DEV001230 Supplementary Material

Files in this Data Supplement:

• Supplemental Table S1 - Adobe PDF

• Supplemental Figure S1 -

  Fig. S1. β-catenin activity represses expression of foregut genes hhex and foxa2. (A,B) At the 32-cell stage, embryos were injected either (A) in the D1 cell with stabilized pt-β-catenin RNA (250 pg) or (B) in the D4 cell with Gsk3β (500 pg) to repress Wnt signaling. At gastrula stage, either anterior endoderm (AE) or posterior endoderm (PE) was isolated and assayed after only 4 hours by RT-PCR for expression of hhex, foxa2 and the pan-endodermal marker sox17α. Bar charts showing normalized relative mRNA expression levels. (C-E) In situ hybridization of bisected stage-18 embryos with a foxa2 probe. Xenopus foxa2 is expressed throughout the endoderm but is enriched in the foregut. (D) Injection of stabilized pt-β-catenin RNA (250 pg) in D1 at the 32-cell stage results in partial repression of foxa2 in the foregut, whereas (E) injection of Gsk3β (500 pg) in D4 upregulates foxa2 expression in the posterior endoderm.

• Supplemental Figure S2 -

  Fig. S2. Vent2 is expressed in the posterior endoderm and can repress foregut development from blastula to early somite stages. (A) In situ hybridization of bisected Xenopus embryos (anterior left) shows that vent2 RNA is robustly expressed in the posterior endoderm from gastrula stage 11 to stage 22, in a reciprocal pattern to hhex. Throughout this period of development, wnt8 RNA is expressed in the lateral mesoderm (red arrows) adjacent to the vent2-expressing posterior endoderm. (B) Injection of vent2 RNA (500 pg) into the anterior D1 cells of the 32-cell stage embryo results in repression of foxa2 expression in the foregut endoderm at stage 18. (C) RNA encoding a hormone inducible GR-Vent2 fusion protein, with the hormone-binding domain of the glucocorticoid receptor fused to Vent2, was injected into the D1 anterior cells of 32-cell stage embryos. Dexamethasone (Dex, 10^-6 M) was added to the media at the indicated stages, embryos were cultured until stage 35-37 and assayed by for1 and pdx1 in situ hybridization. Ectopic GR-Vent2 can repress foregut development up until stage 20, which is similar to the period when ectopically activated β-catenin can repress foregut development. No effect was observed on uninjected embryos treated with Dex, or on injected embryos without Dex.