No abstract

In this study, we demonstrate that the pervasive xenobiotic methoxyacetic acid and the commonly prescribed anticonvulsant valproic acid, both short-chain fatty acids (SCFAs), dramatically increase cellular sensitivity to estrogens, progestins, and other nuclear hormone receptor ligands. These compounds do not mimic endogenous hormones but rather act to enhance the transcriptional efficacy of ligand activated nuclear hormone receptors by up to 8-fold in vitro and in vivo. Detailed characterization of their mode of action revealed that these SCFAs function as both activators of p42͞p44 mitogen-activated protein kinase and as inhibitors of histone deacetylases at doses that parallel known exposure levels. Our results define a class of compounds that possess a dual mechanism of action and function as hormone sensitizers. These findings prompt an evaluation of previously unrecognized drugdrug interactions in women who are administered exogenous hormones while exposed to certain xenobiotic SCFAs. Furthermore, our study highlights the need to structure future screening programs to identify additional hormone sensitizers.

No abstract

Carnitine palmitoyltransferase-I (CPT-I) catalyzes the rate-controlling step of fatty acid oxidation. CPT-I converts long-chain fatty acyl-CoAs to acylcarnitines for translocation across the mitochondrial membrane. The mRNA levels and enzyme activity of the liver isoform, CPT-I␣, are greatly increased in the liver of hyperthyroid animals. Thyroid hormone (T3) stimulates CPT-I␣ transcription far more robustly in the liver than in nonhepatic tissues. We have shown that the thyroid hormone receptor (TR) binds to a thyroid hormone response element (TRE) located in the CPT-I␣ promoter. In addition, elements in the first intron participate in the T3 induction of CPT-I␣ gene expression, but the CPT-I␣ intron alone cannot confer a T3 response. We found that deletion of sequences in the first intron between ؉653 and ؉744 decreased the T3 induction of CPT-I␣. Upstream stimulatory factor (USF) and CCAAT enhancer binding proteins (C/EBPs) bind to elements within this region, and these factors are required for the T3 response. The binding of TR and C/EBP to the CPT-I␣ gene in vivo was shown by the chromatin immunoprecipitation assay. We determined that TR can physically interact with USF-1, USF-2, and C/EBP␣. Transgenic mice were created that carry CPT-I␣-luciferase transgenes with or without the first intron of the CPT-I␣ gene. In these mouse lines, the first intron is required for T3 induction as well as high levels of hepatic expression. Our data indicate that the T3 stimulates CPT-I␣ gene expression in the liver through a T3 response unit consisting of the TRE in the promoter and additional factors, C/EBP and USF, bound in the first intron.

The peroxisome proliferator-activated receptors (PPARalpha, PPARdelta, and PPARgamma) constitute a family of nuclear receptors that regulates metabolic processes involved in lipid and glucose homeostasis. Although generally considered to function as ligand-regulated receptors, all three PPARs exhibit a high level of constitutive activity that may result from their stimulation by intracellularly produced endogenous ligands. Consequently, complete inhibition of PPAR signaling requires the development of inverse agonists. However, the currently available small molecule antagonists for the PPARs function only as partial agonists, or their efficacy is not sufficient to inhibit the constitutive activity of these receptors. Due to the lack of efficacious antagonists that interact with the ligand-binding domain of the PPARs, we decided to target an interaction that is central to nuclear receptor-mediated gene transcription: the nuclear receptor-coactivator interaction. We utilized phage display technology to identify short LXXLL-containing peptides that bind to the PPARs. Analysis of these peptides revealed a consensus binding motif consisting of HPLLXXLL. Cross-screening of these peptides for binding to other nuclear receptors enabled the identification of a high-affinity PPAR-selective peptide that has the ability to repress PPARgamma1-dependent transcription of transfected reporter genes. Most importantly, when introduced into HepG2 cells, the peptide inhibited the expression of endogenous PPARgamma1 target genes, adipose differentiation-related protein and mitochondrial 3-hydroxy-3-methylglutaryl coenzyme A synthase 2. This work lends support for the rational development of peptidomimetics that block receptor-mediated transcription by targeting the nuclear receptor-coactivator interaction surface.