Abstract
Genetic and epigenetic changes contribute to deregulation of gene expression and development of human cancer. Changes in DNA methylation are key epigenetic factors regulating gene expression and genomic stability. Recent progress in microarray technologies resulted in developments of high resolution platforms for profiling of genetic, epigenetic and gene expression changes. OS is a pediatric bone tumor with characteristically high level of numerical and structural chromosomal changes. Furthermore, little is known about DNA methylation changes in OS. Our objective was to develop an integrative approach for analysis of high-resolution epigenomic, genomic, and gene expression profiles in order to identify functional epi/genomic differences between OS cell lines and normal human osteoblasts. A combination of Affymetrix Promoter Tilling Arrays for DNA methylation, Agilent array-CGH platform for genomic imbalance and Affymetrix Gene 1.0 platform for gene expression analysis was used. As a result, an integrative high-resolution approach for interrogation of genome-wide tumour-specific changes in DNA methylation was developed. This approach was used to provide the first genomic DNA methylation maps, and to identify and validate genes with aberrant DNA methylation in OS cell lines. This first integrative analysis of global cancer-related changes in DNA methylation, genomic imbalance, and gene expression has provided comprehensive evidence of the cumulative roles of epigenetic and genetic mechanisms in deregulation of gene expression networks.
Highlights
The hallmark of cancer is the deregulation of gene expression profiles and disruption of molecular networks [1]
To assess genome wide changes in DNA methylation related to OS that may play a role in deregulation of gene expression, as well as to delineate the potential genomic imbalance contributions, an integrative and functional approach for the analysis of DNA methylation, genomic imbalance and gene expression was created, using two OS cell lines, U2OS and MG63, and normal human osteoblasts
12.5 kb regions (2.5 Kb 39 and 7.5–10 Kb) of 25,500 human gene promoters, with an average tilling resolution of 35 nucleotides. This platform was previously used for profiling of genomic histone acetylation in a breast cancer model of environmental exposures [39]. We utilized this platform in combination with the methylated DNA immunoprecipitation (Me-DIP) to develop a comprehensive approach for detection of hypo- and hypermethylation changes at high resolution, and used it to detect such changes in human OS cells in relation to the normal osteoblasts
Summary
The hallmark of cancer is the deregulation of gene expression profiles and disruption of molecular networks [1]. Mutation and genomic instability provide tumours with sufficient diversity, so that cells with adaptive and proliferative selective advantage can evolve in a Darwinian manner. It has become evident that epigenetic factors, heritable changes in DNA methylation, may confer additional and more diverse advantage to tumours including deregulation of gene expression and destabilization of chromatin. Malignant cells can show major disruptions in DNA methylation profiles, which manifest as aberrant hypermethylation and hypomethylation of gene promoters, as well as global genomic hypomethylation [3]. Many genes with aberrant promoter hypermethylation have been identified in tumours, including cell cycle regulators, DNA repair genes, genes associated with apoptosis, hormonal regulation, detoxification, metastasis, angiogenesis, and many others [4,5]
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