PIAS3 suppresses NF-kappaB-mediated transcription by interacting with the p65/RelA subunit.
Nuclear factor-kappaB (NF-kappaB) is a transcription factor critical for key cellular processes, including immune response, apoptosis, and cell cycle progression. A yeast two-hybrid screening, using the Rel homology domain (RHD) of the p65 subunit (RelA) of NF-kappaB as bait, led to the isolation of PIAS3, previously identified as a specific ... inhibitor of STAT3. We show that PIAS3 can directly associate with p65 using an in vitro pull-down and in vivo coimmunoprecipitation assays. When overexpressed, PIAS3 inhibits NF-kappaB-dependent transcription induced by treatment with tumor necrosis factor alpha (TNF-alpha) or interleukin-1beta or by overexpression of TNF family receptors such as RANK, TNFR1, and CD30 or signal transducers of TNF receptor-associated factors (TRAFs), including TRAF2, TRAF5, and TRAF6. Downregulation of PIAS3 by RNA interference reverses its effect on TNF-alpha-mediated NF-kappaB activation. We found that an N-terminal region of PIAS3 is necessary for both the interaction with p65 and the transcriptional suppression activity. In addition, we found that an LXXLL coregulator signature motif located within the N-terminal region of PIAS3 is the minimal requirement for the interaction with p65. Furthermore, we demonstrate that PIAS3 interferes with p65 binding to the CBP coactivator, thereby resulting in a decreased NF-kappaB-dependent transcription. Taken together, these data suggest that PIAS3 may function in vivo as a modulator in suppressing the transcriptional activity of p65.
Mesh Terms:
Amino Acid Motifs, Animals, Antigens, CD, Antigens, CD30, Blotting, Western, Carrier Proteins, Cell Cycle, Cell Line, Dose-Response Relationship, Drug, Enzyme Activation, Glutathione Transferase, Glycoproteins, Humans, Interleukin-1, Luciferases, NF-kappa B, Osteoprotegerin, Plasmids, Precipitin Tests, Protein Binding, Protein Structure, Tertiary, Proteins, RNA Interference, RNA, Small Interfering, Receptors, Cytoplasmic and Nuclear, Receptors, Tumor Necrosis Factor, Receptors, Tumor Necrosis Factor, Type I, Recombinant Fusion Proteins, Signal Transduction, TNF Receptor-Associated Factor 5, TNF Receptor-Associated Factor 6, Time Factors, Transcription Factor RelA, Transcription, Genetic, Transfection, Tumor Necrosis Factor-alpha, Two-Hybrid System Techniques
Amino Acid Motifs, Animals, Antigens, CD, Antigens, CD30, Blotting, Western, Carrier Proteins, Cell Cycle, Cell Line, Dose-Response Relationship, Drug, Enzyme Activation, Glutathione Transferase, Glycoproteins, Humans, Interleukin-1, Luciferases, NF-kappa B, Osteoprotegerin, Plasmids, Precipitin Tests, Protein Binding, Protein Structure, Tertiary, Proteins, RNA Interference, RNA, Small Interfering, Receptors, Cytoplasmic and Nuclear, Receptors, Tumor Necrosis Factor, Receptors, Tumor Necrosis Factor, Type I, Recombinant Fusion Proteins, Signal Transduction, TNF Receptor-Associated Factor 5, TNF Receptor-Associated Factor 6, Time Factors, Transcription Factor RelA, Transcription, Genetic, Transfection, Tumor Necrosis Factor-alpha, Two-Hybrid System Techniques
J. Biol. Chem.
Date: Jun. 04, 2004
PubMed ID: 15140884
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