Proteomic Analysis on Human Islets Suggests Nucleocytoplasmic Transport as a Mechanism of PERK Attenuation Effects in Diabetes

Partial downregulation of pancreatic endoplasmic reticulum kinase (PERK) activity recovered insulin content in human islets exposed to glucolipotoxicity (GLT), resulting in improved insulin secretion and glucose-lowering effects in a mouse model of type 2 diabetes. We conducted this study to elucidate the beta-cell-enhancing mechanisms of PERK attenuation. Pancreatic islets isolated from non-diabetic living donors were divided into three groups: control, GLT mimicking diabetes conditions, and GLT with treatment of a PERK inhibitor (PERKi, GSK2606414) for 24 h. Proteomic analysis was conducted on these samples. Differentially expressed proteins (DEPs) altered by GLT and reversed by PERKi were analyzed using bioinformatics. Validation studies were followed using western blotting, RT-PCR, and immunocytochemistry. Using nine islet samples pooled from seven participants, 161 DEPs were identified among 5513 quantifiable proteins across the three groups. On a subset of 42 proteins that were downregulated by GLT and upregulated by PERKi, GO, and KEGG analyses highlighted nucleocytoplasmic transport (NCT) as a key pathway, involving genes such as XPO4, KPNA4, NUP43, and NUP58. The involvement of NCT, particularly XPO4, was further supported by a replication proteomic analysis using islets from four independent donors. Based on these findings, we examined the NCT of representative β-cell transcription factors, including PDX1 and FOXO1. PERKi significantly increased their nuclear localization under GLT conditions (both p < 0.05), accompanied by heightened expression of their target genes, such as FBXW5. These results suggest that PERKi-mediated modulation of NCT may enhance the functional activity of PDX1 and FOXO1. In conclusion, proteomic analysis revealed that PERKi appears to modulate NCT of human islets under metabolic stress, thereby contributing to the restoration of β-cell function through regulation of relevant transcription factors. These findings suggested a novel mechanism of low-dose PERKi as a therapeutic approach to diabetes, in addition to the canonical unfolded protein response.