Fig. 6

Non-shed extra-embryonic Sdc2 is sufficient for embryonic organ primordia migration and fibrillogenesis. (A) Schematic of the GFP-sdc2 construct with an arrow marking the location of the putative juxtamembrane cleavage site. TM, transmembrane domain. (B,C) Confocal microscopy images reveal GFP (green) expression in embryonic tissue (arrow) with injection of the GFP-sdc2 construct at the single-cell stage (B) but restriction of GFP expression to the YSL/yolk in embryos (n=10) injected at the dome stage (C; arrow shows where embryonic tissue is located). The distribution of co-injected rhodamine-tagged control morpholino (red) correlates with the GFP expression. Insets in B and C show GFP distribution at higher magnification. e, embryo; y, yolk. (D) Schematic of the sdc2-hCD4JM construct. In this construct, 15 juxtamembrane amino acids, including the putative cleavage sequence for proteoglycan shedding, were replaced with a noncleavable sequence from the juxtamembrane domain of human CD4 (hCD4). (E) Co-injection of sdc2 morpholino and sdc2-hCD4JM mRNA into yolk at the dome stage (injection pattern shown in the circular diagram) rescued fibronectin fibrillogenesis in 7/9 embryos (fibronectin fibrils in green) and cardiac migration in 60% of embryos (n=70). (F) Schematic of the sdc2-ΔGAG construct that contains the N-terminal membrane localization sequence but lacks all GAG attachment sites. (G) In contrast to sdc2-hCD4JM, co-injection of sdc2 morpholino and sdc2-ΔGAG mRNA into the yolk at the dome stage did not rescue cardiac primordia migration (normal in only 39% of embryos, n=118).