RESUMEN
Mitchell-Riley syndrome (MRS) is caused by recessive mutations in the regulatory factor X6 gene (RFX6) 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 RFX6 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 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 hybridisation 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 individuals with MRS specifically impairs formation of endocrine cells of the pancreas head and tail.
Asunto(s)
Diferenciación Celular , Diabetes Mellitus/genética , Diabetes Mellitus/patología , Endodermo/embriología , Enfermedades de la Vesícula Biliar/genética , Enfermedades de la Vesícula Biliar/patología , Células Madre Pluripotentes Inducidas/patología , Atresia Intestinal/genética , Atresia Intestinal/patología , Mutación/genética , Páncreas/embriología , Factores de Transcripción del Factor Regulador X/genética , Alelos , Secuencia de Bases , Diferenciación Celular/genética , Cromatina/metabolismo , Consanguinidad , Diabetes Mellitus/diagnóstico por imagen , Embrión de Mamíferos/metabolismo , Desarrollo Embrionario , Familia , Femenino , Enfermedades de la Vesícula Biliar/diagnóstico por imagen , Genoma Humano , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Atresia Intestinal/diagnóstico por imagen , Masculino , Linaje , Transcripción Genética , Transcriptoma/genética , Microtomografía por Rayos XRESUMEN
Fibroblast growth factors (FGFs) comprise a family of signalling molecules with essential roles in early embryonic development across animal species. The role of FGFs in mesoderm formation and patterning in Xenopus has been particularly well studied. However, little is known about FGF16 in Xenopus. Using in situ hybridisation, we uncover the expression pattern of FGF16 during early Xenopus laevis development, which has not been previously described. We show that the zygotic expression of FGF16 is activated in the mesoderm of the early gastrula as a ring around the blastopore, with its first accumulation at the dorsal side of the embryo. Later, FGF16 expression is found in the otic vesicle, the branchial arches and the anterior pituitary, as well as in the chordal neural hinge region of the tailbud. In addition, we show that FGF16 can activate the MAPK pathway and expression of sp5 and sp5l. Like FGF16, sp5 is expressed in the otic vesicle and the branchial arches, with all three of these genes being expressed in the tailbud. These data provide evidence that FGF16 is present in the early mesoderm and can activate the expression of developmentally important transcription factors.