Figure 4.

EGCs derived from cKit^V558Δ PGCs recapitulate survival defects and display lower levels of surface cKIT. (A) EGC cell lines were derived from PGCs of E11.5 embryos harboring Oct4ΔPE:GFP as schematized and cultured on STO feeder cells (gray). (B) By annexin V staining, apoptosis was slightly increased in cKit^{V558Δ/V558Δ} EGCs compared with WT or heterozygotes. (C) Similar levels of cKit transcript in all EGC genotypes were verified by qRT-PCR. (D) cKIT protein levels did not differ among all EGC genotypes by immunoblotting. (E–H) Live immunostaining with cKIT antibody revealed successively reduced surface expression on cKit^V558Δ/+ and cKit^{V558Δ/V558Δ} EGCs. SCF-induced downregulation of cKIT was comparable in all EGC genotypes (F). Similarly, cell surface cKIT reduction was observed in cKit^V558Δ/+ and cKit^{V558Δ/V558Δ} PGCs (G) but not on cKit^S830R/+ PGCs (H). *P < 0.05; bars represent mean ± SEM.