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Article: Transcriptional implications of intragenic DNA methylation in the oestrogen receptor alpha gene in breast cancer cells and tissues

TitleTranscriptional implications of intragenic DNA methylation in the oestrogen receptor alpha gene in breast cancer cells and tissues
Authors
KeywordsBreast cancer
Breast cancer campaign tissue bank
Breast epithelial cells
Breast milk
DNA methylation
Issue Date2015
PublisherBioMed Central Ltd. The Journal's web site is located at http://www.biomedcentral.com/bmccancer/
Citation
BMC Cancer, 2015, v. 15 n. 1, article no. 337 How to Cite?
AbstractBackground: DNA methylation variability regions (MVRs) across the oestrogen receptor alpha (ESR1) gene have been identified in peripheral blood cells from breast cancer patients and healthy individuals. In contrast to promoter methylation, gene body methylation may be important in maintaining active transcription. This study aimed to assess MVRs in ESR1 in breast cancer cell lines, tumour biopsies and exfoliated epithelial cells from expressed breast milk (EBM), to determine their significance for ESR1 transcription. Methods: DNA methylation levels in eight MVRs across ESR1 were assessed by pyrosequencing bisulphite-converted DNA from three oestrogen receptor (ER)-positive and three ER-negative breast cancer cell lines. DNA methylation and expression were assessed following treatment with DAC (1M), or DMSO (controls). ESR1 methylation levels were also assayed in DNA from 155 invasive ductal carcinoma biopsies provided by the Breast Cancer Campaign Tissue Bank, and validated with DNA methylation profiles from the TCGA breast tumours (n=356 ER-pos, n=109 ER-neg). DNA methylation was profiled in exfoliated breast epithelial cells from EBM using the Illumina 450K (n=36) and pyrosequencing in a further 53 donor samples. ESR1 mRNA levels were measured by qRT-PCR. Results: We show that ER-positive cell lines had unmethylated ESR1 promoter regions and highly methylated intragenic regions (median, 80.45%) while ER-negative cells had methylated promoters and lower intragenic methylation levels (median, 38.62%). DAC treatment increased ESR1 expression in ER-negative cells, but significantly reduced methylation and expression of ESR1 in ER-positive cells. The ESR1 promoter was unmethylated in breast tumour biopsies with high levels of intragenic methylation, independent of ER status. However, ESR1 methylation in the strongly ER-positive EBM DNA samples were very similar to ER-positive tumour cell lines. Conclusion: DAC treatment inhibited ESR1 transcription in cells with an unmethylated ESR1 promoter and reduced intragenic DNA methylation. Intragenic methylation levels correlated with ESR1 expression in homogenous cell populations (cell lines and exfoliated primary breast epithelial cells), but not in heterogeneous tumour biopsies, highlighting the significant differences between the in vivo tumour microenvironment and individual homogenous cell types. These findings emphasise the need for care when choosing material for epigenetic research and highlights the presence of aberrant intragenic methylation levels in tumour tissue. © 2015 Shenker et al.
Persistent Identifierhttp://hdl.handle.net/10722/274913
ISSN
2017 Impact Factor: 3.288
2015 SCImago Journal Rankings: 1.627
ISI Accession Number ID

 

DC FieldValueLanguage
dc.contributor.authorShenker, NS-
dc.contributor.authorFlower, KJ-
dc.contributor.authorWilhelm-Benartzi, CS-
dc.contributor.authorDai, W-
dc.contributor.authorBell, E-
dc.contributor.authorGore, E-
dc.contributor.authorEl Bahrawy, M-
dc.contributor.authorWeaver, G-
dc.contributor.authorBrown, R-
dc.contributor.authorFlanagan, JM-
dc.date.accessioned2019-09-10T02:31:29Z-
dc.date.available2019-09-10T02:31:29Z-
dc.date.issued2015-
dc.identifier.citationBMC Cancer, 2015, v. 15 n. 1, article no. 337-
dc.identifier.issn1471-2407-
dc.identifier.urihttp://hdl.handle.net/10722/274913-
dc.description.abstractBackground: DNA methylation variability regions (MVRs) across the oestrogen receptor alpha (ESR1) gene have been identified in peripheral blood cells from breast cancer patients and healthy individuals. In contrast to promoter methylation, gene body methylation may be important in maintaining active transcription. This study aimed to assess MVRs in ESR1 in breast cancer cell lines, tumour biopsies and exfoliated epithelial cells from expressed breast milk (EBM), to determine their significance for ESR1 transcription. Methods: DNA methylation levels in eight MVRs across ESR1 were assessed by pyrosequencing bisulphite-converted DNA from three oestrogen receptor (ER)-positive and three ER-negative breast cancer cell lines. DNA methylation and expression were assessed following treatment with DAC (1M), or DMSO (controls). ESR1 methylation levels were also assayed in DNA from 155 invasive ductal carcinoma biopsies provided by the Breast Cancer Campaign Tissue Bank, and validated with DNA methylation profiles from the TCGA breast tumours (n=356 ER-pos, n=109 ER-neg). DNA methylation was profiled in exfoliated breast epithelial cells from EBM using the Illumina 450K (n=36) and pyrosequencing in a further 53 donor samples. ESR1 mRNA levels were measured by qRT-PCR. Results: We show that ER-positive cell lines had unmethylated ESR1 promoter regions and highly methylated intragenic regions (median, 80.45%) while ER-negative cells had methylated promoters and lower intragenic methylation levels (median, 38.62%). DAC treatment increased ESR1 expression in ER-negative cells, but significantly reduced methylation and expression of ESR1 in ER-positive cells. The ESR1 promoter was unmethylated in breast tumour biopsies with high levels of intragenic methylation, independent of ER status. However, ESR1 methylation in the strongly ER-positive EBM DNA samples were very similar to ER-positive tumour cell lines. Conclusion: DAC treatment inhibited ESR1 transcription in cells with an unmethylated ESR1 promoter and reduced intragenic DNA methylation. Intragenic methylation levels correlated with ESR1 expression in homogenous cell populations (cell lines and exfoliated primary breast epithelial cells), but not in heterogeneous tumour biopsies, highlighting the significant differences between the in vivo tumour microenvironment and individual homogenous cell types. These findings emphasise the need for care when choosing material for epigenetic research and highlights the presence of aberrant intragenic methylation levels in tumour tissue. © 2015 Shenker et al.-
dc.languageeng-
dc.publisherBioMed Central Ltd. The Journal's web site is located at http://www.biomedcentral.com/bmccancer/-
dc.relation.ispartofBMC Cancer-
dc.rightsBMC Cancer. Copyright © BioMed Central Ltd.-
dc.rightsThis work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.-
dc.subjectBreast cancer-
dc.subjectBreast cancer campaign tissue bank-
dc.subjectBreast epithelial cells-
dc.subjectBreast milk-
dc.subjectDNA methylation-
dc.titleTranscriptional implications of intragenic DNA methylation in the oestrogen receptor alpha gene in breast cancer cells and tissues-
dc.typeArticle-
dc.identifier.emailDai, W: weidai2@hku.hk-
dc.identifier.authorityDai, W=rp02146-
dc.description.naturepublished_or_final_version-
dc.identifier.doi10.1186/s12885-015-1335-5-
dc.identifier.pmid25927974-
dc.identifier.scopuseid_2-s2.0-84929173797-
dc.identifier.hkuros302885-
dc.identifier.volume15-
dc.identifier.issue1-
dc.identifier.spagearticle no. 337-
dc.identifier.epagearticle no. 337-
dc.identifier.isiWOS:000354164700003-
dc.publisher.placeUnited Kingdom-

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