title: Epigenetic regulation of p14ARF and p16INK4A expression in cutaneous and uveal melanoma.
abstract: Inactivation of p14ARF and p16INK4A by epigenetic changes in cutaneous and uveal melanoma has been here investigated. Compared with melanocytes, p14ARF mRNA reduction and p16INK4A inactivation were frequently noticed. No association between p14ARF promoter methylation and mRNA levels was found, whereas aberrant p16INK4A methylation was associated with gene silencing (p<0.001). Comparative analysis within melanomas of different Breslow's thicknesses showed that drastic reductions in p14ARF and p16INK4A expression appeared at the level of thin/intermediate and intermediate/thick transitions. The effects of 5-aza-2'-deoxycytidine (5-aza-dC) and suberanilohydroxamic acid (SAHA) on in vivo binding of DNA methyltransferases (DNMTs) and acetyl histone H3/H4 to p14ARF and p16INK4A promoters were tested together with the impact of ectopic expression of p14ARF and p16INK4A on cell proliferation, migration, and invasion. SAHA treatment induced H3 and H4 hyperacetylation at the p14ARF promoter followed by increased p14ARF expression, whereas exposure to 5-aza-dC decreased the recruitment of DNMT1 and DNMT3b at the p16INK4A promoter and reactivated p16INK4A. Studies on promoter-associated di-methyl histone H3 (Lys4) levels ruled out an involvement of this epigenetic trait on p14ARF and p16INK4A expression. The enforced expression of p14ARF or p16INK4A and, even more so, their co-expression, significantly reduced cell proliferation, migration and invasion. Our data pinpoint: i) a frequent impairment of p14ARF and p16INK4A gene expression by epigenetic modifications in melanoma; ii) histone hypoacetylation as the dominant mechanism of p14ARF silencing; and iii) 5' CpG promoter methylation as the major mechanism of p16INK4A gene inactivation. Collectively, our data suggest that selected epi-drugs may be useful in melanoma treatment.
authors.affiliation: Department of Experimental, Specialized Medical-Surgical and Odontostomatological Sciences, University of Messina, Messina, Italy.
authors.forename: Mario
authors.initials: M
authors.lastname: Venza
authors.affiliation: Department of Clinical and Experimental Medicine, University of Messina, Messina, Italy.
authors.forename: Maria
authors.initials: M
authors.lastname: Visalli
authors.affiliation: Department of Pediatric, Gynecological, Microbiological and Biomedical Sciences, University of Messina, Messina, Italy.
authors.forename: Carmelo
authors.initials: C
authors.lastname: Biondo
authors.affiliation: Department of Human Pathology, University of Messina, Messina, Italy.
authors.forename: Maria
authors.initials: M
authors.lastname: Lentini
authors.affiliation: Department of Clinical and Experimental Medicine, University of Messina, Messina, Italy.
authors.forename: Teresa
authors.initials: T
authors.lastname: Catalano
authors.affiliation: Department of Clinical and Experimental Medicine, University of Messina, Messina, Italy. Electronic address: dteti@unime.it.
authors.forename: Diana
authors.initials: D
authors.lastname: Teti
authors.affiliation: Department of Experimental, Specialized Medical-Surgical and Odontostomatological Sciences, University of Messina, Messina, Italy.
authors.forename: Isabella
authors.initials: I
authors.lastname: Venza
mesh_terms: D000328:Adult; D001374:Azacitidine / Q000008:administration & dosage / Q000031:analogs & derivatives; D045744:Cell Line, Tumor; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000096:biosynthesis*; D019175:DNA Methylation / Q000235:genetics; D000077209:Decitabine; D044127:Epigenesis, Genetic; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D020868:Gene Silencing; D006655:Histone Deacetylases / Q000235:genetics; D006801:Humans; D008545:Melanoma / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D011401:Promoter Regions, Genetic; D025803:Tumor Suppressor Protein p14ARF / Q000096:biosynthesis*; D014604:Uveal Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D000098943:Uveal Melanoma
chemical_list: D019941:Cyclin-Dependent Kinase Inhibitor p16; D025803:Tumor Suppressor Protein p14ARF; D000077209:Decitabine; D006655:Histone Deacetylases; D001374:Azacitidine
keywords: Epi-drug; Histone deacetylation; Human melanoma; Promoter methylation; p14(ARF); p16(INK4A)