Human regulates endoderm patterning at the primitive gut tube stage.

Transcriptional factor is known to be a causal gene of Mitchell-Riley syndrome (MRS), an autosomal recessive neonatal diabetes associated with pancreatic hypoplasia and intestinal atresia/malformation. The morphological defects are limited to posterior foregut and mid-hindgut endodermal lineages and do not occur in the anterior foregut lineage; the mechanism remains to be fully elucidated. In this study, we generated heterozygous knockin and homozygous knockin/knockout human-induced pluripotent stem cell (hiPSC) lines and performed in vitro endoderm differentiation to clarify the role of RFX6 in early endoderm development. expression was found to surge at the primitive gut tube (PGT) stage in comparison with that in the undifferentiated or definitive endoderm stage. At the PGT stage, the expression of and , posterior foregut and mid-hindgut master regulators, respectively, was decreased by the RFX6 deficit. PDX1 and CDX2 cells were mostly green fluorescent protein (GFP) in hiPSCs, but their cell number was markedly decreased in hiPSCs. The expression of , an anterior foregut marker, was not affected by the RFX6 deficit. In addition, we found a putative RFX6-binding X-box motif using cap analysis of gene expression-seq and the motif-containing sequences in the enhancer regions of and bound to RFX6 in vitro. Thus, regulates the ParaHox genes and but does not affect in early endodermal differentiation, suggesting that defects in early stage endoderm patterning account for the morphological pathology of MRS.