title: RANKL expression in myeloma cells is regulated by a network involving RANKL promoter methylation, DNMT1, microRNA and TNFα in the microenvironment.
abstract: We studied the regulation of RANKL expression in myeloma by promoter DNA methylation. Methylation-specific polymerase chain reaction showed complete methylation of RANKL promoter in WL-2 myeloma cells but partial methylation in eight other lines. 5-AzadC treatment of WL-2 cells led to demethylation and re-expression of RANKL. Transwell and contact co-culture of WL-2 cells with normal bone marrow-derived mesenchymal stromal cells (BMSCs) resulted in comparable repression of DNA methyltransferase-1 (DNMT1) and re-expression of RANKL in WL-2 cells. Moreover, treatment of WL-2 cells with TNFα led to repression of DNMT1 and re-expression of RANKL in association with upregulation of miR-140-3p and miR-126, which are partially offset by addition of anti-TNFα antibody to transwell-coculture of WL2 with BMSC. Taken together, our results showed that TNFα in the marrow microenvironment led to RANKL demethylation and re-expression in myeloma cells through DNMT1 repression and upregulation of miR-126-3p and miR-140, both known to repress DNMT1 translation.
authors.affiliation: Department of Medicine, Queen Mary Hospital, University of Hong Kong, Hong Kong, China; Department of Paediatrics and Adolescent Medicine, Faculty of Medicine, The University of Hong Kong, Hong Kong, China.
authors.forename: Lingqing
authors.initials: L
authors.lastname: Yuan
authors.affiliation: Department of Medicine, Queen Mary Hospital, University of Hong Kong, Hong Kong, China; Department of Paediatrics and Adolescent Medicine, Faculty of Medicine, The University of Hong Kong, Hong Kong, China.
authors.forename: Godfrey Chi Fung
authors.initials: GC
authors.lastname: Chan
authors.affiliation: Department of Medicine, Queen Mary Hospital, University of Hong Kong, Hong Kong, China; Department of Paediatrics and Adolescent Medicine, Faculty of Medicine, The University of Hong Kong, Hong Kong, China.
authors.forename: Kwong Lam
authors.initials: KL
authors.lastname: Fung
authors.affiliation: Department of Medicine, Queen Mary Hospital, University of Hong Kong, Hong Kong, China; Department of Paediatrics and Adolescent Medicine, Faculty of Medicine, The University of Hong Kong, Hong Kong, China. Electronic address: jcschim@hku.hk.
authors.forename: Chor Sang
authors.initials: CS
authors.lastname: Chim
mesh_terms: D018920:Coculture Techniques; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000378:metabolism*; D019175:DNA Methylation / Q000235:genetics*; D015972:Gene Expression Regulation, Neoplastic; D053263:Gene Regulatory Networks; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000378:metabolism / Q000473:pathology; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D009101:Multiple Myeloma / Q000235:genetics*; D011401:Promoter Regions, Genetic; D053245:RANK Ligand / Q000235:genetics* / Q000378:metabolism; D059016:Tumor Microenvironment / Q000235:genetics*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: C527111:MIRN126 microRNA, human; D035683:MicroRNAs; C544302:Mirn140 microRNA, human; D053245:RANK Ligand; D014409:Tumor Necrosis Factor-alpha; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; C000619380:DNMT1 protein, human
keywords: DNA methylation; DNMT1; MicroRNA; Myeloma microenvironment; RANKL; TNFα