Integrated Analysis of Gene Expression, CpG Island Methylation, and Gene Copy Number in Breast Cancer Cells by Deep Sequencing

Abstract

We used deep sequencing technology to profile the transcriptome, gene copy number, and CpG island methylation status simultaneously in eight commonly used breast cell lines to develop a model for how these genomic features are integrated in estrogen receptor positive (ER+) and negative breast cancer. Total mRNA sequence, gene copy number, and genomic CpG island methylation were carried out using the Illumina Genome Analyzer. Sequences were mapped to the human genome to obtain digitized gene expression data, DNA copy number in reference to the non-tumor cell line (MCF10A), and methylation status of 21,570 CpG islands to identify differentially expressed genes that were correlated with methylation or copy number changes. These were evaluated in a dataset from 129 primary breast tumors. Gene expression in cell lines was dominated by ER-associated genes. ER+ and ER− cell lines formed two distinct, stable clusters, and 1,873 genes were differentially expressed in the two groups. Part of chromosome 8 was deleted in all ER− cells and part of chromosome 17 amplified in all ER+ cells. These loci encoded 30 genes that were overexpressed in ER+ cells; 9 of these genes were overexpressed in ER+ tumors. We identified 149 differentially expressed genes that exhibited differential methylation of one or more CpG islands within 5 kb of the 5′ end of the gene and for which mRNA abundance was inversely correlated with CpG island methylation status. In primary tumors we identified 84 genes that appear to be robust components of the methylation signature that we identified in ER+ cell lines. Our analyses reveal a global pattern of differential CpG island methylation that contributes to the transcriptome landscape of ER+ and ER− breast cancer cells and tumors. The role of gene amplification/deletion appears to more modest, although several potentially significant genes appear to be regulated by copy number aberrations.

Ribosome-omics of the human ribosome

“…Note that either overexpression or underexpression of individual RPs can lead to accumulation of p53 and apoptosis, and in some cases to tumorigenesis (for review, see Raiser et al 2014). It is interesting that one of several breast tumor lines, BT474, has a 10-fold amplification of a segment of chromosome 17 that includes RPL17 and RPL23, accompanied by a 10-fold excess of the mRNA for each (Sun et al 2011). It would be interesting to know if such an excess has any effect on the cell.…”

iReckon: Simultaneous isoform discovery and abundance estimation from RNA-seq data

“…Finally, we use iReckon to explore the transcriptomes of two breast cancer data sets-a patient sample recently sequenced at the BC Genome Sciences Center (Shah et al 2012) and the MCF7 cell line (accession no. SRX040504) (Sun et al 2011).…”

Notch Signaling Activation Is Associated with Patient Mortality and Increased FGF1-Mediated Invasion in Squamous Cell Carcinoma of the Oral Cavity

“…mRNA was analyzed by the UAB NanoString Laboratory (www.uab.edu/ medicine/radonc/en/nanostring; ref. 19). Samples were processed for analysis on the NanoString nCounter Flex system using the PanCancer Pathways Plus panel as per the manufacturer's instructions (NanoString Technologies).…”