Tominaga Takashi, Goto Motomitsu, Onoue Takeshi, Mizoguchi Akira, Sugiyama Mariko, Tsunekawa Taku, Hagiwara Daisuke, Morishita Yoshiaki, Ito Yoshihiro, Iwama Shintaro, Suga Hidetaka, Banno Ryoichi, Arima Hiroshi
Department of Endocrinology and Diabetes, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan.
Department of Endocrinology and Diabetes, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan.
Neurosci Lett. 2017 Aug 24;656:103-107. doi: 10.1016/j.neulet.2017.06.014. Epub 2017 Jun 13.
Sequestosome 1 (SQSTM1) also known as ubiquitin-binding protein p62 (p62) is a cargo protein involved in the degradation of misfolded proteins via selective autophagy. Disruption of autophagy and resulting accumulation of misfolded proteins in the endoplasmic reticulum (ER) leads to ER stress. ER stress is implicated in several neurodegenerative diseases and obesity. As knockout of p62 (p62KO) reportedly induces obesity in mice, we examined how p62 contributes to ER stress and the ensuing unfolded protein response (UPR) in hypothalamus using mouse organotypic cultures in the present study. Cultures from p62KO mice showed significantly reduced formation of LC3-GFP puncta, an index of autophagosome formation, in response to the chemical ER stressor thapsigargin compared to wild-type (WT) cultures. Hypothalamic cultures from p62KO mice exhibited higher basal expression of the UPR/ER stress markers CHOP mRNA and ATF4 mRNA than WT cultures. Thapsigargin enhanced CHOP, ATF4, and BiP mRNA as well as p-eIF2α protein expression in both WT and p62KO cultures, but all peak values were greater in p62KO cultures. A proteasome inhibitor increased p62 expression in WT cultures and upregulated the UPR/ER stress markers CHOP mRNA and ATF4 mRNA in both genotypes, but to a greater extent in p62KO cultures. Therefore, p62 deficiency disturbed autophagosome formation and enhanced both basal and chemically induced ER stress, suggesting that p62 serves to prevent ER stress in mouse hypothalamus by maintaining protein folding capacity.
聚集体蛋白1（SQSTM1），也被称为泛素结合蛋白p62，是一种货物蛋白，通过选择性自噬参与错误折叠蛋白的降解。自噬功能紊乱以及由此导致的错误折叠蛋白在内质网（ER）中的积累会引发内质网应激。内质网应激与多种神经退行性疾病和肥胖症有关。据报道，敲除p62（p62基因敲除）会诱导小鼠肥胖，因此在本研究中，我们使用小鼠器官型培养物研究了p62如何影响下丘脑的内质网应激以及随之而来的未折叠蛋白反应（UPR）。与野生型（WT）培养物相比，来自p62基因敲除小鼠的培养物在受到化学内质网应激诱导剂毒胡萝卜素刺激时，自噬体形成指标LC3 - GFP斑点的形成显著减少。来自p62基因敲除小鼠的下丘脑培养物中，UPR/内质网应激标志物CHOP mRNA和ATF4 mRNA的基础表达高于野生型培养物。毒胡萝卜素增强了野生型和p62基因敲除培养物中CHOP、ATF4和BiP mRNA以及p - eIF2α蛋白的表达，但p62基因敲除培养物中的所有峰值都更高。蛋白酶体抑制剂增加了野生型培养物中p62的表达，并上调了两种基因型中UPR/内质网应激标志物CHOP mRNA和ATF4 mRNA的表达，但在p62基因敲除培养物中的上调程度更大。因此，p62缺乏会干扰自噬体形成，并增强基础和化学诱导的内质网应激，这表明p62通过维持蛋白质折叠能力来预防小鼠下丘脑的内质网应激。
