申请人及所在课题组的前期研究发现Nix蛋白介导了线粒体自噬在脑缺血/复灌中的神经保护作用，而且适度的内质网应激可以通过增强线粒体自噬最终发挥抗缺血性脑损伤作用，但其机制尚不明确。申请人利用TAP结合蛋白质谱技术，发现并鉴定了一种与内质网相关的Nix结合蛋白Calreticulin（CRT）。预实验已初步发现沉默CRT加重缺血/复灌诱导的细胞损伤，而共转染CRT和Nix则可以进一步促进线粒体自噬，提示CRT可能是脑缺血过程中内质网应激诱导线粒体自噬发挥神经保护作用的关键，且其作用机制可能与Nix的相互作用有关。因此，本研究拟采用体内、外脑缺血模型，利用药理学、RNA干扰、质粒过表达等手段，结合Nix基因敲除小鼠，从多个层面明确CRT在缺血性脑损伤中的作用，并阐明其对Nix介导的线粒体自噬的调控机制。本项目的研究结果将为寻找与激活内质网应激相关药物靶点治疗缺血性脑损伤提供重要的线索与实验依据。

Our previous research demonstrated that mitophagy protects against ischemia/reperfusion (I/R)-induced brain injury, which is regulated by Nix protein. Furthermore, moderate endoplasmic reticulum (ER) stress could protect against ischemic brain injury by enhancing mitophagy in mice, however, the underlying mechanisms is still unclear. By tandem affinity purification assay we found a novel ER-related Nix-binding protein, Calreticulin (CRT). Our preliminary data showed that knockdown of CRT significantly increased I/R-induced cell demise and co-transfection of CRT and Nix can promote mitophagy. This evidence suggests that CRT is a critical component of ER stress-induced mitophagy during brain I/R through the interaction with Nix. So we aimed to investigate the neuroprotective effect of CRT in the context of brain I/R and intend to explore the underlying mechanisms related to the Nix-mediated mitophagy. These investigations will provide significant experimental evidence in taking ER stress as novel therapeutic target in cerebral ischemia.