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Endogenous Retinoic Acid Receptor (RAR)-Retinoid X Receptor (RXR) Heterodimers Are the Major Functional Forms Regulating Retinoid-responsive Elements in Adult Human Keratinocytes

1995; Elsevier BV; Volume: 270; Issue: 7 Linguagem: Inglês

10.1074/jbc.270.7.3001

1083-351X

Jia‐Hao Xiao, Béatrice Durand, Pierre Chambon, John J. Voorhees,

Antioxidant Activity and Oxidative Stress

We have examined how retinoic acid receptors (RARs) and retinoid X receptors (RXRs) at physiological concentrations regulate distinct retinoid-responsive elements, hRARβ2/βRARE (DR5) and rCRBPII/RXRE (DR1), in keratinocytes from human skin, a major retinoid target. In vitro, endogenous RAR γ and RXRs bound to these elements as heterodimers (RAR•RXR) but not homodimers (RAR•RAR or RXR•RXR). In cultured keratinocytes, all-trans retinoic acid, 9-cis retinoic acid, and CD367 activated βRARE but not RXRE via endogenous RAR•RXR (ED50 = 2.3, 3.8, and 0.3 nM, respectively) whereas SR11237 showed no significant effect. All-trans retinoic acid, 9-cis retinoic acid, and SR11237 activated RXRE via overexpressed RXR•RXR (ED50 = 110, 120, and 11 nM, respectively), indicating interconversion between retinoic acid isomers, whereas co-overexpression of RARα or RAR γ suppressed this activation. Unlike 9cRA, CD367 neither induced formation of nor activated RXR•RXR. Overexpression of RAR or RXR mutated in transactivation domain AF-2 suppressed endogenous receptor activity over βRARE. Our data suggest that 1) in keratinocytes, RAR•RXR-mediated pathway dominates over that mediated by RXR•RXR; 2) RAR-selective CD367 and RXR-selective SR11237 can be used to identify these two distinct pathways, respectively; 3) βRARE is mainly regulated by RAR•RXR, in which RAR alone confers ligand inducibility whereas AF-2 of unliganded RXR is required for transactivation by liganded RAR AF-2; 4) lack of RXRE activity in keratinocytes is due to low endogenous levels of RXR•RXR and inhibition by RAR•RXR; and 5) interaction among RXRs is much lower than that between RAR and RXR. We have examined how retinoic acid receptors (RARs) and retinoid X receptors (RXRs) at physiological concentrations regulate distinct retinoid-responsive elements, hRARβ2/βRARE (DR5) and rCRBPII/RXRE (DR1), in keratinocytes from human skin, a major retinoid target. In vitro, endogenous RAR γ and RXRs bound to these elements as heterodimers (RAR•RXR) but not homodimers (RAR•RAR or RXR•RXR). In cultured keratinocytes, all-trans retinoic acid, 9-cis retinoic acid, and CD367 activated βRARE but not RXRE via endogenous RAR•RXR (ED50 = 2.3, 3.8, and 0.3 nM, respectively) whereas SR11237 showed no significant effect. All-trans retinoic acid, 9-cis retinoic acid, and SR11237 activated RXRE via overexpressed RXR•RXR (ED50 = 110, 120, and 11 nM, respectively), indicating interconversion between retinoic acid isomers, whereas co-overexpression of RARα or RAR γ suppressed this activation. Unlike 9cRA, CD367 neither induced formation of nor activated RXR•RXR. Overexpression of RAR or RXR mutated in transactivation domain AF-2 suppressed endogenous receptor activity over βRARE. Our data suggest that 1) in keratinocytes, RAR•RXR-mediated pathway dominates over that mediated by RXR•RXR; 2) RAR-selective CD367 and RXR-selective SR11237 can be used to identify these two distinct pathways, respectively; 3) βRARE is mainly regulated by RAR•RXR, in which RAR alone confers ligand inducibility whereas AF-2 of unliganded RXR is required for transactivation by liganded RAR AF-2; 4) lack of RXRE activity in keratinocytes is due to low endogenous levels of RXR•RXR and inhibition by RAR•RXR; and 5) interaction among RXRs is much lower than that between RAR and RXR.