Abstract
Cellular signal transduction pathways are usually studied following administration of an external stimulus. However, disease-associated aberrant activity of the pathway is often due to misregulation of the equilibrium state. The transcription factor NF-kappaB is typically described as being held inactive in the cytoplasm by binding its inhibitor, IkappaB, until an external stimulus triggers IkappaB degradation through an IkappaB kinase-dependent degradation pathway. Combining genetic, biochemical, and computational tools, we investigate steady-state regulation of the NF-kappaB signaling module and its impact on stimulus responsiveness. We present newly measured in vivo degradation rate constants for NF-kappaB-bound and -unbound IkappaB proteins that are critical for accurate computational predictions of steady-state IkappaB protein levels and basal NF-kappaB activity. Simulations reveal a homeostatic NF-kappaB signaling module in which differential degradation rates of free and bound pools of IkappaB represent a novel cross-regulation mechanism that imparts functional robustness to the signaling module.
Publication types
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Comparative Study
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Research Support, N.I.H., Extramural
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Animals
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Blotting, Western
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Cells, Cultured / metabolism
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Computer Simulation*
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Electrophoretic Mobility Shift Assay
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Fibroblasts / metabolism
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Half-Life
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Homeostasis / physiology*
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I-kappa B Kinase / deficiency
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I-kappa B Kinase / genetics
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I-kappa B Kinase / physiology
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I-kappa B Proteins / genetics
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I-kappa B Proteins / metabolism*
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Kinetics
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Leupeptins / pharmacology
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Mice
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Mice, Knockout
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Models, Biological*
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NF-KappaB Inhibitor alpha
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NF-kappa B / metabolism*
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Phosphorylation
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Proteasome Endopeptidase Complex / metabolism
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Protein Binding
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Protein Interaction Mapping
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Protein Processing, Post-Translational
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Proto-Oncogene Proteins / genetics
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Proto-Oncogene Proteins / metabolism*
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Signal Transduction / physiology*
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Tumor Necrosis Factor-alpha / pharmacology
Substances
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I kappa B beta protein
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I-kappa B Proteins
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Leupeptins
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NF-kappa B
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Nfkbia protein, mouse
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Nfkbie protein, mouse
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Proto-Oncogene Proteins
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Tumor Necrosis Factor-alpha
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NF-KappaB Inhibitor alpha
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I-kappa B Kinase
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Proteasome Endopeptidase Complex
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benzyloxycarbonylleucyl-leucyl-leucine aldehyde