Mitchell-Riley syndrome iPSC exhibit reduced pancreatic endoderm differentiation due to an <i>RFX6</i> mutation

Abstract

Mitchell-Riley syndrome (MRS) is caused by recessive mutations in the regulatory factor X6 gene (<i>RFX6</i>) and is characterised by pancreatic hypoplasia and neonatal diabetes. To determine why individuals with MRS specifically lack pancreatic endocrine cells, we micro-CT imaged a 12-week-old foetus homozygous for the nonsense mutation <i>RFX6</i> c.1129C>T, which revealed loss of the pancreas body and tail. From this foetus, we derived iPSCs and show that differentiation of these cells <i>in vitro</i> proceeds normally until generation of pancreatic endoderm, which is significantly reduced. We additionally generated an <i>RFX6^HA</i> reporter allele by gene targeting in wild-type H9 cells to precisely define RFX6 expression and in parallel performed <i>in situ</i> hybridisation for <i>RFX6</i> in the dorsal pancreatic bud of a Carnegie stage 14 human embryo. Both <i>in vitro</i> and <i>in vivo</i>, we find that RFX6 specifically labels a subset of PDX1-expressing pancreatic endoderm. In summary, <i>RFX6</i> is essential for efficient differentiation of pancreatic endoderm, and its absence in individuals with MRS specifically impairs formation of endocrine cells of the pancreas head and tail.