Abstract
The level of TGF-β/bone morphogenetic protein (BMP) signaling through Smad is tightly regulated to ensure proper embryonic patterning and homeostasis. Here we show that Smad activation by TGF-β/BMP is blocked by a highly conserved phosphorylation event in the α-helix 1 region of Smad [T312 in Drosophila Smad1 (MAD)]. α-helix 1 phosphorylation reduces Smad interaction with TGF-β/BMP receptor kinase and affects all receptor-activated Smads except Smad3. Tissue culture and transgenic studies in Drosophila further demonstrate that the biological activity of MAD is repressed by T312 phosphorylation in vivo. Through RNAi screening of the kinome, we have identified Misshapen (Msn) and the mammalian orthologs TNIK, MINK1, and MAP4K4 as the kinases responsible for α-helix 1 phosphorylation. Targeted expression of an active form of Msn in the wing imaginal disk disrupted activation of endogenous MAD by Dpp and expression of the Dpp/MAD target gene. Msn kinases belong to the Ste20 kinase family that has been shown to act as MAP kinase kinase kinase kinase (MAP4K). Our findings thus reveal a function of Msn independent of its impact on MAP kinase cascades. This Smad inhibition mechanism by Msn likely has important implications for development and disease.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Animals
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Animals, Genetically Modified
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Binding Sites
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Bone Morphogenetic Protein Receptors / metabolism
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Bone Morphogenetic Proteins / metabolism
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Cell Line
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DNA-Binding Proteins / antagonists & inhibitors
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism
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Drosophila Proteins / antagonists & inhibitors
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Drosophila Proteins / deficiency
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Drosophila Proteins / genetics
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Drosophila Proteins / metabolism*
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Drosophila melanogaster / genetics
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Drosophila melanogaster / growth & development
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Drosophila melanogaster / metabolism
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Genes, Insect
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Humans
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Molecular Sequence Data
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Phosphorylation
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Protein Serine-Threonine Kinases / antagonists & inhibitors
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Protein Serine-Threonine Kinases / deficiency
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism*
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Protein Structure, Secondary
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RNA Interference
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Sequence Homology, Amino Acid
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Signal Transduction
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Smad Proteins / antagonists & inhibitors*
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Smad Proteins / chemistry
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Smad Proteins / genetics
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Smad Proteins / metabolism
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Transcription Factors / antagonists & inhibitors
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Transcription Factors / genetics
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Transcription Factors / metabolism
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Transforming Growth Factor beta / metabolism
Substances
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Bone Morphogenetic Proteins
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DNA-Binding Proteins
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Drosophila Proteins
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MAD protein, Drosophila
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Smad Proteins
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Transcription Factors
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Transforming Growth Factor beta
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msn protein, Drosophila
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Protein Serine-Threonine Kinases
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Bone Morphogenetic Protein Receptors