Alterations in endoplasmic reticulum (ER) calcium (Ca2+) levels arising from reduced activity of the Sarcoendoplasmic Reticulum Ca2+ ATPase (SERCA) pump are associated with impaired insulin secretion and reduce β-cell survival in both major forms of diabetes. However, at present, the mechanisms of how reduced ER Ca2+ impairs β-cell function remain incompletely understood. To test this, we analyzed mice with β-cell specific SERCA2 deletion (βS2KO mice) and S2KO INS1 cells. βS2KO mice exhibited age-dependent glucose intolerance, reduced glucose-stimulated insulin secretion, and increased proinsulin secretion without evidence of impaired insulin sensitivity. Immunoblot analysis of βS2KO islets and S2KO INS-1 cells revealed reduced active forms of the proinsulin processing enzymes, PC1/3, PC2, and CPE, while immunofluorescence analysis suggested impaired ER-Golgi trafficking of the convertase enzymes and proinsulin. Because the ER serves as the main site of cellular lipid synthesis and changes in the β-cell lipidome have been linked with diabetes pathophysiology, we hypothesized that ER Ca2+ loss may alter protein trafficking in the secretory pathway through changes in ER lipid content. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was performed on the ER fractions of control and S2KO INS1 cells and revealed increased levels of 16:0, 24:0, 24:1, and total ceramides in S2KO cells. SERCA2 restoration partially rescued convertase enzyme expression, proinsulin processing, and ceramide content. Consistent with these findings, RNA sequencing analysis of βS2KO islets revealed changes in pathways related to vesicle transport, protein processing, and lipid metabolism. We conclude that SERCA2 in the β-cell plays a central role in the maintenance of insulin biosynthesis as well as ceramide metabolism.
T. Kono: None. H. Iida: None. C. Lee: None. P. Krishnan: None. P. Arvan: None. I. Lindberg: None. C. Evans-molina: Advisory Panel; Self; Provention Bio, Inc., Consultant; Self; Dompe, Other Relationship; Self; Bristol-Myers Squibb Company, Nimbus Pharmaceuticals, Pfizer Inc.
National Institutes of Health (R01DK093954)