Abstract
TGF-beta induces phosphorylation of the transcription factors Smad2 and Smad3 at the C terminus as well as at an interdomain linker region. TGF-beta-induced linker phosphorylation marks the activated Smad proteins for proteasome-mediated destruction. Here, we identify Nedd4L as the ubiquitin ligase responsible for this step. Through its WW domain, Nedd4L specifically recognizes a TGF-beta-induced phosphoThr-ProTyr motif in the linker region, resulting in Smad2/3 polyubiquitination and degradation. Nedd4L is not interchangeable with Smurf1, a ubiquitin ligase that targets BMP-activated, linker-phosphorylated Smad1. Nedd4L limits the half-life of TGF-beta-activated Smads and restricts the amplitude and duration of TGF-beta gene responses, and in mouse embryonic stem cells, it limits the induction of mesoendodermal fates by Smad2/3-activating factors. Hierarchical regulation is provided by SGK1, which phosphorylates Nedd4L to prevent binding of Smad2/3. Previously identified as a regulator of renal sodium channels, Nedd4L is shown here to play a broader role as a general modulator of Smad turnover during TGF-beta signal transduction.
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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Cell Line
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Cells, Cultured
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Embryonic Stem Cells / cytology
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Embryonic Stem Cells / drug effects
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Embryonic Stem Cells / metabolism
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Endosomal Sorting Complexes Required for Transport / genetics
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Endosomal Sorting Complexes Required for Transport / metabolism*
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HeLa Cells
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Humans
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Immediate-Early Proteins / genetics
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Immediate-Early Proteins / metabolism
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Immunoblotting
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Mice
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Molecular Sequence Data
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Nedd4 Ubiquitin Protein Ligases
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Phosphorylation / drug effects
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Polyubiquitin / metabolism
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Protein Binding
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism
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RNA Interference
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Sequence Homology, Amino Acid
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Signal Transduction
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Smad2 Protein / genetics
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Smad2 Protein / metabolism*
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Smad3 Protein / genetics
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Smad3 Protein / metabolism*
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Transforming Growth Factor beta / pharmacology*
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Ubiquitin-Protein Ligases / genetics
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Ubiquitin-Protein Ligases / metabolism*
Substances
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Endosomal Sorting Complexes Required for Transport
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Immediate-Early Proteins
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SMAD2 protein, human
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SMAD3 protein, human
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Smad2 Protein
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Smad3 Protein
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Transforming Growth Factor beta
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Polyubiquitin
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Nedd4 Ubiquitin Protein Ligases
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Nedd4L protein, human
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Nedd4l protein, mouse
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SMURF1 protein, human
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Ubiquitin-Protein Ligases
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Protein Serine-Threonine Kinases
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serum-glucocorticoid regulated kinase