 cells showed inefficient secretion of a model protein. These results suggest that Rip-mediated activation of ATF6 is important for the homeostasis of the ER in not only ER-stressed but also unstressed cells.
哺乳动物转录因子ATF6最初作为一种嵌入内质网（ER）的II型跨膜蛋白被组成性合成。当内质网应激时未折叠蛋白在内质网中积累，通过一种称为调节性膜内蛋白水解（Rip）的过程，ATF6被激活。在此过程中，ATF6从内质网转运至高尔基体，在那里它被位点1和位点2蛋白酶依次切割。从高尔基体膜释放的ATF6的胞质转录因子结构域进入细胞核，在那里它激活内质网定位的分子伴侣和折叠酶的转录，从而维持内质网的稳态。在这里，我们分析了M19细胞，这是一种缺乏位点2蛋白酶的中国仓鼠卵巢细胞突变体。先前已表明，M19细胞在诱导编码主要内质网伴侣BiP的mRNA方面存在缺陷。在M19细胞中，ATF6并未如预期那样从膜结合前体形式转化为切割后的核形式。此外，一些ATF6组成性地重新定位到高尔基体，在那里它被位点1蛋白酶切割，并与高尔基体保持关联，这表明M19细胞的内质网处于组成性应激状态。与此观点一致，另外两种内质网应激反应介质IRE1和PERK在M19细胞中也被组成性激活。M19细胞显示出模型蛋白分泌效率低下。这些结果表明，Rip介导的ATF6激活不仅对内质网应激细胞，而且对非应激细胞的内质网稳态都很重要。
