Impaired proinsulin processing is observed in both type 1 and type 2 diabetes. We have previously shown that reductions in endoplasmic reticulum (ER) calcium (Ca2+) in the pancreatic β cell arising from impaired activity of the Sarcoendoplasmic Reticulum Ca2+ ATPase (SERCA) pump are associated with increased proinsulin secretion. However, the mechanisms responsible for reduced proinsulin processing in the context of SERCA2 deficiency remain incompletely understood. To test this, we developed mice with β cell specific SERCA2 deletion (βS2KO mice) and S2KO INS1 cells. βS2KO mice exhibited age-dependent glucose intolerance and reduced glucose-stimulated insulin secretion without evidence of impaired insulin sensitivity. ER Ca2+ levels in islets from βS2KO mice were significantly reduced, while serum proinsulin/insulin (PI/I) ratios and whole pancreas PI/I content were elevated. Immunoblot analysis of βS2KO islets and S2KO INS-1 cells revealed reduced active forms of the proinsulin processing enzymes, PC1/3, PC2 and CPE. Restoration of SERCA2b via adenoviral transduction in S2KO INS1 cells was sufficient to restore PC1/3 and PC2 maturation and enzyme activity. Brefeldin A treatment in INS1 cells recapitulated the impairments in PC1/3 and PC2 maturation observed in S2KO cells, suggesting a disturbance in protein trafficking between the ER and Golgi. Consistent with this, trafficking assays were performed using a vesicular stomatitis virus G (VSVG) protein construct and revealed a significantly slower rate of VSVG movement from the ER to the Golgi in S2KO INS1 cells. Moreover, pancreas sections from βS2KO mice showed increased co-localization of proinsulin and ProPC2 in the early compartments of the secretory pathway. Taken together, these data suggest that loss of SERCA2 activity and ER Ca2+ loss in the pancreatic β cell leads to impaired proinsulin processing via reduced maturation and trafficking of proinsulin processing enzymes.
T. Kono: None. H. Iida: None. T.S. Jarvela: None. C. Lee: None. X. tong: None. P. Arvan: None. I. Lindberg: None. C. Evans-Molina: Consultant; Self; Bristol-Myers Squibb.
National Institutes of Health (R01DK093954); U.S. Department of Veterans Affairs (2-I01BX001733)