Insulin storage and glucose homeostasis in mice null for the granule zinc transporter ZnT8 and studies of the type 2 diabetes-associated variants.
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.


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.
Nicolson
Tamara J
TJ
Section of Cell Biology, Division of Medicine, Imperial College London, London, U.K.
Bellomo
Elisa A
EA
Wijesekara
Nadeeja
N
Loder
Merewyn K
MK
Baldwin
Jocelyn M
JM
Gyulkhandanyan
Armen V
AV
Koshkin
Vasilij
V
Tarasov
Andrei I
AI
Carzaniga
Raffaella
R
Kronenberger
Katrin
K
Taneja
Tarvinder K
TK
da Silva Xavier
Gabriela
G
Libert
Sarah
S
Froguel
Philippe
P
Scharfmann
Raphael
R
Stetsyuk
Volodymir
V
Ravassard
Philippe
P
Parker
Helen
H
Gribble
Fiona M
FM
Reimann
Frank
F
Sladek
Robert
R
Hughes
Stephen J
SJ
Johnson
Paul R V
PR
Masseboeuf
Myriam
M
Burcelin
Remy
R
Baldwin
Stephen A
SA
Liu
Ming
M
Lara-Lemus
Roberto
R
Arvan
Peter
P
Schuit
Frans C
FC
Wheeler
Michael B
MB
Chimienti
Fabrice
F
Rutter
Guy A
GA
D000818:Animals; D001786:Blood Glucose / Q000378:metabolism*; D027682:Cation Transport Proteins / Q000235:genetics / Q000378:metabolism*; D003594:Cytoplasmic Granules / Q000378:metabolism; D003924:Diabetes Mellitus, Type 2 / Q000453:epidemiology / Q000235:genetics / Q000378:metabolism*; D005089:Exocytosis / Q000502:physiology; D005260:Female; D015870:Gene Expression / Q000502:physiology; D006367:HeLa Cells; D006706:Homeostasis / Q000502:physiology; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000378:metabolism / Q000473:pathology / Q000502:physiology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D011110:Polymorphism, Genetic; D012307:Risk Factors; D015032:Zinc / Q000378:metabolism*; D000075703:Zinc Transporter 8
D001786:Blood Glucose; D027682:Cation Transport Proteins; D007328:Insulin; C492522:SLC30A8 protein, human; C516348:Slc30a8 protein, mouse; D000075703:Zinc Transporter 8; D015032:Zinc