Abstract
Smads proteins play a key role in the intracellular signaling of the transforming growth factor (TGF)-beta family of growth factors, which exhibits a diverse set of cellular responses, including cell proliferation and differentiation. In particular, Smad7 acts as an antagonist of TGF-beta signaling, which could determine the intensity or duration of its signaling cascade. In this study we identified a protein inhibitor of activated STAT (signal transducers and activators of transcription), PIASy, as a novel interaction partner of Smad7 by yeast two-hybrid screening using the MH2 domain of Smad7 as bait. The association of Smad7 and PIASy was confirmed using co-expressed tagged proteins in 293T cells. Moreover, we found that other Smads including Smad3 also associated with PIASy through its MH2 domain, and PIASy suppressed TGF-beta-mediated activation of Smad3. PIASy also stimulated the sumoylation of Smad3 in vivo. Furthermore, endogenous PIASy expression was induced by TGF-beta in Hep3B cells. These findings provide the first evidence that a PIAS family protein, PIASy, associates with Smads and involves the regulation of TGF-beta signaling using the negative feedback loop.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Blotting, Northern
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COS Cells
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Carrier Proteins / metabolism*
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Cell Differentiation
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Cell Division
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Cell Line
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DNA-Binding Proteins / chemistry
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DNA-Binding Proteins / metabolism*
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Fluorescent Antibody Technique, Indirect
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Gene Expression Regulation*
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Humans
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Immunoblotting
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Intracellular Signaling Peptides and Proteins*
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Poly-ADP-Ribose Binding Proteins
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Protein Binding
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Protein Inhibitors of Activated STAT
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Protein Structure, Tertiary
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Recombinant Fusion Proteins / metabolism
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Recombinant Proteins / metabolism
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Reverse Transcriptase Polymerase Chain Reaction
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SUMO-1 Protein / metabolism
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Saccharomyces cerevisiae / metabolism
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Signal Transduction*
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Smad3 Protein
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Smad6 Protein
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Smad7 Protein
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Time Factors
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Trans-Activators / chemistry
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Trans-Activators / metabolism*
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Transcription, Genetic
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Transfection
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Transforming Growth Factor beta / metabolism*
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Tumor Cells, Cultured
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Two-Hybrid System Techniques
Substances
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Carrier Proteins
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DNA-Binding Proteins
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Intracellular Signaling Peptides and Proteins
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PIAS4 protein, human
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Poly-ADP-Ribose Binding Proteins
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Protein Inhibitors of Activated STAT
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Recombinant Fusion Proteins
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Recombinant Proteins
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SMAD3 protein, human
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SMAD6 protein, human
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SMAD7 protein, human
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SUMO-1 Protein
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Smad3 Protein
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Smad6 Protein
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Smad7 Protein
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Trans-Activators
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Transforming Growth Factor beta