title: Regulation of miRNA-21 by reactive oxygen species-activated ERK/NF-κB in arsenite-induced cell transformation.
abstract: After acute exposure of cells to arsenic, reactive oxygen species mediate changes in cell behavior, including activation of proliferative signaling. For chronic exposure to arsenic, however, the function of reactive oxygen species in cell transformation remains poorly understood. Although microRNA-21 (miR-21) has been implicated in various aspects of carcinogenesis, its functions and molecular mechanisms in carcinogen-induced tumorigenesis are unclear. The purpose of this study was to determine if miR-21 is involved in arsenite-induced malignant transformation and to characterize the associated signaling pathways. During arsenite-induced transformation of human embryo lung fibroblast (HELF) cells, miR-21 was upregulated, and the extracellular signal-regulated kinase (ERK)/nuclear factor-κB (NF-κB) signal pathway was activated. Moreover, superoxide radical dismutase (a scavenger of superoxide) and catalase (a scavenger of hydroperoxides) blocked the arsenite-induced effects in HELF cells and mouse embryonic fibroblasts. Blockage of ERK by the inhibitor U0126 or inhibition of NF-κB p65 by siRNA or Bay 11-7082 prevented the increases in miR-21 and the decreases in Spry1, Pten, and Pdcd4, the target proteins of miR-21, induced by arsenite. As determined by a ChIP-qPCR assay, NF-κB p65 regulated miR-21 expression by binding directly to the promoter of miR-21. Further, anti-miR-21 downregulated miR-21 expression and prevented the arsenite-induced activation of ERK via the increase in Spry1, indicating that miR-21 has a feedback effect in regulating ERK activation. Overexpression of miR-21 with an miR-21 mimic and feedback activation of ERK and NF-κB via the decrease in Spry1 promoted the malignancy of HELF cells exposed to arsenite, but knockdown of miR-21 with anti-miR-21 and feedback blockage of ERK and NF-κB activation through an increase in Spry1 decreased anchorage-independent growth of arsenite-transformed cells. Thus, the transformation of HELF cells induced by chronic exposure to arsenite is mediated by increased miR-21 expression, which, in turn, is mediated by reactive oxygen species activation of the ERK/NF-κB pathway.
authors.affiliation: Institute of Toxicology, Nanjing Medical University, Nanjing 210029, Jiangsu, People's Republic of China.
authors.forename: Min
authors.initials: M
authors.lastname: Ling
authors.forename: Yuan
authors.initials: Y
authors.lastname: Li
authors.forename: Yuan
authors.initials: Y
authors.lastname: Xu
authors.forename: Ying
authors.initials: Y
authors.lastname: Pang
authors.forename: Lu
authors.initials: L
authors.lastname: Shen
authors.forename: Rongrong
authors.initials: R
authors.lastname: Jiang
authors.forename: Yue
authors.initials: Y
authors.lastname: Zhao
authors.forename: Xiaojun
authors.initials: X
authors.lastname: Yang
authors.forename: Jianping
authors.initials: J
authors.lastname: Zhang
authors.forename: Jianwei
authors.initials: J
authors.lastname: Zhou
authors.forename: Xinru
authors.initials: X
authors.lastname: Wang
authors.forename: Qizhan
authors.initials: Q
authors.lastname: Liu
mesh_terms: D000818:Animals; D018053:Arsenites / Q000633:toxicity*; D001483:Base Sequence; D002461:Cell Line, Transformed; D047369:Chromatin Immunoprecipitation; D017931:DNA Primers; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005260:Female; D006801:Humans; D051379:Mice; D008813:Mice, Inbred ICR; D035683:MicroRNAs / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism*; D015538:Nitrosation; D017382:Reactive Oxygen Species / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D018053:Arsenites; D017931:DNA Primers; C524940:MIRN21 microRNA, human; C531950:MIRN21 microRNA, mouse; D035683:MicroRNAs; D016328:NF-kappa B; D017382:Reactive Oxygen Species; D048049:Extracellular Signal-Regulated MAP Kinases; C015001:arsenite