Abstract
Here we report the use of fluorescence recovery after photobleaching (FRAP) to examine the intranuclear dynamics of fluorescent oestrogen receptor-alpha (ER). After bleaching, unliganded ER exhibits high mobility (recovery t1/2 < 1 s). Agonist (oestradiol; E2) or partial antagonist (4-hydroxytamoxifen) slows ER recovery (t1/2 approximately 5-6 s), whereas the pure antagonist (ICI 182,780) and, surprisingly, proteasome inhibitors each immobilize ER to the nuclear matrix. Dual FRAP experiments show that fluorescent ER and SRC-1 exhibit similar dynamics only in the presence of E2. In contrast to reports that several nuclear proteins show uniform dynamics, ER exhibits differential mobility depending upon several factors that are linked to its transcription function.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Bacterial Proteins / analysis
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Biological Transport / drug effects
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Biological Transport / genetics*
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Cysteine Endopeptidases / metabolism*
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Cysteine Proteinase Inhibitors / pharmacology
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Dactinomycin / pharmacology
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Estradiol / analogs & derivatives*
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Estradiol / pharmacology*
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Estrogen Antagonists / pharmacology*
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Estrogen Receptor alpha
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Fulvestrant
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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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Ligands
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Luminescent Proteins / analysis
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Microscopy, Fluorescence / methods
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Multienzyme Complexes / antagonists & inhibitors
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Multienzyme Complexes / metabolism*
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Nuclear Matrix / drug effects
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Nuclear Matrix / metabolism*
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Nuclear Receptor Coactivator 1
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Proteasome Endopeptidase Complex
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Protein Structure, Tertiary / drug effects
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Protein Structure, Tertiary / physiology
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Receptors, Estrogen / drug effects
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Receptors, Estrogen / genetics
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Receptors, Estrogen / metabolism*
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Tamoxifen / analogs & derivatives*
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Tamoxifen / pharmacology
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Transcription Factors / drug effects
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Transcription Factors / metabolism
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Transcription, Genetic / drug effects
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Transcription, Genetic / physiology*
Substances
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Bacterial Proteins
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Cysteine Proteinase Inhibitors
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Estrogen Antagonists
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Estrogen Receptor alpha
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Leupeptins
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Ligands
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Luminescent Proteins
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Multienzyme Complexes
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Receptors, Estrogen
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Transcription Factors
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yellow fluorescent protein, Bacteria
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Tamoxifen
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afimoxifene
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Dactinomycin
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Fulvestrant
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Estradiol
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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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Cysteine Endopeptidases
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Proteasome Endopeptidase Complex
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benzyloxycarbonylleucyl-leucyl-leucine aldehyde