Figure 9. Hox genes effect on axis elongation involves Brachyury regulation downstream of the Wnt/βcatenin pathway.
(A) Consecutive electroporation of PM precursors with Cherry and then with Venus together with T (left panel), Hoxa13 (middle), or a combination of the two vectors (right). Arrowheads: anterior boundary of Cherry (red) and Venus (green) domains. (B) Ratio of Venus over Cherry domains. Dots: electroporated embryos. Bar indicates the mean. (C) Axis elongation velocity of embryos electroporated with control, Hoxa13, or co-electroporated with Hoxa13 and either high or low dose of T. (D) Q-RT PCR quantification of T expression in control or Hoxa13-expressing PM progenitor cells. (E–F) Luciferase activity (RLU) after over-expression of cTprLuc and CMV-Renilla together with either (E) control, Hoxc11 or Hoxa13 or (F) control, βcatLEF, Hoxa13 or Hoxa13+βcatLEF. (G–I) Luciferase assay measuring Wnt/βcatenin activity after over-expression of BATLuc and CMV-Renilla and (G) Hoxa13mutH, Hoxa13dn, Hoxa13+Hoxa13mutH or Hoxa13+Hoxa13dn. (H) Hoxd10mutH, Hoxd10dn, Hoxd10+Hoxd10mutH or Hoxd10+Hoxd10dn, (I) Hoxc11mutH, Hoxc11dn, Hoxc11+Hoxc11mutH or Hoxc11+Hoxc11dn. (J) Luciferase assay measuring Wnt/βcatenin activity from 28-somite stage dissected tail-buds after over-expression of BATLuc and CMV-Renilla constructs and either Hoxa13mutH or Hoxa13dn, or Hoxa13mutH with Hoxc11mutH or Hoxa13dn with Hoxc11dn, or Hoxa13mutH with Hoxc11mutH and Hoxd10mutH or Hox13dn with Hoxc11dn and Hoxd10dn. (K, L) Q-RT PCR quantification of T, Axin2 (K), and Fzd2 (L) expression in PM progenitors co-expressing either Hoxa13mutH with Hoxc11mutH and Hoxd10mutH or Hoxa13dn with Hoxc11dn and Hoxd10dn. Stars: p-value of the two-tailed Student's t-test applied between the different conditions. *p < 0.05; **p < 0.01; ***p < 0.005. Error bars: standard error to the mean (SEM).
Figure 9—figure supplement 1. Overexpression of T has no effect on Wnt activity.
