title: Genome-wide DNA methylation analysis reveals estrogen-mediated epigenetic repression of metallothionein-1 gene cluster in breast cancer.
abstract: BACKGROUND
  Recent genome-wide analysis has shown that DNA methylation spans long stretches of chromosome regions consisting of clusters of contiguous CpG islands or gene families. Hypermethylation of various gene clusters has been reported in many types of cancer. In this study, we conducted methyl-binding domain capture (MBDCap) sequencing (MBD-seq) analysis on a breast cancer cohort consisting of 77 patients and 10 normal controls, as well as a panel of 38 breast cancer cell lines.
  
  
  RESULTS
  Bioinformatics analysis determined seven gene clusters with a significant difference in overall survival (OS) and further revealed a distinct feature that the conservation of a large gene cluster (approximately 70 kb) metallothionein-1 (MT1) among 45 species is much lower than the average of all RefSeq genes. Furthermore, we found that DNA methylation is an important epigenetic regulator contributing to gene repression of MT1 gene cluster in both ERα positive (ERα+) and ERα negative (ERα-) breast tumors. In silico analysis revealed much lower gene expression of this cluster in The Cancer Genome Atlas (TCGA) cohort for ERα + tumors. To further investigate the role of estrogen, we conducted 17β-estradiol (E2) and demethylating agent 5-aza-2'-deoxycytidine (DAC) treatment in various breast cancer cell types. Cell proliferation and invasion assays suggested MT1F and MT1M may play an anti-oncogenic role in breast cancer.
  
  
  CONCLUSIONS
  Our data suggests that DNA methylation in large contiguous gene clusters can be potential prognostic markers of breast cancer. Further investigation of these clusters revealed that estrogen mediates epigenetic repression of MT1 cluster in ERα + breast cancer cell lines. In all, our studies identify thousands of breast tumor hypermethylated regions for the first time, in particular, discovering seven large contiguous hypermethylated gene clusters.
authors.affiliation: Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center at San Antonio, STRF, Room 225, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA.
authors.forename: Rohit R
authors.initials: RR
authors.lastname: Jadhav
authors.affiliation: Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center at San Antonio, STRF, Room 225, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA.
authors.forename: Zhenqing
authors.initials: Z
authors.lastname: Ye
authors.affiliation: Department of Obstetrics and Gynecology, Taipei Medical University Shuang Ho Hospital, New Taipei City, 23561 Taiwan.
authors.forename: Rui-Lan
authors.initials: RL
authors.lastname: Huang
authors.affiliation: Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center at San Antonio, STRF, Room 225, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA.
authors.forename: Joseph
authors.initials: J
authors.lastname: Liu
authors.affiliation: Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center at San Antonio, STRF, Room 225, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA.
authors.forename: Pei-Yin
authors.initials: PY
authors.lastname: Hsu
authors.affiliation: Department of Obstetrics and Gynecology, Medical College of Wisconsin, 9200 West Wisconsin Avenue, Froedtert Medical College Lab Building (FMCLB) 258, Milwaukee, 53226 WI USA.
authors.forename: Yi-Wen
authors.initials: YW
authors.lastname: Huang
authors.affiliation: Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center at San Antonio, STRF, Room 225, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA ; Department of Pharmaceutical Sciences, Biotechnology Program/RENORBIO, Health Sciences Center, Universidade Federal do Espirito Santo, Av. Marechal Campos, 1468, Maruipe, 29040-090 Vitoria ES Brazil ; Programa Ciencias Sem Fronteiras, CNPq, Brasilia, Brazil.
authors.forename: Leticia B
authors.initials: LB
authors.lastname: Rangel
authors.affiliation: Department of Obstetrics and Gynecology, Taipei Medical University Shuang Ho Hospital, New Taipei City, 23561 Taiwan ; School of Medicine, Taipei Medical University, No. 250, Wu-Hsing Street, Taipei, 110 Taiwan ; Graduate Institute of Life Sciences, Department and Graduate Institute of Biochemistry, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Hung-Cheng
authors.initials: HC
authors.lastname: Lai
authors.affiliation: Departamento de Pathologı'a, Universidad de la Frontera, Claro Solar 115, Temuco, Chile.
authors.forename: Juan Carlos
authors.initials: JC
authors.lastname: Roa
authors.affiliation: Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center at San Antonio, STRF, Room 225, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA ; Cancer Therapy and Research Center, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA ; Department of Epidemiology and Biostatistics, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA.
authors.forename: Nameer B
authors.initials: NB
authors.lastname: Kirma
authors.affiliation: Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center at San Antonio, STRF, Room 225, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA ; Cancer Therapy and Research Center, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA ; Department of Epidemiology and Biostatistics, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA.
authors.forename: Tim Hui-Ming
authors.initials: TH
authors.lastname: Huang
authors.affiliation: Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center at San Antonio, STRF, Room 225, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA ; Cancer Therapy and Research Center, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA ; Department of Epidemiology and Biostatistics, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, 78229 TX USA.
authors.forename: Victor X
authors.initials: VX
authors.lastname: Jin
keywords: DNA methylation; MT1