Cell-Type Specific DNA Methylation Patterns Define Human Breast Cellular Identity

Petr Novak,Bernard W Futscher,Mathias Ehrich,James C Garbe,Martha R Stampfer,Taylor J Jensen,Jose L Munoz-Rodriguez,William B Coleman

doi:10.1371/journal.pone.0052299

Abstract

DNA methylation plays a role in a variety of biological processes including embryonic development, imprinting, X-chromosome inactivation, and stem cell differentiation. Tissue specific differential methylation has also been well characterized. We sought to extend these studies to create a map of differential DNA methylation between different cell types derived from a single tissue. Using three pairs of isogenic human mammary epithelial and fibroblast cells, promoter region DNA methylation was characterized using MeDIP coupled to microarray analysis. Comparison of DNA methylation between these cell types revealed nearly three thousand cell-type specific differentially methylated regions (ctDMRs). MassARRAY was performed upon 87 ctDMRs to confirm and quantify differential DNA methylation. Each of the examined regions exhibited statistically significant differences ranging from 10–70%. Gene ontology analysis revealed the overrepresentation of many transcription factors involved in developmental processes. Additionally, we have shown that ctDMRs are associated with histone related epigenetic marks and are often aberrantly methylated in breast cancer. Overall, our data suggest that there are thousands of ctDMRs which consistently exhibit differential DNA methylation and may underlie cell type specificity in human breast tissue. In addition, we describe the pathways affected by these differences and provide insight into the molecular mechanisms and physiological overlap between normal cellular differentiation and breast carcinogenesis.

Highlights

DNA methylation is an epigenetic mark located on the carbon-5 position of cytosine residues in mammalian genomes, primarily on the cytosine within a cytosine-guanine sequence in differentiated cells [1]
Alterations in DNA methylation have been demonstrated to occur in human mammary epithelial cells (HMEC) that have overcome a first defined HMEC stress-associated senescence barrier; only HMEC able of responding to stress through the upregulation of p16 and defined as pre-stasis were used for these studies [26,29,38]
The identified cell-type specific differentially methylated regions (ctDMRs) are often associated with genes involved in development and differentiation and these genes often act as transcription factors implicated in cell patterning

Summary

Introduction

DNA methylation is an epigenetic mark located on the carbon-5 position of cytosine residues in mammalian genomes, primarily on the cytosine within a cytosine-guanine sequence in differentiated cells [1]. This epigenetic modification of genomic DNA plays a role in a variety of biological processes including embryonic development, imprinting, and X-chromosome inactivation [2–4]. The differentiation of pluripotent stem cells to comprise the various tissues and cell types within the body is thought to be controlled by epigenetic mechanisms including DNA methylation [5]. In the case of breast cancer, aberrant DNA methylation is a known contributor to the disease [17–19]; the molecular mechanisms associated with disease development and progression are still not well understood

Methods

Results

Conclusion