Tobacco Smoke Induces Polycomb-Mediated Repression of Dickkopf-1 in Lung Cancer Cells

Abstract

Limited information is available about epigenetic mechanisms by which cigarette smoke enhances the initiation and progression of lung cancer. To examine this issue, A549 and Calu-6 lung cancer cells were cultured in normal media with or without tobacco smoke condensate (TSC) under clinically relevant exposure conditions. Ten-day TSC exposure dramatically increased the tumorigenicity of lung cancer cells in nude mice. Microarray and quantitative reverse transcription-PCR (RT-PCR) experiments revealed that this phenomenon coincided with diminished expression of Dickkopf-1 (Dkk-1). Western blot, chromatin immunoprecipitation, methylationspecific PCR, and pyrosequencing experiments showed that repression of Dkk-1 coincided with decreased H4K16Ac, increased H3K27me3, and recruitment of SirT1, EZH2, SUZ12, and Bmi1 without DNA hypermethylation within the Dkk-1 promoter despite prolonged TSC exposures. Removal of TSC from culture media resulted in loss of promoterassociated polycomb repressor complexes and reexpression of Dkk-1. siRNA-mediated knockdown of EZH2 and SirT1 partially abrogated TSC-mediated inhibition of Dkk-1 expression. Western blot and quantitative RT-PCR array experiments showed that TSC exposure as well as knockdown of Dkk-1 activated Wnt signaling and significantly up-regulated Wnt5a in lung cancer cells. Knockdown of Dkk-1 recapitulated the dramatic protumorigenic effects of TSC exposure in Calu-6 cells. Despite the transient nature of Dkk-1 repression following TSC exposure in vitro, Dkk-1 remained silenced in tumor xenografts derived from TSC-treated Calu-6 cells. Collectively, these data provide evidence that cigarette smoke directly engages polycomb machinery to activate a signaling network implicated in maintenance of cancer stem cells.

Cigarette Smoke Induces Epidermal Growth Factor Receptor-Dependent Redistribution of Apical MUC1 and Junctional β-Catenin in Polarized Human Airway Epithelial Cells

“…10 The effects of CS on normal nontransformed airway cells appear to differ from those that are malignantly-transformed. For example, Hussain et al showed that CS directly engaged polycomb machinery to activate canonical Wnt signaling in the maintenance of cancer stem cells, 37 while Marsit et al reported CS-induced Wnt signaling through epigenetic silencing of the extracellular Wnt antagonist, soluble frizzled receptor protein (SFRP). 38 The complexity of mechanisms regulating canonical Wnt signaling in response to CS might reflect a stage-specific control of Wnt signaling activation during the multistep process of malignant transformation.…”

Polycomb (PcG) Proteins, BMI1 and SUZ12, Regulate Arsenic-induced Cell Transformation

“…PcG complexes comprise the machinery of transcriptional regulatory processing in homeotic (HOX) gene expression (12,30), and several cellular responses, including cell cycle (31), senescence (20), cancer (11,(32)(33)(34), cell fate decisions (33) and stem cell differentiation or maintenance (11,35). PcG consists of two distinct protein complexes, PRC1 and PRC2, which are best characterized in the repression of transcription.…”

“…PRC2 maintains and establishes tumor suppressor gene silencing during carcinogenesis (36 -37). Increases in SUZ12, a key component of PRC2, coincide with transformation and many cancers (14,34,36,38). PRC1 recognizes the histone H3K27 methylation and recruits PRC2 to PcG target genes resulting in transcriptional repression (15).…”

Distinctive expression of the polycomb group proteins Bmi1 polycomb ring finger oncogene and enhancer of zeste homolog 2 in nonsmall cell lung cancers and their clinical and clinicopathologic significance

“…Recently, Hussain et al reported that tobacco smoke engaged polycomb repressor complexes that contained EZH2 to mediate epigenetic silencing of Dickkopf-1 (a Wnt signaling antagonist) and enhanced the malignant phenotype of lung cancer cells. 37 Although the mechanism by which EZH2 mediates tumor aggressiveness is unclear, recent studies indicate that EZH2 mediates transcriptional silencing of a tumor suppressor gene E-cadherin (by trimethylation of H3 lysine 27). 38,39 Yu et al demonstrated that EZH2 may mediate increased invasiveness and metastasis by silencing several downstream targets in addition to E-cadherin, including the adrenergic receptor (ADR) ADRB2.…”