title: Epigenetic regulation of microRNA genes and the role of miR-34b in cell invasion and motility in human melanoma.
abstract: Invasive melanoma is the most lethal form of skin cancer. The treatment of melanoma-derived cell lines with 5-aza-2'-deoxycytidine (5-Aza-dC) markedly increases the expression of several miRNAs, suggesting that the miRNA-encoding genes might be epigenetically regulated, either directly or indirectly, by DNA methylation. We have identified a group of epigenetically regulated miRNA genes in melanoma cells, and have confirmed that the upstream CpG island sequences of several such miRNA genes are hypermethylated in cell lines derived from different stages of melanoma, but not in melanocytes and keratinocytes. We used direct DNA bisulfite and immunoprecipitated DNA (Methyl-DIP) to identify changes in CpG island methylation in distinct melanoma patient samples classified as primary in situ, regional metastatic, and distant metastatic. Two melanoma cell lines (WM1552C and A375 derived from stage 3 and stage 4 human melanoma, respectively) were engineered to ectopically express one of the epigenetically modified miRNA: miR-34b. Expression of miR-34b reduced cell invasion and motility rates of both WM1552C and A375, suggesting that the enhanced cell invasiveness and motility observed in metastatic melanoma cells may be related to their reduced expression of miR-34b. Total RNA isolated from control or miR-34b-expressing WM1552C cells was subjected to deep sequencing to identify gene networks around miR-34b. We identified network modules that are potentially regulated by miR-34b, and which suggest a mechanism for the role of miR-34b in regulating normal cell motility and cytokinesis.
authors.affiliation: Sanford Burnham Medical Research Institute, Orlando, Florida, United States of America.
authors.forename: Joseph
authors.initials: J
authors.lastname: Mazar
authors.forename: Divya
authors.initials: D
authors.lastname: Khaitan
authors.forename: Dan
authors.initials: D
authors.lastname: DeBlasio
authors.forename: Cuncong
authors.initials: C
authors.lastname: Zhong
authors.forename: Subramaniam S
authors.initials: SS
authors.lastname: Govindarajan
authors.forename: Sharmila
authors.initials: S
authors.lastname: Kopanathi
authors.forename: Shaojie
authors.initials: S
authors.lastname: Zhang
authors.forename: Animesh
authors.initials: A
authors.lastname: Ray
authors.forename: Ranjan J
authors.initials: RJ
authors.lastname: Perera
mesh_terms: D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism; D015152:Blotting, Northern; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D002465:Cell Movement; D018899:CpG Islands; D019175:DNA Methylation; D057890:Epigenomics; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D006801:Humans; D008545:Melanoma / Q000235:genetics* / Q000556:secondary*; D035683:MicroRNAs / Q000235:genetics*; D009361:Neoplasm Invasiveness; D020411:Oligonucleotide Array Sequence Analysis; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012878:Skin Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D014945:Wound Healing
chemical_list: D014408:Biomarkers, Tumor; C522425:MIRN34 microRNA, human; D035683:MicroRNAs; D012333:RNA, Messenger