Deficiencies in pancreatic ß-cell Ca2+ ATPase SPCA1 alter Golgi Ca2+ Homeostasis
Parimanath, Elizabeth   (Indiana University - Indianapolis)
Mentor: Molina, Carmella
Within the ß-cell, Ca2+ is an important cofactor for a number of processes. Intracellular Ca2+ is dynamically stored within secretory pathway organelles at higher concentrations than the cytosol. While altered ER Ca2+ levels have been found to contribute to beta cell dyshomeostasis, the contribution of Golgi Ca2+ to diabetes pathophysiology is unknown. Intra- Golgi stores of Ca2+ are maintained by the Secretory Pathway Calcium ATPase (SPCA1). SPCA1 expression was measured in islets from cadaveric human donors with T1D and T2D, and SPCA1 levels were significantly lower compared to non-diabetic controls. We hypothesized that loss of SPCA1 in the ß-cell would lead to altered Golgi Ca2+ dyshomeostasis, impaired insulin secretion, and increased Golgi stress. Utilizing Golgi subcompartment specific probes and fluorescence lifetime imaging microscopy, we found INS-1 cells lacking SPCA1 (SPCA1KO) cells exhibit reduced intra-Golgi Ca2+ levels compared to WT INS-1 cells and restoration of SPCA1 normalized intra-Golgi Ca2+. Immunofluorescent staining for the Golgi markers Syntaxin6 and Giantin revealed no difference in Golgi volume between WT and SPCA1KO cells. To assess the effect of SPCA1 in primary tissue, islets were isolated from SPCA1 haploinsufficient (SPCA1+/-) and WT littermate C57BL/6J mice and cytosolic Ca2+ was labeled with Fura2. Islets from 8wk old male SPCA1+/- mice exhibited altered Ca2+ oscillations, and dysfunction was exacerbated in islets from 24wk old male SPCA1+/- mice following 16 weeks high fat diet. Collectively, our data suggests that SPCA1 loss dysregulates Golgi Ca2+ homeostasis impacting proper ß-cell function.