| |
Figure 1. IGFBP-5 Is Expressed Dorsally and Has Anteriorizing Activity(A) SDS-PAGE of conditioned medium from 293Tcells labeled with 35S-cysteine and -methionine and transfected with GFP (control) or xIGFBP-5 cDNA. Protein bands at 38 kDa and 18 kDa correspond to xIGFBP-5 full-length protein and proteolytic cleavage products, respectively.(B) Schematic structure of IGFBP-5. SP, N-terminal signal peptide; IB, IGF binding domain; TY, thyroglobulin type I repeat. The GCGCCXXC sequence critical for IGF binding (Kalus et al., 1998) is indicated.(C) Sequence alignment of Xenopus and human IGFBP-5 proteins. The arrowhead indicates the predicted signal peptide cleavage site and the red and blue bars indicate the IG and TY, respectively.(D–H) Whole-mount in situ hybridization of xIGFBP-5 transcripts.(D) Animal view at the 4-cell stage.(E) Dorsal view at neural plate stage showing xIGFBP-5 expression in the dorsal midline.(F) Transverse section of the embryo at neural plate stage; expression is seen in floor plate (fp), notochord (nc), and dorsal endoderm (de).(G) Lateral view of tailbud tadpole.(H) Transverse section of tailbud embryo showing xIGFBP-5 expression in dorsal fin (df), floor plate (fp), and hypochord (hy).(I) Uninjected control.(J) Embryo radially injected with a total of 400 pg xIGFBP-5 mRNA into the animal pole of embryos at the 4- to 8-cell stage (93%, n = 106). Note the enlargement of the cement gland.(K) Uninjected embryo stained for Otx-2 at neurula stage.(L) Sibling injected dorsally with 100 pg of IGFBP-5 mRNA.(M) Ectopic expression of Otx-2 after ventral injection of 100 pg IGFBP-5 mRNA |
