Rfx6 in pancreatic and intestinal endocrine cell differentiation and function – RFX-PancInt

Recently the transcription factor RFX6 has been identified as a novel causal gene in monogenic diabetes. Indeed, loss of function mutations in RFX6 transcription factor have been reported to be responsible for a rare autosomal recessive disease syndrome of neonatal diabetes and small bowel obstruction associated with severe intestinal malabsorption and diarrhea (Smith et al, Nature 2010).

We initially identified RFX6 as a downstream target of the proendocrine transcription factor Neurog3 when studying the molecular and cellular mechanisms controlling islet cell development in the mouse pancreas (Soyer et al Development 2010). In the adult, RFX6 is expressed in islet cells as well as in endocrine cells of the intestine such as L and K cells which secrete the gluco-incretin hormones GLP1 and GIP (unpublished data). Rfx6-deficient mice fail to develop hormone expressing alpha-, beta- and delta-cells, and die post-partum (Smith et al, Nature 2010). Thus Rfx6 is an essential gene for islet cell development including for the production of insulin in beta cell. Because of the early lethality of the constitutive knock-out mouse, the role of Rfx6 in adult beta cell and enteroendocrine cells including in insulin secretion and the regulation of glucose homeostasis could not be studied. Similarly the mechanisms for beta cell defect in human patients are unknown.

For these reasons the main goals of the RFX-PancInt research proposal are to decipher - the role of Rfx6 in the control of glucose homeostasis in mouse and human - the mechanisms underlying the pathophysiology of diabetes and intestinal failure in relation with RFX6 mutations.
To reach these goals three research teams, with complementary expertise in -mouse models to study islet and endocrine cell differentiation and function, -ex vivo system to study human beta cells and -human genetics in diabetes will join their efforts: The Gradwohl lab (coordinator, IGBMC, Illkirch) the Scharfmann lab (Inserm, Paris) and the Vaxillaire/Froguel lab (CNRS, Institut Pasteur, Lille).

We will use loss of function and functional genomics approaches, in mouse and in human beta cell lines, to address the role of RFX6 in beta cell differentiation and function. We have generated a conditional knock-out mouse to specifically inactive Rfx6 in the developing and adult beta cells. Consequences on beta cell differentiation, maintenance and regulation of glucose homeostasis will be studied. We will dissect RFX6 role using a unique model of functional human beta cell lines. In this system, we will knock-down RFX6 and assess insulin content and secretion. To identify RFX6 direct targets we will perform ChIPSeq and microarrays experiments in the mouse and human experimental systems. By specifically inactivating Rfx6 in the adult mouse intestine we will address the role of this gene in the differentiation and function of the enteroendocrine lineage and explore particularly intestinal absorption, glucose and energy metabolism. Finally, we have identified a novel RFX6 mutation in a patient with adult onset of diabetes. Our goals are to assess the clinical features associated with the mutation to define a causal link with diabetes and to screen for other Rfx6 mutation(s) in additional diabetic patients. We will also determine the molecular characteristics and functional defects of the mutant RFX6 protein. Finally we also aim to sequence novel candidate genes in the Neurog3 and RFX6-pathways for monogenic diabetes in human.

The strengths of this proposal are – the relevance of the study to understand the mechanisms of beta cell function and glucose homeostasis and thus failure in diabetes – the use of novel mouse and human experimental models – the study of a novel mutation in human – and the complementary expertise of the investigators. With this study we expect to further dissect the determinants of normal glycemia, which may have significant implications for the discovery of novel therapeutic targets.