Endogenous patterns of TGFbeta superfamily signaling during early Xenopus development.
Transforming growth factor beta (TGFbeta) superfamily signaling has been implicated in patterning of the early Xenopus embryo. Upon ligand stimulation, TGFbeta receptors phosphorylate Smad proteins at carboxy-terminal SS(V/M)S consensus motifs. Smads 1/5/8, activated by bone morphogenetic protein (BMP) signaling, induce ventral mesoderm whereas Smad2, activated by activin-like ligands, induces dorsal mesoderm. Although ectopic expression studies are consistent with roles for TGFbeta signals in early Xenopus embryogenesis, when and where BMP and activin-like signaling pathways are active endogenously has not been directly examined. In this study, we investigate the temporal and spatial activation of TGFbeta superfamily signaling in early Xenopus development by using antibodies specific for the type I receptor-phosphorylated forms of Smad1/5/8 and Smad2. We find that Smad1/5/8 and two distinct isoforms of Smad2, full-length Smad2 and Smad2(delta)exon3, are phosphorylated in early embryos. Both Smad1/5/8 and Smad2/Smad2(delta)exon3 are activated after, but not before, the mid-blastula transition (MBT). Endogenous activation of Smad2/Smad2(delta)exon3 requires zygotic transcription, while Smad1/5/8 activation at MBT appears to involve transcription-independent regulation. We also find that the competence of embryonic cells to respond to TGF(delta) superfamily ligands is temporally regulated and may be a determinant of early patterning. Levels of phospho-Smad1/5/8 and of phospho-Smad2/Smad2(delta)exon3 are asymmetrically distributed across both the animal-vegetal and dorsoventral axes. The timing of the development of these asymmetries differs for phospho-Smad1/5/8 and for phospho-Smad2/Smad2(delta)exon3, and the spatial distribution of phosphorylation of each Smad changes dramatically as gastrulation begins. We discuss the implications of our results for endogenous functions of BMP and activin-like signals as candidate morphogens regulating primary germ layer formation and dorsoventral patterning of the early Xenopus embryo.
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Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA. mwhitman@hms.harvard.edu
Lee
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Keller
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ten Dijke
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Whitman
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D000546:Amanitins / Q000494:pharmacology; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001755:Blastocyst / Q000378:metabolism; D019521:Body Patterning; D019485:Bone Morphogenetic Proteins / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005091:Exons; D005775:Gastrula / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D051381:Rats; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D051785:Smad Proteins; D051898:Smad1 Protein; D051899:Smad2 Protein; D051902:Smad5 Protein; D051904:Smad8 Protein; D013997:Time Factors; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D014981:Xenopus / Q000196:embryology*; D029867:Xenopus Proteins
D000546:Amanitins; D019485:Bone Morphogenetic Proteins; D004268:DNA-Binding Proteins; D008024:Ligands; C497402:MXD1 protein, Xenopus; D019384:Nucleic Acid Synthesis Inhibitors; D010750:Phosphoproteins; D020033:Protein Isoforms; C497399:SMAD1 protein, human; C497782:SMAD2 protein, human; C107514:SMAD9 protein, human; D051785:Smad Proteins; D051898:Smad1 Protein; D051899:Smad2 Protein; C497785:Smad2 protein, Xenopus; C497784:Smad2 protein, rat; D051902:Smad5 Protein; C497800:Smad5 protein, Xenopus; D051904:Smad8 Protein; C491622:Smad8 protein, Xenopus; C109462:Smad9 protein, rat; D015534:Trans-Activators; D016212:Transforming Growth Factor beta; D029867:Xenopus Proteins