XB-ART-20663Proc Natl Acad Sci U S A. October 25, 1994; 91 (22): 10255-9.
A truncated bone morphogenetic protein receptor affects dorsal-ventral patterning in the early Xenopus embryo.
Bone morphogenetic proteins (BMPs), which are members of the transforming growth factor beta (TGF-beta) superfamily, have been implicated in bone formation and the regulation of early development. To better understand the roles of BMPs in Xenopus laevis embryogenesis, we have cloned a cDNA coding for a serine/threonine kinase receptor that binds BMP-2 and BMP-4. To analyze its function, we attempted to block the BMP signaling pathway in Xenopus embryos by using a dominant-negative mutant of the BMP receptor. When the mutant receptor lacking the putative serine/threonine kinase domain was expressed in ventral blastomeres of Xenopus embryos, these blastomeres were respecified to dorsal mesoderm, eventually resulting in the formation of a secondary body axis. These findings suggest that endogenous BMP-2 and BMP-4 are involved in the dorsal-ventral specification in the embryo and that ventral fate requires induction rather than resulting from an absence of dorsal specification.
PubMed ID: 7937936
PMC ID: PMC44998
Article link: Proc Natl Acad Sci U S A.
Genes referenced: actn1 bmp2 bmp4 bmpr1a bmpr1b gsc post tbxt tgfb1
Article Images: [+] show captions
|FIG. 1. Amino acid sequence and homology of mTFR11. (A) Translated amino acid sequence of the mTFR11 cDNA. The signal peptide and transmembrane domains are indicated by a single underline. Potential sites of N-linked glycosylation are indicated by asterisks. The ends of the kinase domain are indicated by arrows. Ten of the conserved cysteine residues in the extracellular domain are in reverse print. Type I box is indicated by a double underline. An arrowhead is under the tyrosine residue that changes to stop codon in the dominant negative BMP receptor. (B) Homology of the mTFR11/hALK-3 to other receptors for ligands of the TGF-(3 superfamily. Percent amino acid identity is indicated. Light-shaded boxes, extracellular domain; black boxes, transmembrane domain; dark-shaded boxes, type I box; hatched boxes, intracellular kinase domain. ALK-2, -3, -5, and -6, activin receptor-like kinases 2, 3, 5, and 6(26); xTFR11, Xenopus TFR11 (accession no. D32066); mTGFf3RI, mouse TGF-(3 type I receptor (27); mActRI, mouse activin type I receptor (28); mActRII, mouse activin type II receptor (13); mTGF-fBRII, mouse TGF-f type II receptor (29);daf4, C. elegans daf-4 gene product (30).|
|FIG. 2. Specific binding of iodinated BMP-4 to COS cells transfected with mTFR11. (A) 'MI-BMP4 binding to COS cells transfected with mTFR11. Binding was performed on cell monolayers as described below and was competitively inhibited with unlabeled BMP-4. (Inset) Scatchard analysis. o, pMV2; *, mTfr11/pMV2. (B) Specificity of mI-BMPA binding to COS cells transfected with mTFR11. Binding of 125I-BMP4 to COS cells transfected with mTFR11 was performed with or without increasing concentrations ofunlabeled ligands. *, BMP-2; o, BMP-4;m, activin A; *, TGF-.l1. (C) Chemical cross-linking ofiodinated BMP-4 to COS cells transfected with mTFR11/pMV2 orpMV2. COS cells were transfected with either pMV2 (lane 1) or mTFR11/pMV2 (lanes 2-5) and incubated with 5 ng of 125I-BMP-4 per ml (all lanes) and 500 ng ofBMP-4 per ml (lane 3), 500 ng of BMP-2 per ml (lane 4), or 500 ng of activin per ml (lane 5). Molecular masses are indicated in kDa.|
|FIG. 3. Truncated BMP receptor inhibits BMP signaling pathway in early Xenopus embryo. At the four-cell stage, embryos were injected with the indicated samples into the dorsal marginal region and allowed to develop until stage 37 or 38. (A) Distilled H20. (B) BMP-4 mRNA, 200 pg. (C) AmnTFR11 mRNA, 200 pg. (D) BMP-4 plus AmTFR11 mRNA, 200 pg of each.|
|FIG. 5. Effect of inhibition of BMP signaling pathway on expression of early mesodermal markers. AmTFR11 mRNA was injected into the marginal region of ventral blastomeres at the four-cell stage. Uninjected control embryo (A, C, and E), and AmTFR11-injected embryos (B, D, and F) were developed until early gastrula stage and subjected to whole-mount in situ analysis. The probe for goosecoid was used in A and B; the probe for Xpo was used in C and D; the probe for Xenopus brachyury was used in E and F. Views of the vegetal pole are shown and dorsal is up. Note that the expression of Xpo gene disappeared in the ventral part of embryo that received truncated BMP receptor mRNA (arrowheads) but that of brachyury gene, normally expressed through mesoderm, was not affected.|