Fig. 4

Talin 1 is required for endothelial cell elongation cell-autonomously. (A) Schematic representation of transplantation procedure. For these experiments, we transplanted cells at blastula stage from Tg(ve-cad:ve-cad-TS)^uq11bh donors to Tg(kdrl:Hsa.HRAS-mCherry)^s916 recipients. (B) Wild-type dorsal aorta (magenta) of recipient embryos at 2 dpf containing either wild-type donor ECs (top, yellow) or tln1^uq1al−/− mutant ECs (bottom, yellow). Scale bars: 10 µm. Greyscale images show donor ECs only. (C) Wild-type recipient dorsal aorta (magenta) of recipient embryos at 3 dpf containing wild-type donor ECs (top, yellow) or tln1^uq1al−/− mutant ECs (bottom, yellow). Scale bars: 10 µm. Greyscale images show donor ECs only. (D) Quantification of EC elongation of transplanted ECs during dorsal aorta maturation from 2 dpf to 3 dpf, n=4 replicates; 2 dpf data, n=15 sibling ECs and n=14 tln1^uq1al−/− mutant ECs; 3 dpf data, n=15 sibling ECs and n=15 tln1^uq1al−/− mutant ECs. *P<0.05 for 2 dpf sibling versus tln1^uq1al−/− mutant (Mann-Whitney test). ***P<0.001 for 3 dpf sibling versus 3 dpf tln1^uq1al−/− mutant (unpaired t-test). **P<0.01 for 2 dpf sibling versus 3 dpf sibling (Mann-Whitney test). **P<0.01 for 2 dpf tln1^uq1al−/− mutant versus 3 dpf tln1^uq1al−/− mutant (Mann-Whitney test). (E) Schematic representation of complementary transplantation experiments. We transplanted cells from Cascade Blue-injected tln1 crispants carrying two transgenes: Tg(ve-cad:ve-cad-TS)^uq11bh and Tg(fli1ep:nls-mCherry)^uq37bh. Cells were transplanted into Tg(ve-cad:ve-cad-TS)^uq11bh wild-type embryos at blastula stages. (F) A small graft of Tln1-deficient ECs [indicated by Cascade Blue and Tg(fli1ep:nls-mCherry)^uq37bh expression] within a wild-type DA at 2 dpf. Scale bar: 25 µm. (G) Endothelial-restricted Tln1-deficient ECs, imaged at 3 dpf. Scale bar: 25 µm. (H) Quantifications of EC shape of Tln1-deficient donor cells versus neighbouring wild-type ECs at 2 and 3 dpf, showing that Tln1-deficient cells fail to elongate while neighbouring ECs are unaffected and elongate significantly over time. Graphs show replicate averages (large circles); smaller circles indicate individual data points of each replicate (colour matched), n=2 replicates; 2 dpf data, n=25 wild-type ECs and n=11 tln1^uq1al−/− mutant ECs; 3 dpf data, n=14 wild-type ECs and n=10 tln1^uq1al−/− mutant ECs. Mann-Whitney test for all comparisons (*P<0.01, **P<0.005, ****P<0.0001).