Fig. S4

Fig. S4 A furin-resistant form of Vegfc (Vegfc^ss) is secreted and can partially rescue lymphatic development in vegfc^um18 mutant embryos. (A) Vegfc structure showing location of the furin cleavage domain and the conserved amino acid sequences from zebrafish (Dr), human (Hs), mouse (Mm) and rat (Rn) Vegfc proteins. The amino acid substitutions to prevent furin cleavage are shown. (B) The proportion of embryos with indicated genotypes that displayed TD formation following injection of an endothelial driven egfp- 2Acherry or vegfc^ss-2Acherry transgene. (C) Zebrafish Vegfc structure showing location of the hemagglutinin (HA) epitope tag at the C-terminus used to detect expression of furin-resistant Vegfc (Vegfc^ss). Western blot shows expression of wild-type zebrafish Vegfc or Vegfc^ss in cell lysates or extracellular media from transiently transfected NIH3T3 cells. In cell lysates, full-length Vegfc can be detected in both cases. Interestingly, we can detect a small amount of full-length wild-type Vegfc in the medium, along with the C-terminal domain that has been cleaved. Furthermore, we are able to detect furin-resistant Vegfc^ss in the medium, the vast majority of which remains unprocessed.