| |
Fig. 3. BMP-3b and BMP-3 have apparently different dorsalizing activities in Xenopus embryos. (A–C) Xenopus embryos injected with BMP-3b or BMP-3
mRNA. Overexpression of BMP-3b led to ectopic head formation, whereas BMP-3 generated aberrant tail formation. Synthetic mRNAs (1 ng each) encoding
xBMP-3b (A), xBMP-3 (B), or -galactosidase ( -gal; C) were injected into marginal zone of two ventral blastomeres at the four cell stage. (D–G)
Differentiation of ventral marginal zone explants (VMZs) injected with BMP-3b or BMP-3. Typical explants are shown in circles. (D) VMZs injected with
rBMP-3b RNA generated head-like structures, consisting of an eye and cement glands (68%, n 15). (E) VMZs injected with rBMP-3 elongated and formed
melanocytes (93%, n 15). (F) Control VMZs developed cylindrical structures. (G) These VMZs were harvested at stage 41 and scored by RT-PCR.
Injection of BMP-3b and BMP-3 caused expression of dorsal markers (cardiac actin and NCAM), and repression of a ventral marker ( globin). BMP-3b
also elevated anterior tissue markers such as cement gland (xAG1), lens ( B1-crystallin), and retina (rhodopsin). EF1 is an internal loading control. Emb
RT and RT are controls for cDNA synthesis and for genomic DNA contamination, respectively. (H) Analysis of early markers in embryos coinjected
with -gal (100 pg), and either BMP-3b or BMP-3 (1 ng). Light blue staining represents area injected with BMP-3b (left) or BMP-3 (center). Brown staining
represents expression of Otx2 (top panel), and Xwnt-8 (middle panel), and cerberus (bottom panel). All panels are vegetal views of embryos with dorsal aspect
top. Injection of BMP-3b mRNA triggered ectopic Otx2 expression (top left), whereas injection of BMP-3b and BMP-3 reduced endogenous Xwnt8
expression (middle). BMP-3b did not induce cerberus expression, indicating that BMP-3b is involved in head formation independently of, or downstream
from, cerberus. |
