Fig. 2

Temporally and Spatially Controlled Perturbation of the Tail Organizer

(A and B) Western blot for pSmad (A) and qRT-PCR for nascent transcription (B) of pooled, dissected tail buds from embryos treated with DMH1 or DMSO at the 5-somite stage. pSmad levels (A) and bmp4 and eve1 transcription are reduced 1–5 h post-DMH1 treatment (hpt) relative to DMSO-treated controls. n = 3 replicates for each time point.

(C–E) A local perturbation of the tail organizer is introduced in transient transgenics that express eve1 along with GFP using the tbx6l enhancer. On the right, an experimental image of a tail organizer is shown in which nuclei of all cells are labeled with nls-RFP (red) and transgenic cells are labeled with GFP (green). Transgenic expression of eve1 increases pSmad levels (D) as well as bmp2b, bmp4, and eve1 transcription (E). Blocking the Bmp receptor with DMH1 eliminates the increase in pSmad (D) and bmp4 and eve1 transcription, but not bmp2b transcription (E). Measurements were made 3 h post-DMH1 treatment. n = 3 experimental replicates for each condition.

(F) Representative images of body elongation phenotypes following perturbation of the tail organizer.

(G) A summary of the percent of embryos with body elongation defects. Total number of embryos from at least three experimental replicates are indicated.

Error bars denote the standard error.