Fig. 2

Phospho-Smad 1, 5, 8 (pSmad) distribution. A-C, E: Normal pattern of pSmad labeling. D, F: Pattern of pSmad following overexpression of Bmp2b. A: Pattern of phosphorylated Smad 1, 5, and 8 (red) in the anterior somites of 12S stage embryo. pSmad is distributed throughout the somite. A subset of lateral pSmad+ cells also express Pax7 (green, arrowheads). B: Pattern of pSmad (red) in the posterior somites of a 12S-stage embryo. C-F: Transverse sections labeled for pSmad (red) and Pax7 (green). C: At 20S, pSmad is found in Pax7+ myogenic precursors at the dorsal and ventral regions of the somite (arrowheads). Insets show a Pax7+ nucleus (green) that is also pSmad+ (red). pSmad labeling is also seen in the medial portion of the somite. D: pSmad in a 20S embryo following heat-shock induction of Bmp2b overexpression at 17S. Ectopic activation of BMP signaling is still evident at this time, including in the Pax7-expressing population (green). Because of the mosaic distribution of the plasmid DNA following injection, the pattern of ectopic activation can vary. In this case, high levels of ectopic pSmad (red) are seen in the ventral somite. E: pSmad labeling in a 24-hr embryo. A subset of Pax7+ myogenic precursors (green) at the dorsal and ventral regions of the somite are also pSmad+ (red). Insets indicate cells that are both Pax7+ and pSmad+. F: Distribution of pSmad at 24 hr following heat-shock induction of Bmp2b overexpression at 17S. A high level of ectopic BMP activation is evident throughout the somite (pSmad, red), including a high proportion of the Pax7-expressing myogenic precursors (green). Scale bars = 50 μm in B (for A,B), 25 μm in D (for C-F).