Genome-wide screen of promoter methylation identifies novel markers in melanoma

  1. Yasuo Koga1,7,
  2. Mattia Pelizzola2,7,
  3. Elaine Cheng3,
  4. Michael Krauthammer4,
  5. Mario Sznol5,
  6. Stephan Ariyan6,
  7. Deepak Narayan6,
  8. Annette M. Molinaro2,
  9. Ruth Halaban3,8 and
  10. Sherman M. Weissman1,8
  1. 1 Department of Genetics, Yale University School of Medicine, New Haven, Connecticut 06520-8059, USA;
  2. 2 Department of Epidemiology and Public Health, Yale University School of Medicine, New Haven, Connecticut 06520-8059, USA;
  3. 3 Department of Dermatology, Yale University School of Medicine, New Haven, Connecticut 06520-8059, USA;
  4. 4 Department of Pathology, Yale University School of Medicine, New Haven, Connecticut 06520-8059, USA;
  5. 5 Comprehensive Cancer Center Section of Medical Oncology, Yale University School of Medicine, New Haven, Connecticut 06520-8059, USA;
  6. 6 Department of Surgery, Yale University School of Medicine, New Haven, Connecticut 06520-8059, USA

    1. ↵7 These authors contributed equally to this work.

    Abstract

    DNA methylation is an important component of epigenetic modifications, which influences the transcriptional machinery aberrant in many human diseases. In this study we present the first genome-wide integrative analysis of promoter methylation and gene expression for the identification of methylation markers in melanoma. Genome-wide promoter methylation and gene expression of eight early-passage human melanoma cell strains were compared with newborn and adult melanocytes. We used linear mixed effect models (LME) in combination with a series of filters based on the localization of promoter methylation relative to the transcription start site, overall promoter CpG content, and differential gene expression to discover DNA methylation markers. This approach identified 76 markers, of which 68 were hyper- and eight hypomethylated (LME, P < 0.05). Promoter methylation and differential gene expression of five markers (COL1A2, NPM2, HSPB6, DDIT4L, MT1G) were validated by sequencing of bisulfite-modified DNA and real-time reverse transcriptase PCR, respectively. Importantly, the incidence of promoter methylation of the validated markers increased moderately in early and significantly in advanced-stage melanomas, using early-passage cell strains and snap-frozen tissues (n = 18 and n = 24, respectively) compared with normal melanocytes and nevi (n = 11 and n = 9, respectively). Our approach allows robust identification of methylation markers that can be applied to other studies involving genome-wide promoter methylation. In conclusion, this study represents the first unbiased systematic effort to determine methylation markers in melanoma and revealed several novel genes regulated by promoter methylation that were not described in cancer cells before.

    Footnotes

    • ↵8 Corresponding authors.

      E-mail ruth.halaban{at}yale.edu; fax (203) 785-7637.

      E-mail sherman.weissman{at}yale.edu; fax (203) 737-2286.

    • [Supplemental material is available online at www.genome.org. The microarray data from this study have been submitted to Gene Expression Omnibus (GEO) (http://www.ncbi.nlm.nih.gov/geo) under accession no. GSE13706.]

    • Article published online before print. Article and publication date are at http://www.genome.org/cgi/doi/10.1101/gr.091447.109.

      • Received January 20, 2009.
      • Accepted May 20, 2009.
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    1. Published in Advance June 2, 2009, doi: 10.1101/gr.091447.109 Genome Res. 19: 1462-1470 Copyright © 2009 by Cold Spring Harbor Laboratory Press

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