Abstract
Nuclear receptor coactivators (NRCoAs) are nuclear hormone receptor-associated regulatory proteins that interact with members of the nuclear receptor superfamily in the presence of their cognate ligand, enhancing their transcriptional activity. The identification of ubiquitin-proteasome pathway proteins as coactivators provides evidence that ubiquitin-proteasome-mediated protein degradation plays an integral role in eukaryotic gene transcription. It has also been observed that nuclear receptors themselves are ubiquitinated and degraded in a hormone-dependent manner and that ubiquitin-proteasome function is essential for most nuclear receptors to function as transactivators. Here, we show that specific ubiquitin-proteasome pathway enzymes target specific NRCoA proteins in vivo and in vitro. First, using a temperature-sensitive cell line that contains a thermolabile ubiquitin-activating E1 enzyme, we confirmed that NRCoA proteins are targets of the ubiquitin-proteasome pathway. Then using coimmunoprecipitation studies, we also demonstrate that in vivo, NRCoA proteins are ubiquitinated. Finally, we illustrate that in vitro, NRCoA ubiquitination and degradation depend on the ubiquitin-activating enzyme (E1) and on specific ubiquitin-conjugating enzymes (E2) for each of the coactivators.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Binding Sites
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CREB-Binding Protein
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Cysteine Endopeptidases / metabolism*
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Cysteine Proteinase Inhibitors / pharmacology
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HeLa Cells
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Histone Acetyltransferases
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Humans
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Leupeptins / pharmacology
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Multienzyme Complexes / antagonists & inhibitors
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Multienzyme Complexes / metabolism*
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Mutation
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Nuclear Proteins / drug effects
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Nuclear Proteins / metabolism
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Nuclear Receptor Coactivator 1
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Nuclear Receptor Coactivator 2
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Nuclear Receptor Coactivator 3
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Proteasome Endopeptidase Complex
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Proteins / metabolism*
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Receptors, Cytoplasmic and Nuclear / metabolism*
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Trans-Activators / drug effects
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Trans-Activators / metabolism
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Transcription Factors / drug effects
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Transcription Factors / metabolism
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Ubiquitin / metabolism*
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Ubiquitin-Activating Enzymes / genetics
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Ubiquitin-Activating Enzymes / metabolism*
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Ubiquitin-Conjugating Enzymes / genetics
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Ubiquitin-Conjugating Enzymes / metabolism*
Substances
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Cysteine Proteinase Inhibitors
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Leupeptins
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Multienzyme Complexes
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Nuclear Proteins
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Nuclear Receptor Coactivator 2
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Proteins
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Receptors, Cytoplasmic and Nuclear
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Trans-Activators
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Transcription Factors
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Ubiquitin
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CREB-Binding Protein
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CREBBP protein, human
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Histone Acetyltransferases
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NCOA1 protein, human
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Nuclear Receptor Coactivator 1
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Nuclear Receptor Coactivator 3
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Ubiquitin-Conjugating Enzymes
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Cysteine Endopeptidases
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Proteasome Endopeptidase Complex
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Ubiquitin-Activating Enzymes
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benzyloxycarbonylleucyl-leucyl-leucine aldehyde