Characterization of a novel PXR isoform with potential dominant-negative properties.
BACKGROUND & AIMS
The nuclear Pregnane X Receptor (PXR, NR1I2) plays a pivotal role in xenobiotic metabolism. Here, we sought to characterize a new PXR isoform (hereafter called small PXR or sPXR) stemming from alternative transcription starting sites downstream of a CpG Island located near exon 3 of the human PXR gene.


METHODS
Quantitative RT-PCR, western blot, methylation-specific PCR, luciferase reporter assays, electro-mobility shift assays, and stable sPXR overexpression were used to examine sPXR expression and function in hepatocellular cell lines, healthy human liver (n=99), hepatocellular adenomas (HCA, n=91) and hepatocellular carcinoma samples (HCC, n=213).


RESULTS
Liver sPXR mRNA expression varied importantly among individuals and encodes a 37kDa nuclear protein consisting of the ligand-binding domain of PXR that behaves as a dominant-negative of PXR transactivation properties. In vitro methylation of the sPXR upstream promoter abolished its activity, while the demethylation agent 5-aza-2-deoxycytidine increased sPXR mRNA expression in several cell lines. Finally, we observed that sPXR mRNA expression displayed significant differences related to HCA or HCC biology.


CONCLUSIONS
This novel PXR isoform, displaying a dominant-negative activity and regulated by DNA methylation, is associated with outcomes of patients with HCC treated by resection, suggesting that it represents a key modulator of PXR.
['Breuker|Cyril|C|', 'Planque|Chris|C|', 'Rajabi|Fatemeh|F|', 'Nault|Jean-Charles|JC|', 'Couchy|Gabrielle|G|', 'Zucman-Rossi|Jessica|J|', 'Evrard|Alexandre|A|', 'Kantar|Jovana|J|', 'Chevet|Eric|E|', 'Bioulac-Sage|Paulette|P|', 'Ramos|Jeanne|J|', 'Assenat|Eric|E|', 'Joubert|Dominique|D|', 'Pannequin|Julie|J|', 'Hollande|Frédéric|F|', 'Pascussi|Jean Marc|JM|']
D018248:Adenoma, Liver Cell D006528:Carcinoma, Hepatocellular D045744:Cell Line, Tumor D002478:Cells, Cultured D019175:DNA Methylation D006498:Hepatectomy D006801:Humans D066298:In Vitro Techniques D008099:Liver D008113:Liver Neoplasms D000077297:Pregnane X Receptor D020033:Protein Isoforms D012333:RNA, Messenger D011987:Receptors, Steroid D016896:Treatment Outcome
Drug metabolism Epigenetic regulation Hepatocyte Nuclear receptor