DEV02579 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure 1 -

  Fig. S1. TSKs siRNA depleted the level of TSK protein production. COS-7 cells were transfected with TSKs, TSKA or control1 siRNA plus TSKA-Myc or TSKB-Myc expression plasmid, and then the level of TSKA and TSKB proteins were examined by immunoblotting.

• Supplemental Figure 2 -

  Fig. S2. TSK depletion doesn’t effect on the expression of chordin and goosecoid. (A) The siRNA solution including Fast Green was microinjected in the space between the vitelline membrane and epiblast. (B) After electroporation, Hensen’s node and the anterior fourth of the primitive streak were surgically removed. GFP was expressed in the central area of the embryo after 6 hours incubation. (C-N) After electroporation of control1 (D,H,L), TSKA (E,I,M) or TSKs (F,J,N) siRNA, Hensen’s node and the anterior fourth of the primitive streak were surgically removed and the expression of TSK (F), chordin (G-J) and goosecoid (K-N) were examined by in situ hybridization. (C,G,K) Hensen’s node and the anterior fourth of the primitive streak of control embryos were surgically removed, and then examined the expression of Tsukushi (C), chordin (G) and goosecoid (K) without electroporation. The endogenous and upregulated expression of TSK (arrowhead in F) is nearly fully depleted in the presence of TSKs siRNA, but TSK is ectopically induced at both sides of the stump of the primitive streak (arrows in F).

• Supplemental Figure 3 -

  Fig. S3. Misexpression of VG1 in the area pellucida induces TSK. Cells secreting Vg1 were grafted into the anterior third of the area pellucida of stage XI-XII embryos, and these were cultured for 12 or 24 hours before in situ hybridization for TSK. Arrows indicate ectopic expression of TSK in the host embryos.