Mot-2-Mediated Cross Talk between Nuclear Factor-KB and P53 Is Involved Is Arsenite-Induced Tumorigenesis of Human Embryo Lung Fibroblast Cells

Article excerpt

BACKGROUND: Inactivation of p53 is involved in arsenite-induced tumorigenesis; however, the molecular mechanisms remain poorly understood.

OBJECTIVE: We investigated the molecular mechanisms underlying the inactivation of p53 and neoplastic transformation induced by arsenite in human embryo lung fibroblast (HELF) cells.

METHODS: Anchorage-independent growth assays were performed, and tumorigenicity in intact animals was assessed to confirm arsenite-induced neoplastic transformation. We determined the levels and functions of p53, nuclear factor-kappa B (NF-kB; a key transcriptional regulator), and mot-2 (a p53 inhibitor) and their relationships in arsenite-induced transformed HELF cells by two-dimensional electrophoresis, reverse-transcriptase polymerase chain reaction, Western blot, immunofluorescence, and co-immunoprecipitation assays.

RESULTS: Exposure of HELF cells to low levels of arsenite increased their proliferation rate and anchorage-independent growth and disrupted normal contact inhibition. When introduced into nude mice, transformed cells were tumorigenic. We used proteomic analysis to identify proteins with altered expression between untreated and arsenite-exposed cells. We found decreased expression of NF-kB repressing factor (NKRF; an inhibitor of NF-kB-mediated gene transcription), increased expression of mot-2, and increased activation of NF-kB. Changes in cells exposed to 1.0 [micro]M arsenite were more marked than changes in cells exposed to 0.5 or 2.0 [micro]M arsenite. Inactivation of NF-kB prevented malignant transformation induced by 1.0 [micro]M arsenite. Moreover, we also identified a mechanism whereby NF-kB regulated p53. Specifically, activation of NF-kB up-regulated mot-2 expression, which prevented nuclear translocation of p53 and switched the binding preference of the p53 and NF-kB coactivator CBP [cyclic AMP-responsive element binding protein (CREB) binding protein] from p53 to NF-kB.

CONCLUSIONS: mot-2--mediated cross talk between NF-kB and p53 appears to be involved in arsenite-induced tumorigenesis of HELF cells.

KEY WORDS: arsenite, mot-2, NF-kB, p53, signal transduction, tumorigenesis. Environ Health Perspect 118:936-942 (2010). doi:10.1289/ehp.0901677 [Online 3 March 2010]

Arsenite is an environmental toxicant that has been associated with numerous human health problems, including dermatosis, diabetes mellitus, cardiovascular disorders, and cancer (Tapio and Grosche 2006). Epidemiologic evidence implicates exposure to arsenite in causing human cancers of the skin, lung, and bladder [International Agency for Research on Cancer (IARC) 2004]. Although skin is thought to be the most sensitive site for arsenite toxicity, the lung is now recognized as a specific target (Chen et al. 2004). In vitro, arsenite causes malignant transformation of various cells, including human keratinocytes and small airway epithelial cells (Pi et al. 2008; Wen et al. 2008). Arsenite induces tumorigenesis by activating signal pathways and transcription factors, such as mitogen-activated protein kinases, activating factor-1, and nuclear factor-KB (NF-kB), which are involved in promoting cell proliferation and malignant transformation (Huang et al. 2004; Salnikow and Zhitkovich 2008). Although arsenite is carcinogenic, the mechanisms involved remain unclear.

In mammalian cells, p53 (often called the "guardian of the genome") is involved in the maintenance of genome stability. In response to diverse cellular stresses, p53 protein transactivates downstream target genes required for DNA repair, cell cycle arrest, and apoptosis (Pietsch et al. 2008). People carrying one dysfunctional p53 gene in their germline have a high probability of developing a tumor. Restoration of p53 function inhibits ultraviolet B--induced skin carcinogenesis. Thus, functional inactivation of p53 is a common characteristic of tumors (Konstantakou et al. 2009; Lim and Park 2009). …