title: Epigenetic DNA methylation of antioxidative stress regulator NRF2 in human prostate cancer.
abstract: Epigenetic control of NRF2, a master regulator of many critical antioxidative stress defense genes in human prostate cancer (CaP), is unknown. Our previous animal study found decreased Nrf2 expression through promoter CpG methylation/histone modifications during prostate cancer progression in TRAMP mice. In this study, we evaluated CpG methylation of human NRF2 promoter in 27 clinical prostate cancer samples and in LNCaP cells using MAQMA analysis and bisulfite genomic DNA sequencing. Prostate cancer tissue microarray (TMA) containing normal and prostate cancer tissues was studied by immunohistochemistry. Luciferase reporter assay using specific human NRF2 DNA promoter segments and chromatin immunoprecipitation (ChIP) assay against histone modifying proteins were performed in LNCaP cells. Three specific CpG sites in the NRF2 promoter were found to be hypermethylated in clinical prostate cancer samples (BPH<ADT-RCaP<AS-CaP). NRF2 staining in human prostate cancer TMA showed a decreasing trend for both intensity and percentage of positive cells from normal tissues to advanced-stage prostate cancer (Gleason score from 3-9). Reporter assays in the LNCaP cells containing these three CpG sites showed methylation-inhibited transcriptional activity of the NRF2 promoter. LNCaP cells treated with 5-aza/TSA restored the expression of NRF2 and NRF2 downstream target genes, decreased expression levels of DNMT and HDAC proteins, and ChIP assays showed increased RNA Pol II and H3Ac with a concomitant decrease in H3K9me3, MBD2, and MeCP2 at CpG sites of human NRF2 promoter. Taken together, these findings suggest that epigenetic modification may contribute to the regulation of transcription activity of NRF2, which could be used as prevention and treatment target of human prostate cancer.
authors.affiliation: Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey.
authors.forename: Tin Oo
authors.initials: TO
authors.lastname: Khor
authors.affiliation: Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey.
authors.forename: Francisco
authors.initials: F
authors.lastname: Fuentes
authors.affiliation: Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey.
authors.forename: Limin
authors.initials: L
authors.lastname: Shu
authors.affiliation: Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey.
authors.forename: Ximena
authors.initials: X
authors.lastname: Paredes-Gonzalez
authors.affiliation: Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey.
authors.forename: Anne Yuqing
authors.initials: AY
authors.lastname: Yang
authors.affiliation: Department of Chemical Biology, Susan Lehman Cullman Laboratory for Cancer Research, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey.
authors.forename: Yue
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York.
authors.forename: Dominic J
authors.initials: DJ
authors.lastname: Smiraglia
authors.affiliation: Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University, Baltimore, Maryland.
authors.forename: Srinivasan
authors.initials: S
authors.lastname: Yegnasubramanian
authors.affiliation: Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University, Baltimore, Maryland.
authors.forename: William G
authors.initials: WG
authors.lastname: Nelson
authors.affiliation: Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, New Jersey. KongT@pharmacy.rutgers.edu.
authors.forename: Ah-Ng Tony
authors.initials: AN
authors.lastname: Kong
mesh_terms: D000818:Animals; D015153:Blotting, Western; D047369:Chromatin Immunoprecipitation; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation; D044127:Epigenesis, Genetic / Q000235:genetics*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007124:Immunoenzyme Techniques; D008297:Male; D051379:Mice; D051267:NF-E2-Related Factor 2 / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011471:Prostatic Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
chemical_list: D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D012333:RNA, Messenger