Evidence for antagonism of BMP-4 signals by MAP kinase during Xenopus axis determination and neural specification.
We have previously shown that mitogen-activated protein (MAP) kinase activity is required for neural specification in Xenopus. In mammalian cells, the BMP-4 effector Smad1 is inhibited by phosphorylation at MAP kinase sites (Kretzschmar et al., 1997). To test the hypothesis that MAP kinase inhibits the BMP-4/Smad1 pathway during early Xenopus development, we have generated a Smad1 mutant lacking the MAP kinase phosphorylation sites (M4A-Smad1) and compared the effects of wild-type (WT)- and M4A-Smad1 on axial pattern and neural specification in Xenopus embryos. Although overexpression of either WT- or M4A-Smad1 produced ventralized embryos, at each mRNA concentration, M4A-Smad1 had a greater ventralizing effect than WT-Smad1. Interestingly, overexpression of either form of Smad1 in ventral blastomeres disrupted posterior pattern and morphogenesis; again, more severe defects were produced by expression of M4A-Smad1 than by equal amounts of WT-Smad1. Ectodermal expression of M4A-Smad1 disrupted expression of the anterior neural gene otx2 in vivo and inhibited neural specification in response to endogenous signals in mesoderm-ectoderm recombinates. In contrast, overexpression of WT-Smad1 at identical levels had little effect on either neural specification or otx2 expression. Comparisons of protein levels following overexpression of either WT- or M4A-Smad1 indicate that WT-Smad1 may be slightly more stable than M4A-Smad1; thus, differences in stability cannot account for the increased effectiveness of M4A-Smad1. Our results demonstrate that mutations disrupting the MAPK phosphorylation sites act collectively as a gain-of-function mutation in Smad1 and that inhibitory phosphorylation of Smad1 may be a significant mechanism for the regulation of BMP-4/Smad1 signals during Xenopus development.
Sater
Amy K
AK
Division of Molecular and Cell Biology, Department of Biology and Biochemistry, University of Houston, Houston, TX 77204, USA. asater@uh.ca
El-Hodiri
Heithem M
HM
Goswami
Mousumi
M
Alexander
Tara B
TB
Al-Sheikh
Oday
O
Etkin
Laurence D
LD
Akif Uzman
J
J
D000818:Animals; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D004475:Ectoderm / Q000737:chemistry; D015870:Gene Expression; D008648:Mesoderm / Q000737:chemistry; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism* / Q000494:pharmacology; D009420:Nervous System / Q000196:embryology* / Q000254:growth & development; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D051785:Smad Proteins; D015534:Trans-Activators / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D029867:Xenopus Proteins / Q000378:metabolism*; D014982:Xenopus laevis / Q000196:embryology* / Q000235:genetics / Q000378:metabolism
D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D004268:DNA-Binding Proteins; C497402:MXD1 protein, Xenopus; D012333:RNA, Messenger; D051785:Smad Proteins; D015534:Trans-Activators; D029867:Xenopus Proteins; C525812:bmp4 protein, Xenopus; D020928:Mitogen-Activated Protein Kinases