title: DNA methylation contributes toward silencing of antioncogenic microRNA-203 in human and canine melanoma cells.
abstract: Melanoma is a poor-prognosis cancer in both humans and dogs. We have elucidated the antitumor mechanisms of antioncogenic microRNA (miR)-203 which is downregulated in human melanoma, as well as in canine melanoma. The aim of this study was to clarify the mechanism of this downregulation. We focused on epigenetic aberration of miR-203 transcription. Treatment with 5-aza-2'-deoxycitidine (5-aza) markedly upregulated the expression level of miR-203 in almost all of the cell lines tested. Furthermore, bisulfite sequencing or methylation-specific PCR showed DNA methylation of CpG islands upstream of the miR-203 coding region (MIR203) in both human and canine melanoma cells, as well as in canine clinical specimens, but not in human normal melanocytes. The results of a luciferase activity assay showed obvious suppression of the transcription of miR-203 by DNA methylation. The use of the luciferase activity assay for CREB1 and an inhibition assay of miR-203 function performed with an miR-203 inhibitor confirmed the contribution of miR-203 upregulation toward the negative regulation of the target gene of miR-203. These results indicate that canine melanoma might be a preclinical model of human melanoma for epigenetic studies. In addition, this study suggests that agents that can demethylate MIR203 could be a common promising therapeutic agent for the treatment of human and canine melanomas.
authors.affiliation: aLaboratory of Molecular Diagnostics and Therapeutics bLaboratory of Small Animal Clinical Science (Surgical Division), Joint Faculty of Veterinary Medicine cBiomedical Science Center for Translational Research, The United Graduate School of Veterinary Science, Yamaguchi University, Yamaguchi dLaboratory of Veterinary Clinical Oncology, Faculty of Applied Biological Sciences, Gifu University, Gifu eLaboratory of Veterinary Surgery, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo, Japan.
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Noguchi
authors.forename: Takashi
authors.initials: T
authors.lastname: Mori
authors.forename: Takayuki
authors.initials: T
authors.lastname: Nakagawa
authors.forename: Kazuhito
authors.initials: K
authors.lastname: Itamoto
authors.forename: Tomoya
authors.initials: T
authors.lastname: Haraguchi
authors.forename: Takuya
authors.initials: T
authors.lastname: Mizuno
mesh_terms: D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D019175:DNA Methylation / Q000187:drug effects / Q000502:physiology*; D004283:Dog Diseases / Q000235:genetics / Q000473:pathology; D004285:Dogs; D044127:Epigenesis, Genetic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D020868:Gene Silencing / Q000187:drug effects; D016147:Genes, Tumor Suppressor / Q000187:drug effects; D006801:Humans; D008544:Melanocytes / Q000187:drug effects / Q000378:metabolism; D008545:Melanoma / Q000235:genetics* / Q000473:pathology / Q000662:veterinary; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D009062:Mouth Neoplasms / Q000235:genetics / Q000473:pathology / Q000662:veterinary
chemical_list: C546703:MIRN203 microRNA, human; D035683:MicroRNAs; D001374:Azacitidine