Insulin storage and glucose homeostasis in mice null for the granule zinc transporter ZnT8 and studies of the type 2 diabetes-associated variants.
Nicolson TJ., Bellomo EA., Wijesekara N., Loder MK., Baldwin JM., Gyulkhandanyan AV., Koshkin V., Tarasov AI., Carzaniga R., Kronenberger K., Taneja TK., da Silva Xavier G., Libert S., Froguel P., Scharfmann R., Stetsyuk V., Ravassard P., Parker H., Gribble FM., Reimann F., Sladek R., Hughes SJ., Johnson PR., Masseboeuf M., Burcelin R., Baldwin SA., Liu M., Lara-Lemus R., Arvan P., Schuit FC., Wheeler MB., Chimienti F., Rutter GA.
OBJECTIVE: Zinc ions are essential for the formation of hexameric insulin and hormone crystallization. A nonsynonymous single nucleotide polymorphism rs13266634 in the SLC30A8 gene, encoding the secretory granule zinc transporter ZnT8, is associated with type 2 diabetes. We describe the effects of deleting the ZnT8 gene in mice and explore the action of the at-risk allele. RESEARCH DESIGN AND METHODS: Slc30a8 null mice were generated and backcrossed at least twice onto a C57BL/6J background. Glucose and insulin tolerance were measured by intraperitoneal injection or euglycemic clamp, respectively. Insulin secretion, electrophysiology, imaging, and the generation of adenoviruses encoding the low- (W325) or elevated- (R325) risk ZnT8 alleles were undertaken using standard protocols. RESULTS: ZnT8(-/-) mice displayed age-, sex-, and diet-dependent abnormalities in glucose tolerance, insulin secretion, and body weight. Islets isolated from null mice had reduced granule zinc content and showed age-dependent changes in granule morphology, with markedly fewer dense cores but more rod-like crystals. Glucose-stimulated insulin secretion, granule fusion, and insulin crystal dissolution, assessed by total internal reflection fluorescence microscopy, were unchanged or enhanced in ZnT8(-/-) islets. Insulin processing was normal. Molecular modeling revealed that residue-325 was located at the interface between ZnT8 monomers. Correspondingly, the R325 variant displayed lower apparent Zn(2+) transport activity than W325 ZnT8 by fluorescence-based assay. CONCLUSIONS: ZnT8 is required for normal insulin crystallization and insulin release in vivo but not, remarkably, in vitro. Defects in the former processes in carriers of the R allele may increase type 2 diabetes risks.