title: Transcriptome association analysis identifies miR-375 as a major determinant of variable acetaminophen glucuronidation by human liver.
abstract: Acetaminophen is the leading cause of acute liver failure (ALF) in many countries including the United States. Hepatic glucuronidation by UDP-glucuronosyltransferase (UGT) 1A subfamily enzymes is the major route of acetaminophen elimination. Reduced glucuronidation may predispose some individuals to acetaminophen-induced ALF, but mechanisms underlying reduced glucuronidation are poorly understood. We hypothesized that specific microRNAs (miRNAs) may reduce UGT1A activity by direct effects on the UGT1A 3'-UTR shared by all UGT1A enzyme transcripts, or by indirect effects on transcription factors regulating UGT1A expression. We performed an unbiased miRNA whole transcriptome association analysis using a bank of human livers with known acetaminophen glucuronidation activities. Of 754 miRNAs evaluated, 9 miRNAs were identified that were significantly overexpressed (p<0.05; >2-fold) in livers with low acetaminophen glucuronidation activities compared with those with high activities. miR-375 showed the highest difference (>10-fold), and was chosen for further mechanistic validation. We demonstrated using in silico analysis and luciferase reporter assays that miR-375 has a unique functional binding site in the 3'-UTR of the aryl hydrocarbon receptor (AhR) gene. Furthermore overexpression of miR-375 in LS180 cells demonstrated significant repression of endogenous AhR protein (by 40%) and mRNA (by 10%), as well as enzyme activity and/or mRNA of AhR regulated enzymes including UGT1A1, UGT1A6, and CYP1A2, without affecting UGT2B7, which is not regulated by AhR. Thus miR-375 is identified as a novel repressor of UGT1A-mediated hepatic acetaminophen glucuronidation through reduced AhR expression, which could predispose some individuals to increased risk for acetaminophen-induced ALF.
authors.affiliation: Program in Individualized Medicine (PrIMe), Department of Veterinary Clinical Sciences, College of Veterinary Medicine, Washington State University, Pullman, WA, USA. Electronic address: johnp@vetmed.wsu.edu.
authors.forename: Ioannis
authors.initials: I
authors.lastname: Papageorgiou
authors.affiliation: Sackler School of Graduate Biomedical Sciences, Tufts University, Boston, MA, USA. Electronic address: Marina.Freytsis@tufts.edu.
authors.forename: Marina
authors.initials: M
authors.lastname: Freytsis
authors.affiliation: Program in Individualized Medicine (PrIMe), Department of Veterinary Clinical Sciences, College of Veterinary Medicine, Washington State University, Pullman, WA, USA. Electronic address: michael.court@vetmed.wsu.edu.
authors.forename: Michael H
authors.initials: MH
authors.lastname: Court
mesh_terms: D020413:3' Untranslated Regions / Q000187:drug effects; D000082:Acetaminophen / Q000009:adverse effects / Q000378:metabolism*; D018712:Analgesics, Non-Narcotic / Q000009:adverse effects / Q000378:metabolism*; D051792:Basic Helix-Loop-Helix Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D002460:Cell Line; D056486:Chemical and Drug Induced Liver Injury / Q000201:enzymology / Q000378:metabolism; D019295:Computational Biology; D005103:Expert Systems; D005260:Female; D020869:Gene Expression Profiling; D017930:Genes, Reporter / Q000187:drug effects; D020719:Glucuronides / Q000378:metabolism; D014453:Glucuronosyltransferase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008099:Liver / Q000201:enzymology / Q000378:metabolism*; D008297:Male; D050217:Metabolic Detoxication, Phase II; D035683:MicroRNAs / Q000737:chemistry / Q000378:metabolism*; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D018336:Receptors, Aryl Hydrocarbon / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D014015:Tissue Banks; D000099148:UGT1A1 Enzyme
chemical_list: D020413:3' Untranslated Regions; D000082:Acetaminophen; D018712:Analgesics, Non-Narcotic; D051792:Basic Helix-Loop-Helix Proteins; D020719:Glucuronides; D014453:Glucuronosyltransferase; D035683:MicroRNAs; D012313:RNA; D012333:RNA, Messenger; D018336:Receptors, Aryl Hydrocarbon; D000099148:UGT1A1 Enzyme; C111235:AHR protein, human; C540933:MIRN375 microRNA, human; C039919:RNA, recombinant
keywords: Acetaminophen; Acetaminophen-induced hepatotoxicity; Aryl hydrocarbon receptor; UDP-glucuronosyltransferase 1A; miRNA; miRNA transcriptome association analysis