Abstract
How cells integrate multiple patterning signals to achieve early endoderm regionalization remains largely unknown. Between gastrulation and neurulation, retinoic acid (RA) signaling is required, while Wnt/β-catenin signaling has to be repressed for the specification of the pancreas, oesophagus, stomach, and duodenum primordia in Xenopus embryos. In attempt to screen for RA regulated genes in Xenopus endoderm, we identified a direct RA target gene, N-myc downstream regulated gene 1a (ndrg1a) that showed expression early in the archenteron roof endoderm and late in the developing pancreas, oesophagus, stomach, and duodenum. Both antisense morpholino oligonucleotide mediated knockdown of ndrg1a in Xenopus laevis and the transcription activator-like effector nucleases (TALEN) mediated disruption of ndrg1 in Xenopus tropicalis demonstrate that like RA signaling, Ndrg1a is specifically required for the specification of Xenopus pancreas, oesophagus, stomach, and duodenum primordia. Immunofluorescence data suggest that RA-activated Ndrg1a suppresses Wnt/β-catenin signaling in Xenopus archenteron roof endoderm cells. Blocking Wnt/β-catenin signaling rescued Ndrg1a knockdown phenotype. Furthermore, overexpression of the putative Wnt/β-catenin target gene Atf3 phenocopied knockdown of Ndrg1a or inhibition of RA signaling, while Atf3 knockdown can rescue Ndrg1a knockdown phenotype. Lastly, the pancreas/stomach/duodenum transcription factor Pdx1 was able to rescue Atf3 overexpression or Ndrg1a knockdown phenotype. Together, we conclude that RA activated Ndrg1a represses Wnt/β-catenin signaling to allow the specification of pancreas, oesophagus, stomach, and duodenum progenitor cells in Xenopus embryos.
Highlights
The regionalization of endoderm occurs concurrently with its formation during gastrulation
We provide evidences implicating that retinoic acid (RA)-activated Ndrg1a represses Wnt/b-catenin signaling in the archenteron roof endoderm cells and releases the inhibitory effect of Wnt/b-catenin signaling activities on the formation of pancreas, oesophagus, stomach, and duodenum
Endoderm significantly increased upon BMS453 treatment, or MO1 injection (Fig. 6A, B). These data suggest that RA and Ndrg1a repress Wnt/b-catenin signaling in Xenopus archenteron roof endoderm cells
Summary
The regionalization of endoderm occurs concurrently with its formation during gastrulation. Avian, and mice indicate that the RA signaling is required for the formation of dorsal pancreas, oesophagus, stomach, and duodenum primordia [10,11,12,13,14,15]. The endodermal RA target genes that mediate the early activities of RA signaling to specify the pancreas, oesophagus, stomach, and duodenum anlagen proper remain to be identified. In Xenopus, Wnt/b-catenin signaling must be repressed in anterior endoderm between gastrula and early somite stages of development to allow the formation of the liver as well as the pancreas, stomach, and duodenum primordia [22,23]. We provide evidences implicating that RA-activated Ndrg1a represses Wnt/b-catenin signaling in the archenteron roof endoderm cells and releases the inhibitory effect of Wnt/b-catenin signaling activities on the formation of pancreas, oesophagus, stomach, and duodenum
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