Mitchell-Riley syndrome iPSC exhibit reduced pancreatic endoderm differentiation due to an RFX6 mutation.

Mitchell-riley syndrome (MRS) is caused by recessive mutations in the Regulatory Factor X, 6 (RFX6) gene and is characterised by pancreatic hypoplasia and neonatal diabetes. To determine why MRS patients specifically lack pancreatic endocrine cells, we micro-CT imaged a 12-week old foetus homozygous for the nonsense mutation RFX6 c.1129C>T, which revealed loss of the pancreas body and tail. From this foetus, we derived iPSC and show that differentiation of these cells in vitro proceeds normally until generation of pancreatic endoderm, which is significantly reduced. We additionally generated an RFX6HA reporter allele by gene targeting in wild-type H9 cells to precisely define RFX6 expression and in parallel performed in situ hybridization for RFX6 in the dorsal pancreatic bud of a Carnegie Stage 14 human embryo. Both in vitro and in vivo, we find that RFX6 specifically labels a subset of PDX1-expressing pancreatic endoderm. In summary, RFX6 is essential for efficient differentiation of pancreatic endoderm, and its absence in MRS patients specifically impairs formation of endocrine cells of the pancreas head and tail.