title: Expression of a novel mRNA transcript for human microsomal epoxide hydrolase (EPHX1) is regulated by short open reading frames within its 5'-untranslated region.
abstract: Microsomal epoxide hydrolase (mEH, EPHX1) is a critical xenobiotic-metabolizing enzyme, catalyzing both detoxification and bioactivation reactions that direct the disposition of chemical epoxides, including the carcinogenic metabolites of several polycyclic aromatic hydrocarbons. Recently, we discovered that a previously unrecognized and primate-specific EPHX1 transcript, termed E1-b, was actually the predominant driver of EPHX1 expression in all human tissues. In this study, we identify another human EPHX1 transcript, designated as E1-b'. Unusually, both the E1-b and E1-b' mRNA transcripts are generated from the use of a far upstream gene promoter, localized ∼18.5 kb 5'-upstream of the EPHX1 protein-coding region. Although expressed at comparatively lower levels than E1-b, the novel E1-b' transcript is readily detected in all tissues examined, with highest levels maintained in human ovary. The E1-b' mRNA possesses unusual functional features in its 5'-untranslated region, including a GC-rich leader sequence and two upstream AUGs that encode for short peptides of 26 and 17 amino acids in length, respectively. Results from in vitro transcription/translation assays and direct transfection in mammalian cells of either the E1-b' transcript or the encoded peptides demonstrated that the E1-b' upstream open reading frames (uORFs) are functional, with their presence markedly inhibiting the translation of EPHX1 protein, both in cis and in trans configurations. These unique uORF peptides exhibit no homology to any other known uORF sequences but likely function to mediate post-transcription regulation of EPHX1 and perhaps more broadly as translational regulators in human cells.
authors.affiliation: Center for Molecular Toxicology and Carcinogenesis, Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, Pennsylvania 16802, USA.
authors.forename: Hong Loan
authors.initials: HL
authors.lastname: Nguyen
authors.forename: Xi
authors.initials: X
authors.lastname: Yang
authors.forename: Curtis J
authors.initials: CJ
authors.lastname: Omiecinski
mesh_terms: D020121:5' Untranslated Regions; D000595:Amino Acid Sequence; D001483:Base Sequence; D004789:Enzyme Activation; D004851:Epoxide Hydrolases / Q000235:genetics / Q000378:metabolism*; D005091:Exons; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D056945:Hep G2 Cells; D006801:Humans; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D016366:Open Reading Frames; D010053:Ovary / Q000166:cytology / Q000378:metabolism; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D011487:Protein Conformation; D059748:Proteolysis; D012323:RNA Processing, Post-Transcriptional; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D017385:Sequence Homology; D013997:Time Factors; D014158:Transcription, Genetic; D014162:Transfection; D014908:Triticum / Q000378:metabolism
chemical_list: D020121:5' Untranslated Regions; D010455:Peptides; D012333:RNA, Messenger; D004851:Epoxide Hydrolases; C522330:EPHX1 protein, human
keywords: 5′-untranslated region (5′ UTR); EPHX1; gene expression; microsomal epoxide hydrolase; translational regulation; upstream open reading frame (uORF)
references.citation: Mol Cell Biol. 2002 Dec;22(24):8562-70
references.pmid: 12446775
references.citation: BMC Genomics. 2006 Jan 26;7:16
references.pmid: 16438715
references.citation: Biol Cell. 2004 Sep;96(7):479-98
references.pmid: 15380615
references.citation: Cancer Res. 2000 Apr 1;60(7):1849-56
references.pmid: 10766171
references.citation: Toxicol Sci. 2005 May;85(1):624-31
references.pmid: 15716486
references.citation: Mol Cell Biol. 1989 Nov;9(11):5134-42
references.pmid: 2601712
references.citation: Prog Lipid Res. 2005 Jan;44(1):1-51
references.pmid: 15748653
references.citation: Proc Natl Acad Sci U S A. 1994 Jun 7;91(12):5612-6
references.pmid: 7515506
references.citation: Biochem Pharmacol. 1998 Apr 1;55(7):1059-69
references.pmid: 9605429
references.citation: Eur J Biochem. 1982 Aug;126(2):425-31
references.pmid: 7128597
references.citation: Arch Biochem Biophys. 1997 Jan 15;337(2):275-83
references.pmid: 9016823
references.citation: Mol Pharmacol. 2005 Jan;67(1):220-30
references.pmid: 15465926
references.citation: J Biol Chem. 2005 Jun 24;280(25):24252-60
references.pmid: 15851478
references.citation: Biochem Pharmacol. 1998 Jan 15;55(2):169-75
references.pmid: 9448739
references.citation: J Pharmacol Exp Ther. 1994 Apr;269(1):417-23
references.pmid: 7513361
references.citation: Histochem J. 2001 Jun;33(6):329-36
references.pmid: 11758809
references.citation: Cancer Res. 1997 Aug 15;57(16):3562-8
references.pmid: 9270029
references.citation: Chem Biol Interact. 2008 Jun 17;173(3):179-86
references.pmid: 18468591
references.citation: Endocrinology. 2000 Sep;141(9):3353-65
references.pmid: 10965908
references.citation: Gene. 2005 Nov 21;361:13-37
references.pmid: 16213112
references.citation: Oncogene. 1999 Oct 7;18(41):5631-7
references.pmid: 10523842
references.citation: J Pharmacol Exp Ther. 2009 Jul;330(1):23-30
references.pmid: 19364907
references.citation: Mol Cell Biol. 1995 Oct;15(10):5235-45
references.pmid: 7565672
references.citation: Cancer Res. 1996 Dec 1;56(23):5410-6
references.pmid: 8968094
references.citation: Mol Cell Biol. 1997 Sep;17(9):4904-13
references.pmid: 9271370
references.citation: Toxicol Sci. 2007 Jun;97(2):384-97
references.pmid: 17329237
references.citation: Trends Genet. 2001 Dec;17(12):685-7
references.pmid: 11718907
references.citation: Microbiol Rev. 1996 Jun;60(2):366-85
references.pmid: 8801438
references.citation: Nucleic Acids Res. 2005 Sep 26;33(17):5512-20
references.pmid: 16186132
references.citation: Toxicol Appl Pharmacol. 2007 Jul 1;222(1):42-56
references.pmid: 17512962
references.citation: J Biol Chem. 2003 Jul 11;278(28):25716-21
references.pmid: 12730202
references.citation: J Biol Chem. 1999 Aug 20;274(34):23963-8
references.pmid: 10446164
references.citation: Chem Biol Interact. 2000 Dec 1;129(1-2):41-59
references.pmid: 11154734
references.citation: Mol Cell Biol. 2000 Dec;20(23):8635-42
references.pmid: 11073965
references.citation: Carcinogenesis. 1998 Mar;19(3):387-93
references.pmid: 9525271
references.citation: Annu Rev Microbiol. 2009;63:385-409
references.pmid: 19514854
references.citation: Biotechniques. 2006 Jul;41(1):35-6, 38, 40 passim
references.pmid: 16869511
references.citation: Cancer Res. 2000 Feb 1;60(3):517-21
references.pmid: 10676627
references.citation: J Biol Chem. 1994 Aug 12;269(32):20668-73
references.pmid: 8051167
references.citation: Genes Dev. 2006 Apr 15;20(8):915-21
references.pmid: 16618802
references.citation: PLoS One. 2011 Mar 18;6(3):e14749
references.pmid: 21445251