Fig. S3

(A) Zebrafish embryos either mock-treated or treated with RA for in the indicated times and then fixed and stained for Dr-mespa RNA. Shown is a dorsal view of the PSM, with anterior oriented up. (B) Zebrafish embryos were either mock-treated, treated with RA, CHX or both for the indicated times and then stained for Dr-mespb RNA. Shown is a side view of the PSM with anterior oriented up. (C) Zebrafish embryos transgenic for Xl-mespb-GFP were treated for 1 h with RA and then stained for GFP expression. The expression of Xl-mespb-GFP continues to be upregulated after longer time periods of RA treatment (data not shown). (D) The 3.5 kb Xl-mespb-GFP (FL) or the 1.7 kb Xl-mespb-GFP (Stu) transgenes were introduced into a batch of embryos. Each batch was subdivided, either mock-treated or treated with RA for 1.5 h and then fixed and stained for GFP expression in parallel. About half of the embryos prepared with Xl-mespb-GFP-FL (Blue) show transgenic expression, and this frequency does not change upon RA treatment. By contrast, the frequency of transgene expression in embryos prepared with Xl-mespb-GFP-Stu (Red) drops significantly upon RA treatment, indicating that RA is not activating but repressing expression from Xl-mespb-GFP-Stu. At least 100 embryos were scored under each experimental condition.