CpG island methylation affects accessibility of the proximal BRCA1 promoter to transcription factors

Jinhua Xu, Dezheng Huo, Yinghua Chen, Chika Nwachukwu, Cindy Collins, Janelle Rowell, Dennis J. Slamon, Olufunmilayo I. Olopade

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

To understand the mechanism of transcriptional down-regulation of BRCA1 by promoter methylation, we screened 51 breast cancer cell lines and identified HCC38 as another BRCA1 promoter-methylated cell line in addition to UACC3199. There was low expression of BRCA1 mRNA and BRCA1 protein in both cell lines as measured by quantitative RT-PCR and western blot analysis. After transient treatment with 5-aza-2′-deoxycytidine (5-azaCdR) and trichostatin A (TSA), re-expression of BRCA1 mRNA and BRCA1 protein was detected in UACC3199 cells, but not in HCC38 cells. Another demethylating agent, zebularine, did not induce BRCA1 re-expression in either cell line. To test the hypothesis that methylation of CpG sites may affect accessibility of the BRCA1 promoter to transcription factors and consequently cause down-regulation of BRCA1, we analyzed the binding of four transcription factors (CTCF, SpI, E2F1 and E2F6) to the BRCA1 promoter using chromatin immunoprecipitation assay (ChIP) and quantitative PCR. CTCF and E2F were enriched at the unmethylated BRCA1 promoter in MCF-7 cells. In contrast, these two transcription factors were not enriched at the methylated BRCA1 promoter in UACC3199 and HCC38 cells. Following demethylating drug treatment, E2F1 was enriched at the BRCA1 promoter in the demethylated UACC3199 cells. This indicates that reduced accessibility of transcription factors to the methylated promoter is one of the mechanisms for down-regulation of BRCA1 in heavily methylated cancer cells.

Original languageEnglish (US)
Pages (from-to)593-601
Number of pages9
JournalBreast Cancer Research and Treatment
Volume120
Issue number3
DOIs
StatePublished - Apr 1 2010
Externally publishedYes

Fingerprint

CpG Islands
Methylation
Transcription Factors
BRCA1 Protein
Cell Line
Down-Regulation
pyrimidin-2-one beta-ribofuranoside
decitabine
trichostatin A
Polymerase Chain Reaction
Messenger RNA
Chromatin Immunoprecipitation
MCF-7 Cells
Western Blotting
Breast Neoplasms
Pharmaceutical Preparations
Neoplasms

Keywords

  • BRCA1
  • ChIP assay
  • Methylation
  • Promoter
  • Transcription factor

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Cite this

CpG island methylation affects accessibility of the proximal BRCA1 promoter to transcription factors. / Xu, Jinhua; Huo, Dezheng; Chen, Yinghua; Nwachukwu, Chika; Collins, Cindy; Rowell, Janelle; Slamon, Dennis J.; Olopade, Olufunmilayo I.

In: Breast Cancer Research and Treatment, Vol. 120, No. 3, 01.04.2010, p. 593-601.

Research output: Contribution to journalArticle

Xu, Jinhua ; Huo, Dezheng ; Chen, Yinghua ; Nwachukwu, Chika ; Collins, Cindy ; Rowell, Janelle ; Slamon, Dennis J. ; Olopade, Olufunmilayo I. / CpG island methylation affects accessibility of the proximal BRCA1 promoter to transcription factors. In: Breast Cancer Research and Treatment. 2010 ; Vol. 120, No. 3. pp. 593-601.
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