title: The "melanoma-enriched" microRNA miR-4731-5p acts as a tumour suppressor.
abstract: We previously identified miR-4731-5p (miR-4731) as a melanoma-enriched microRNA following comparison of melanoma with other cell lines from solid malignancies. Additionally, miR-4731 has been found in serum from melanoma patients and expressed less abundantly in metastatic melanoma tissues from stage IV patients relative to stage III patients. As miR-4731 has no known function, we used biotin-labelled miRNA duplex pull-down to identify binding targets of miR-4731 in three melanoma cell lines (HT144, MM96L and MM253). Using the miRanda miRNA binding algorithm, all pulled-down transcripts common to the three cell lines (n=1092) had potential to be targets of miR-4731 and gene-set enrichment analysis of these (via STRING v9.1) highlighted significantly associated genes related to the 'cell cycle' pathway and the 'melanosome'. Following miR-4731 overexpression, a selection (n=81) of pull-down transcripts underwent validation using a custom qRT-PCR array. These data revealed that miR-4731 regulates multiple genes associated with the cell cycle (e.g. CCNA2, ORC5L, and PCNA) and the melanosome (e.g. RAB7A, CTSD, and GNA13). Furthermore, members of the synovial sarcoma X breakpoint family (SSX) (melanoma growth promoters) were also down-regulated (e.g. SSX2, SSX4, and SSX4B) as a result of miR-4731 overexpression. Moreover, this down-regulation of mRNA expression resulted in ablation or reduction of SSX4 protein, which, in keeping with previous studies, resulted in loss of 2D colony formation. We therefore speculate that loss of miR-4731 expression in stage IV patient tumours supports melanoma growth by, in part; reducing its regulatory control of SSX expression levels.
authors.affiliation: Dermatology Research Centre, The University of Queensland, School of Medicine, Translational Research Institute, Brisbane, QLD, Australia.|QIMR Berghofer Medical Research Institute, Herston, Brisbane, QLD, Australia.
authors.forename: Mitchell S
authors.initials: MS
authors.lastname: Stark
authors.affiliation: Dermatology Research Centre, The University of Queensland, School of Medicine, Translational Research Institute, Brisbane, QLD, Australia.
authors.forename: Lisa N
authors.initials: LN
authors.lastname: Tom
authors.affiliation: QIMR Berghofer Medical Research Institute, Herston, Brisbane, QLD, Australia.
authors.forename: Glen M
authors.initials: GM
authors.lastname: Boyle
authors.affiliation: School of Biomedical Sciences, Institute of Health and Biomedical Innovation, Queensland University of Technology, at The Translational Research Institute, Brisbane, QLD, Australia.
authors.forename: Vanessa F
authors.initials: VF
authors.lastname: Bonazzi
authors.affiliation: Dermatology Research Centre, The University of Queensland, School of Medicine, Translational Research Institute, Brisbane, QLD, Australia.
authors.forename: H Peter
authors.initials: HP
authors.lastname: Soyer
authors.affiliation: School of Biomedical Sciences, Institute of Health and Biomedical Innovation, Queensland University of Technology, at The Translational Research Institute, Brisbane, QLD, Australia.
authors.forename: Adrian C
authors.initials: AC
authors.lastname: Herington
authors.affiliation: School of Biomedical Sciences, Institute of Health and Biomedical Innovation, Queensland University of Technology, at The Translational Research Institute, Brisbane, QLD, Australia.
authors.forename: Pamela M
authors.initials: PM
authors.lastname: Pollock
authors.affiliation: QIMR Berghofer Medical Research Institute, Herston, Brisbane, QLD, Australia.
authors.forename: Nicholas K
authors.initials: NK
authors.lastname: Hayward
mesh_terms: D000465:Algorithms; D001710:Biotin / Q000737:chemistry; D002453:Cell Cycle; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D015972:Gene Expression Regulation, Neoplastic; D016147:Genes, Tumor Suppressor; D006801:Humans; D008545:Melanoma / Q000378:metabolism*; D035683:MicroRNAs / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D012097:Repressor Proteins / Q000378:metabolism; D015398:Signal Transduction; D012878:Skin Neoplasms / Q000378:metabolism*
chemical_list: D035683:MicroRNAs; D009363:Neoplasm Proteins; D012097:Repressor Proteins; C000626296:microRNA 4731, human; C093962:synovial sarcoma X breakpoint proteins; D001710:Biotin
keywords: PMP22; SSX; melanoma; miR-4731; microRNA