Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark.
Division of Metabolism, Endocrinology, & Diabetes, University of Michigan Medical Center, Ann Arbor, MI.
Diabetes. 2019 Apr;68(4):747-760. doi: 10.2337/db18-0671. Epub 2019 Jan 22.
Although endoplasmic reticulum (ER) chaperone binding to mutant proinsulin has been reported, the role of protein chaperones in the handling of wild-type proinsulin is underinvestigated. Here, we have explored the importance of glucose-regulated protein 94 (GRP94), a prominent ER chaperone known to fold insulin-like growth factors, in proinsulin handling within β-cells. We found that GRP94 coimmunoprecipitated with proinsulin and that inhibition of GRP94 function and/or expression reduced glucose-dependent insulin secretion, shortened proinsulin half-life, and lowered intracellular proinsulin and insulin levels. This phenotype was accompanied by post-ER proinsulin misprocessing and higher numbers of enlarged insulin granules that contained amorphic material with reduced immunogold staining for mature insulin. Insulin granule exocytosis was accelerated twofold, but the secreted insulin had diminished bioactivity. Moreover, GRP94 knockdown or knockout in β-cells selectively activated protein kinase R-like endoplasmic reticulum kinase (PERK), without increasing apoptosis levels. Finally, GRP94 mRNA was overexpressed in islets from patients with type 2 diabetes. We conclude that GRP94 is a chaperone crucial for proinsulin handling and insulin secretion.
尽管内质网（ER）伴侣蛋白与突变前胰岛素原的结合已有报道，但对于野生型前胰岛素原的伴侣蛋白的作用研究甚少。在这里，我们研究了葡萄糖调节蛋白 94（GRP94）在β细胞中前胰岛素原处理中的重要性，它是一种著名的 ER 伴侣蛋白，已知能折叠胰岛素样生长因子。我们发现 GRP94 与前胰岛素原共沉淀，并且抑制 GRP94 的功能和/或表达会降低葡萄糖依赖性胰岛素分泌，缩短前胰岛素原半衰期，并降低细胞内前胰岛素原和胰岛素水平。这种表型伴随着 ER 后前胰岛素原错误处理，以及含有形态不规则物质的增大胰岛素颗粒数量增加，这些物质对成熟胰岛素的免疫金染色减少。胰岛素颗粒胞吐作用加快了两倍，但分泌的胰岛素生物活性降低。此外，β细胞中 GRP94 的敲低或敲除选择性激活蛋白激酶 R 样内质网激酶（PERK），而不会增加细胞凋亡水平。最后，GRP94mRNA 在 2 型糖尿病患者的胰岛中过度表达。我们得出结论，GRP94 是一种对前胰岛素原处理和胰岛素分泌至关重要的伴侣蛋白。
