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Role of Nuclear Receptors and Their Splice Variants in Regulating Boar Taint Metabolism and Steroidogenesis

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Title: Role of Nuclear Receptors and Their Splice Variants in Regulating Boar Taint Metabolism and Steroidogenesis
Author: Gray, Matthew Alexander
Department: Department of Animal and Poultry Science
Program: Animal and Poultry Science
Advisor: Squires, E. James
Abstract: This thesis investigates how activation of the nuclear receptors pCAR, pPXR and pFXR and the presence of their alternatively spliced variants, pCAR-SV2, pPXR-SV1 and pFXR-SV1 affects metabolism of boar taint compounds (androstenone and skatole), steroidogenesis and gene expression in boars. Transactivation of CAR, PXR, or FXR in primary Leydig cells increased the expression of several genes involved in steroidogenesis, including cytochrome B5A (CYB5A) and cytochrome B5 reductase 1 (CYB5R1), as well as hydroxysteroid (17-beta) dehydrogenase 4 (HSD17B4) and retinol dehydrogenase 12 (RDH12). Treatment with CAR or PXR agonists significantly decreased sex steroid production, by approximately 40%, and significantly increased production of 16-androstene steroids by approximately 25%. Treatment with an FXR agonist increased sex steroid production by approximately 46%, but had no effect on 16-androstene steroid synthesis. In primary hepatocytes receptor transactivation resulted in altered expression of several transcripts, including increased expression of CYP2B22 by CAR, CYP2A19, CYP2B22, CYP2C33, and CYP2C49 by PXR, and CYP2C33 and CYP2E1 by FXR. Only transactivation of PXR had a significant effect on androstenone metabolism, while FXR had the greatest effect on skatole metabolism, increasing the production of the key metabolite 6-hydroxy-3-methylindole. The dominant negative effect of pCAR-SV2 was dependent on DNA binding, since mutation of the DNA binding domain abolished the effect pCAR-SV2 had on pCAR-WT; this effect was also found in vivo, with increased expression of pCAR-SV2 correlated with decreased expression of several target genes. The dominant positive effect of pPXR-SV1 on pPXR-WT was dependent on the charged clamp region, which is involved in co-regulatory protein binding, with K246 being the key residue in this region. The pFXR-SV1 dominant positive effect was dependent on the tyrosine residue of the MYTG insert, and the phosphorylation status of this residue may be important for pFXR-SV1 to exert its effect. The dominant positive effects of pPXR-SV1 and pFXR-SV1 were not shown to exist in vivo, though increased pFXR expression did increase expression of CYP2C49 and CYP2C33, and decrease expression of CYP2E1. In summary, transactivation of pCAR, pPXR, and pFXR alters boar taint formation and metabolism, and the dominant negative effect of pCAR-SV2 may be able to affect pCAR transactivation in vivo.
Date: 2013-11
Rights: Attribution-NonCommercial-NoDerivs 2.5 Canada
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Attribution-NonCommercial-NoDerivs 2.5 Canada Except where otherwise noted, this item's license is described as Attribution-NonCommercial-NoDerivs 2.5 Canada