Effects of a GTP-insensitive mutation of glutamate dehydrogenase on insulin secretion in transgenic mice.
Glutamate dehydrogenase (GDH) plays an important role in insulin secretion as evidenced in children by gain of function mutations of this enzyme that cause a hyperinsulinism-hyperammonemia syndrome (GDH-HI) and sensitize beta-cells to leucine stimulation. GDH transgenic mice were generated to express the human GDH-HI H454Y mutation and human wild-type GDH in islets driven by the rat insulin promoter. H454Y transgene expression was confirmed by increased GDH enzyme activity in islets and decreased sensitivity to GTP inhibition. The H454Y GDH transgenic mice had hypoglycemia with normal growth rates. H454Y GDH transgenic islets were more sensitive to leucine- and glutamine-stimulated insulin secretion but had decreased response to glucose stimulation. The fluxes via GDH and glutaminase were measured by tracing 15N flux from [2-15N]glutamine. The H454Y transgene in islets had higher insulin secretion in response to glutamine alone and had 2-fold greater GDH flux. High glucose inhibited both glutaminase and GDH flux, and leucine could not override this inhibition. 15NH4Cl tracing studies showed 15N was not incorporated into glutamate in either H454Y transgenic or normal islets. In conclusion, we generated a GDH-HI disease mouse model that has a hypoglycemia phenotype and confirmed that the mutation of H454Y is disease causing. Stimulation of insulin release by the H454Y GDH mutation or by leucine activation is associated with increased oxidative deamination of glutamate via GDH. This study suggests that GDH functions predominantly in the direction of glutamate oxidation rather than glutamate synthesis in mouse islets and that this flux is tightly controlled by glucose.
Division of Endocrinology, The Children's Hospital of Philadelphia and Diabetes Center, Philadelphia, Pennsylvania 19104, USA.
Changhong
C
Li
Andrea
A
Matter
Andrea
A
Kelly
Thomas J
TJ
Petty
Habiba
H
Najafi
Courtney
C
MacMullen
Yevgeny
Y
Daikhin
Ilana
I
Nissim
Adam
A
Lazarow
Jae
J
Kwagh
Heather W
HW
Collins
Betty Y L
BY
Hsu
Itzhak
I
Nissim
Marc
M
Yudkoff
Franz M
FM
Matschinsky
Charles A
CA
Stanley
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology; D005969:Glutamate Dehydrogenase / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D005973:Glutamine / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006801:Humans; D006946:Hyperinsulinism / Q000201:enzymology / Q000235:genetics / Q000503:physiopathology; D066298:In Vitro Techniques; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D007700:Kinetics; D007930:Leucine / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008954:Models, Biological; D009154:Mutation; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
D007328:Insulin; D011994:Recombinant Proteins; D005973:Glutamine; D000244:Adenosine Diphosphate; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D005969:Glutamate Dehydrogenase; D007930:Leucine; D005947:Glucose