内质网定位的TIGAR调节GRP78在脑预适应中的作用和机制

Our preliminary data showed that TP53-induced glycolysis and apoptosis regulator (TIGAR) relocates to the endoplasmic reticulum (ER) during cerebral preconditioning, but the biological significance of TIGAR’s relocation in ER remains to be elucidated. We hypothesize that during cerebral preconditioning, TIGAR relocates to the ER, where it regulates pentose phosphate pathway (PPP) to remove reactive oxygen species (ROS) and to maintain cell survival. Alternatively, TIGAR may regulate the ER molecular chaperone GRP78 to relieve oxidative stress in ER and ER stress dependent apoptosis. Another possibility is that TIGAR may act on GRP78 to regulate ER-phagy through STX17/Rab1/FAM134B pathway. In this project, we will establish isoflurane, hypoxic and ischemic preconditioning models in mice, primary murine cortical neurons and HT22 hipocampal neuronal cells to characterize the neuroprotective role of ER resident TIGAR and its regulation on GRP78 in cerebral preconditioning and the mechanisms. First, we intend to demonstrate that TIGAR translocates to ER to mediate ischemic tolerance during cerebral preconditioning. This study will extend our knowledge of the organelle distribution and bioactivity of TIGAR. Second, we will test whether TIGAR regulates ER stress, ROS production in ER and ER-phagy through regulation on GRP78 to broaden the bioactivity and signals of ER-phagy. Finally, we want to characterize the molecular mechanism of TIGAR’s ER relocation and its regulation on GRP78. We will use agents acting on TIGAR or GRP78 to mimic preconditioning-induced neuroprotection, thus proposing a therapeutic strategy for ischemic cerebrovascular diseases.

我们的预实验发现脑预适应时TIGAR(TP53诱导糖酵解和细胞凋亡调节子)上调并转位于内质网（ER），但生物意义不明。我们推测在脑预适应时TIGAR转位于ER，促进ER内磷酸戊糖途径，或调节ER分子伴侣GRP78，减轻ER内氧化应激和ER应激依赖的细胞凋亡，或经STX17/Rab1/FAM134B调节内质网自噬，介导缺血耐受。本课题建立整体动物和离体神经细胞预适应模型，应用多种分子生物学方法研究脑预适应中TIGAR分布于ER并调节GRP78产生神经保护作用的机制，旨在：① 研究脑预适应时TIGAR转位于ER介导缺血耐受，扩展TIGAR细胞器分布和生物功能。② 深入研究TIGAR是否通过GRP78调节ER应激，ER内ROS生成和内质网自噬，丰富内质网自噬的生物活性与调节信号。③ 探索神经细胞内TIGAR转位ER并调节GRP78的分子机制，为防治脑缺血提供靶向TIGAR或GRP78的候选药物。

脑预适应或脑缺血再灌时TIGAR(TP53诱导糖酵解和细胞凋亡调节子)上调并转位于内质网（ER），但生物意义不明。我们推测在脑预适应或脑缺血再灌时TIGAR上调，转位于内质网（ER），减轻ER应激或氧化应激，或TIGAR经STX17调节自噬，介导神经保护作用。本课题从这些假设入手，建立小鼠和培养神经细胞预适应和缺血再灌注模型，在整体和细胞水平应用多种分子生物学方法研究了在脑预适应和缺血再灌注中，TIGAR产生神经保护作用的机制，研究发现：①.证实TIGAR参与脑预适应介导的缺血耐受；②.发现脑缺血再灌注中TIGAR转位于内质网（ER）及细胞核，抑制ER应激，以产生神经保护作用的新机制，扩展TIGAR的细胞器分布和生物功能；③.发现TIGAR调节脑缺血再灌注中自噬活性产生神经保护作用，而STX17通过增强自噬流和减轻ER应激依赖性神经元凋亡来抑制缺血性损伤，为防治缺血性脑中风和其他神经退行性疾病提供了新靶点或新思路。