Figure 3

Spry2 gain-of-function inhibits somite myogenesis. Electroporation of pCAB-Spry2 full length or RFP-MKP3 expression constructs into epithelial somites at HH16, as indicated, followed by 24-hour incubation to HH21/22. (A-C) Whole mount in situ hybridisation for MyoD (purple) and GFP or RFP (red), (Ai) is a section of embryo in (A). (A, Ai) Spry2 expression led to loss of MyoD in transfected cells, arrows in (A) indicate the targeted electroporated somites and lines indicate the level of sectioning in (Ai). (B) Electroporation of an empty pCAB-IRES-GFP expression vector (used as a control) into somites which showed normal MyoD expression (arrows). (C) Loss of MyoD was observed in cells electroporated with RFP-Mkp3 (arrows); (Ci) RFP detected by fluorescent filter Alexa-Fluor-465 in the same electroporated somites in (C). (D) Control of unelectroporated embryo showing normal MyoD expression. Magnifications: 20x in (A, B, C & D), 24x in (Ci), 200x in (Ai).