Abstract：
The Unfolded protein response (UPR) is an adaptive signalling pathway which is triggered by accumulation of unfolded/misfolded protein in ER lumen. The UPR consist of three transmembrane proteins-IRE1α, PERK and ATF6 that sense ER stress which leads to activation and downstream signaling from ER lumen to cytosol to restore homeostasis. IRE1α is an evolutionary conserved arm of UPR and acts as an interaction platform for many potential proteins that become activated under ER stress conditions. We investigated potential partners of IRE1 α through MS studies and found EXOSC3 as one of the binding partner of IRE1α. Exosomal complex proteins have 3\'5\' exonuclease properties (EXOSC3) that play an important role in mRNA surveillance. This property of exosomal proteins coincides with IRE1α ribonuclease activities and its mechanism of action is similar to that of IRE1α-RIDD pathway which degrades any unstable mRNA that disrupts cellular homeostasis. At the same time, studies have shown that knockdown of EXOSC3 causes ER stress in human cells, so we speculated that there might be a functional crosstalk between IRE1α and EXOSC3 under ER stress conditions. Therefore, we employed computational tools to predict and explore the stability and dynamics of the IRE1α-EXOSC3 complex. The analysis indicates that IRE1α and EXOSC3 exhibit potential interaction with the involvement of ScanNet, predicting binding pockets between the two proteins. Further, the interaction was validated via co-immunoprecipitation and yeast two-hybrid assays, thus suggesting EXOSC3 as a component of the UPRosome complex. Hence, this functional crosstalk might influence the dynamic functional output of IRE1α.Communicated by Ramaswamy H. Sarma.
摘要：
未折叠蛋白反应(UPR)是适应性信号传导途径,其由未折叠/错误折叠蛋白在ER腔中的积累触发。UPR由三种跨膜蛋白IRE1α组成，PERK和ATF6感测导致激活的ER应激和从ER腔到细胞质的下游信号以恢复体内平衡。IRE1α是UPR的进化保守臂，并充当许多在ER应激条件下被激活的潜在蛋白质的相互作用平台。我们通过MS研究调查了IRE1α的潜在伴侣，发现EXOSC3是IRE1α的结合伴侣之一。外泌体复合蛋白具有3'5'核酸外切酶特性（EXOSC3），在mRNA监测中起重要作用。外泌体蛋白的这种特性与IRE1α核糖核酸酶活性一致，其作用机制类似于IRE1α-RIDD途径，该途径降解任何破坏细胞稳态的不稳定mRNA。同时,研究表明，敲除EXOSC3会导致人体细胞内质网应激，因此，我们推测在ER应力条件下，IRE1α和EXOSC3之间可能存在功能性串扰。因此,我们使用计算工具来预测和探索IRE1α-EXOSC3复合物的稳定性和动力学。分析表明，IRE1α和EXOSC3与ScanNet的参与表现出潜在的相互作用，预测两种蛋白质之间的结合口袋。Further,通过免疫共沉淀和酵母双杂交试验验证了相互作用，因此表明EXOSC3是UPFrosome复合体的组成部分。因此，这种功能串扰可能会影响IRE1α的动态功能输出。由RamaswamyH.Sarma沟通。
