Fig. 4: TGFBR1 in the DML and TGFBR2 in the posterior somite are necessary for fusion.

a–c DML-derived myocytes electroporated with MLC promoter driving expression of constitutively active (CA, b) and dominant-negative (DN, c) TGFBR1 variants, together with membrane GFP (green) and nuclear mCherry (red). d Column graph for a–c showing the population of electroporated myocytes containing the indicated number of nuclei relative to their controls (in %). e–g DML-derived myocytes electroporated with a CRISPR/Cas9 construct targeting the chicken TGFBR1 or TGFBR2 sequences together with membrane GFP (green) and nuclear mCherry (red). h Column graph for e–g showing the population of electroporated myocytes containing the indicated number of nuclei relative to their controls (in %). Solid colour columns indicate overall significant difference with controls. Striped colour columns indicate non-significant difference with controls. i, j Myocytes electroporated in the posterior border of the dermomyotome with a CRISPR/Cas9 construct targeting the chicken TGFBR2 sequence together with membrane GFP (green) and nuclear mCherry (red). k Column graph for i-j showing the population of electroporated myocytes containing the indicated number of nuclei relative to their controls (in %). Embryos in a–c, e–g, i, j and l–n were fixed at E5.5 and stained against GFP and RFP antibodies. l–n Somites electroporated in the posterior border (PL) with cytoplasmic BFP (in blue) as electroporation marker and the TGF β reporter we identified, upstream of eGFP (in green). Somites were analysed 6 h (l), one day (m) and 3 days (n) after electroporation. o Column graph for l–n showing the percentage of electroporated cells that express the reporter. Statistical analyses: CA TGFBR1: \(\bar x\): 1.88; n = 6; Ctrl: \(\bar x\): 2.46; n = 13; P-value =0.0004; DN TGFBR1: \(\bar x\): 3.56; n = 9; Ctrl: \(\bar x\): 2.46; n = 13; P-value < 0.0001; CRISPR DML TGFBR1: \(\bar x\): 3.02; n = 21; Ctrl: \(\bar x\): 2.43; n = 14; P-value < 0.0001; CRISPR DML TGFBR2: \(\bar x\): 2.47; n = 21; Ctrl: \(\bar x\): 2.43; n = 14; P-value: 0.97; CRISPR PB TGFBR2: \(\bar x\): 2.94; n = 48; Ctrl: \(\bar x\): 2.61; n = 32; P-value = 0.0009. Arrowheads point to cell nuclei in select fibres. ***P < 0.001, ns = p > 0.05. Error bars in d, h, k, o: SEM. Scale bars: 50 μm. Source data are provided (see ‘Data availability’).