Endoplasmic reticulum (ER) stress is emerging as a factor for the pathogenesis of granular corneal dystrophy type 2 (GCD2). This study was designed to investigate the molecular mechanisms underlying the protective effects of melatonin on ER stress in GCD2. Our results showed that GCD2 corneal fibroblasts were more susceptible to ER stress‐induced death than were wild‐type cells. Melatonin significantly inhibited GCD2 corneal cell death, caspase‐3 activation, and poly (ADP‐ribose) polymerase 1 cleavage caused by the ER stress inducer, tunicamycin. Under ER stress, melatonin significantly suppressed the induction of immunoglobulin heavy‐chain‐binding protein (BiP) and activation of inositol‐requiring enzyme 1α (IRE1α), and their downstream target, alternative splicing of X‐box binding protein 1(XBP1). Notably, the reduction in BiP and IRE1α by melatonin was suppressed by the ubiquitin‐proteasome inhibitor, MG132, but not by the autophagy inhibitor, bafilomycin A1, indicating involvement of the ER‐associated protein degradation (ERAD) system. Melatonin treatment reduced the levels of transforming growth factor‐β‐induced protein (TGFBIp) significantly, and this reduction was suppressed by MG132. We also found reduced mRNA expression of the ERAD system components HRD1 and SEL1L, and a reduced level of SEL1L protein in GCD2 cells. Interestingly, melatonin treatments enhanced SEL1L levels and suppressed the inhibition of SEL1L N‐glycosylation caused by tunicamycin. In conclusion, this study provides new insights into the mechanisms by which melatonin confers its protective actions during ER stress. The results also indicate that melatonin might have potential as a therapeutic agent for ER stress‐related diseases including GCD2.

中文翻译：

---

褪黑素通过激活内质网相关蛋白降解来减少内质网应激和角膜营养不良相关的TGFBIp

内质网（ER）压力正在成为2型颗粒性角膜营养不良（GCD2）发病机理的一个因素。本研究旨在研究褪黑激素对GCD2内质网应激的保护作用的分子机制。我们的结果表明，与野生型细胞相比，GCD2角膜成纤维细胞更容易受到内质网应激诱导的死亡。褪黑素显着抑制了由ER应激诱导剂衣霉素引起的GCD2角膜细胞死亡，caspase-3活化和多聚（ADP-核糖）聚合酶1裂解。在内质网应激下，褪黑素显着抑制免疫球蛋白重链结合蛋白（BiP）的诱导和肌醇需要酶1α（IRE1α）及其下游靶标的激活，X盒结合蛋白1（XBP1）的选择性剪接。尤其，泛素-蛋白酶体抑制剂MG132抑制了褪黑素对BiP和IRE1α的降低，而自噬抑制剂巴非霉素A1则没有抑制，表明ER相关蛋白降解（ERAD）系统的参与。褪黑素治疗可显着降低转化生长因子-β诱导蛋白（TGFBIp）的水平，而MG132可抑制这种降低。我们还发现ERAD系统组件的mRNA表达降低HRD1和SEL1L以及GCD2细胞中SEL1L蛋白的水平降低。有趣的是，褪黑激素治疗可增强衣霉素引起的SEL1L水平并抑制SEL1L N糖基化。总之，本研究为褪黑激素在内质网应激过程中赋予保护作用的机制提供了新的见解。结果还表明，褪黑激素可能具有作为治疗与ER应激相关疾病（包括GCD2）的作用的潜力。