Abstract
We investigated the role of Heat shock protein 90 (Hsp90) in vitamin D action in Caco-2 cells using geldanamycin (GA) to block Hsp90 function and RNA interference to reduce Hsp90beta expression. When cells were exposed to GA, vitamin D-mediated gene expression and transcriptional activity were inhibited by 69% and 54%, respectively. Gel shift analysis indicated that GA reduced vitamin D-mediated DNA binding activity of the vitamin D receptor (VDR). We tested the specific role of Hsp90beta by knocking down its expression with stably expressed short hairpin RNA. Vitamin D-induced gene expression and transcriptional activity were reduced by 90% and 80%, respectively, in Hsp90beta-deficient cells. Nuclear protein for VDR and RXRalpha, its heterodimer partner, were not reduced in Hsp90beta-deficient cells. These findings indicate that Hsp90beta is needed for optimal vitamin D responsiveness in the enterocyte and demonstrate a specific role for Hsp90beta in VDR signaling.
MeSH terms
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Benzoquinones / pharmacology
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Caco-2 Cells
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Calcitriol / pharmacology*
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DNA / metabolism
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Electrophoretic Mobility Shift Assay
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Enterocytes / drug effects*
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Enterocytes / metabolism*
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Enzyme Inhibitors / pharmacology
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Gene Expression Regulation / drug effects
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HSP90 Heat-Shock Proteins / antagonists & inhibitors
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HSP90 Heat-Shock Proteins / genetics
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HSP90 Heat-Shock Proteins / physiology*
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Humans
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Lactams, Macrocyclic / pharmacology
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Polymerase Chain Reaction
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Protein Binding / drug effects
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RNA, Messenger / metabolism
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RNA, Small Interfering / pharmacology
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Receptors, Calcitriol / metabolism
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Retinoid X Receptor alpha / metabolism
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Signal Transduction / drug effects
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Steroid Hydroxylases / genetics
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Vitamin D3 24-Hydroxylase
Substances
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Benzoquinones
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Enzyme Inhibitors
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HSP90 Heat-Shock Proteins
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HSP90AB1 protein, human
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Lactams, Macrocyclic
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RNA, Messenger
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RNA, Small Interfering
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Receptors, Calcitriol
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Retinoid X Receptor alpha
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DNA
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Steroid Hydroxylases
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Vitamin D3 24-Hydroxylase
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Calcitriol
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geldanamycin