The Transcription Factor Hepatocyte Nuclear Factor-6/Onecut-1 Controls the Expression of Its Paralog Onecut-3 in Developing Mouse Endoderm

During development, the endoderm gives rise to several organs, including the pancreas and liver. This differentiation process requires spatial and temporal regulation of gene expression in the endoderm by a network of tissue-specific transcription factors whose elucidation is far from complete. These factors include the Onecut protein hepatocyte nuclear factor-6 (HNF-6), which controls pancreas and liver development as shown in our previous work on Hnf6 knock-out embryos. In mammals, HNF-6 has two paralogs, Onecut-2 (OC-2) and OC-3, whose patterns of expression in the adult overlap with that of HNF-6. In the present work, we examine the expression profile of the three Onecut factors in the developing mouse endoderm. We show that HNF-6, OC-2, and OC-3 are expressed sequentially, which defines new steps in endoderm differentiation. By analyzing Hnf6 knock-out embryos we find that HNF-6 is required for expression of the Oc3 gene in the endoderm. We show that OC-3 colocalizes with HNF-6 in the endoderm and in embryonic pancreas and liver. Based on transfection, chromatin immunoprecipitation, and whole embryo electroporation experiments, we demonstrate that HNF-6 can bind to and stimulate the expression of the Oc3 gene. This study identifies a regulatory cascade between two paralogous transcription factors, sheds new light on the interpretation of the Hnf6 knock-out phenotype, and broadens the transcription factors network operating during development of the endoderm, liver, and pancreas.