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Fig. 1. Effects of citral and BMS-453 on pronephros marker genes. Embryos were incubated during gastrulation with citral (B, E) or BMS-453 (C, F) or cultured in control medium (A, D). At the late tailbud stage 33, XSMP-30 (A–C) and XPax-2 (D–F) expression was analysed by whole mount in situ hybridization. Panels D′, E′, F′ are high magnification of the pronephric tubules area of panels D, E, F, respectively. XSMP-30 expression is severely reduced in embryos treated with the chemical inhibitors (B, C). XPax-2 expression in connective tubules and nephrostomes (arrowheads in panel D′) is lost in treated embryos (E′, F′) while expression in the duct is not affected. |
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Fig. 2. Ectopic expression of XCyp26 or dnRAR inhibits pronephros formation and expression of pronephros marker genes. Four-cell stage embryos were injected with XCyp26 (D–F) or dnRAR (G–I) encoding mRNA together with LacZ mRNA into the four blastomeres (A–I) or the two blastomeres fated to give rise to the left side (J–L). In the later case, the right side of the embryo is used as an internal control. A–I: In situ hybridization (purple staining) for XPax-2 (A, D, G) XSMP-30 (B, E, H) and XWT-1 (C, F, I). X-gal staining is revealed in red (D–I). In control embryos, XPax-2 is expressed in the tubules and the duct while XSMP-30 is restricted to the tubules and XWT-1 expressed in the glomus. No signal for these genes in either of theses structures was detected. J–L: Transverse histological section of tadpole stage 38. Several pronephric tubules (red circle) are clearly visible on the control side (K, and high magnification in panel J) whereas none is observed on the injected side (K, and high magnification in panel L). |
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Fig. 7. RA signalling is required during gastrulation for pronephros formation. Embryos were treated (B–F) or not (A) with BMS-453 during the time windows indicated in the cartoon, cultured until the late tailbud stage analysed for XPax-2 expression by whole mount in situ hybridization. Panels A′, B′, C′, D′, E′, F′ are tubules area enlargements of panels A, B, C, D, E, F respectively. The effect of BMS-453 on XPax-2 expression in the pronephric tubules is much more severe if embryos are treated during gastrulation (B, C, D) rather than after gastrulation is completed (E, F). |
