Journal
DIABETES
Volume 59, Issue 7, Pages 1674-1685Publisher
AMER DIABETES ASSOC
DOI: 10.2337/db09-0986
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Funding
- Swiss National Science Foundation
- Geneva Cancer League
- Swiss Multiple Sclerosis Society
- National Center of Competence in Research on Neural Plasticity and Repair (NCCR-NEURO)
- Boninchi Foundation
- Fondation Romande pour la Recherche sur le Diabete
- Roche Research Foundation
- Jules Thorne Foundation
- Association de Langue Francaise pour l'Etude du Diabete et des Maladies Metaboliques (ALFEDIAM)
- European Foundation for the Study of Diabetes (EFSD)
- Swiss National Science Foundation [310000-109402, CR3213129987, 3100A0-103867/1]
- Juvenile Diabetes Research Foundation [12007-158]
- EU [222980, C2008-T7]
- NIH/NIDDK [DK072522-01]
- NCCR Frontiers in Genetics
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OBJECTIVE-Pancreatic islets of perinatal mice lacking the transcription factor Rfx3 exhibit a marked reduction in insulin-producing beta-cells. The objective of this work was to unravel the cellular and molecular mechanisms underlying this deficiency. RESEARCH DESIGN AND METHODS-Immunofluorescence studies and quantitative RT-PCR experiments were used to study the emergence of insulin-positive cells, the expression of transcription factors implicated in the differentiation of beta-cells from endocrine progenitors, and the expression of mature beta-cell markers during development in Rfx3(-/-) and pancreas-specific Rfx3-knockout mice. RNA interference experiments were performed to document the consequences of downregulating Rfx3 expression in Min6 beta-cells. Quantitative chromatin immunoprecipitation (ChIP), ChIP sequencing, and bandshift experiments were used to identify Rfx3 target genes. RESULTS-Reduced development of insulin-positive cells in Rfx3(-/-) mice was not due to deficiencies in endocrine progenitors or beta-lineage specification, but reflected the accumulation of insulin-positive beta-cell precursors and defective beta-cells exhibiting reduced insulin, Glut-2, and Gck expression. Similar incompletely differentiated beta-cells developed in pancreas-specific Rfx3-deficient embryos. Defective beta-cells lacking Glut-2 and Gck expression dominate in Rfx3-deficent adults, leading to glucose intolerance. Attenuated Glut-2 and glucokinase expression, and impaired glucose-stimulated insulin secretion, were also induced by RNA interference-mediated inhibition of Rfx3 expression in Min6 cells. Finally, Rfx3 was found to bind in Min6 cells and human islets to two well-known regulatory sequences, Pal-1 and Pal-2, in the neuroendocrine promoter of the glucokinase gene. CONCLUSIONS-Our results show that Rfx3 is required for the differentiation and function of mature beta-cells and regulates the beta-cell promoter of the glucokinase gene. Diabetes 59:16741685, 2010
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