PO-307 Paradoxical decrease in H2AFX copy number yet increasing mRNA expression in human breast cancer

Abstract

IntroductionHistones are basic proteins responsible for DNA packaging in eukaryotic cell nuclei. H2AX is an H2A variant that plays an important role early in the DNA damage response. Although H2AX expression has been investigated, the direct contribution of this abundant chromatin protein in breast cancer is not well understood. The H2AFX gene encodes the same H2AX protein by both a short non-polyadenylated transcript and a longer polyadenylated transcript. We are characterising H2AFX copy number (CN), mRNA expression and protein abundance in breast cancer to discover if this foundation chromatin component contributing to genome stability could play a role in breast cancer.Material and methodsH2AFX mRNA abundance was analysed in a panel of breast cell lines by absolute quantitative PCR. The Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) dataset was used for bioinformatic analysis.Results and discussionsAnalysis of the METABRIC dataset shows that H2AFX copy number alterations involving loss of one allele are very common (41%) and correlate with genome instability. No diploid deletions are observed, and only 2% of samples show H2AFX CN amplifications. No mutations were observed in the H2AFX coding region. Although we observed that the H2AFX short transcript is more abundant than the long transcript in the panel of breast cell lines, we found that the ratio of short and long transcript is constant. This validates the relevance of transcriptomic databases using polyA-selected H2AFX mRNAs that only report long transcript abundance. Analysis of H2AFX mRNA levels in METABRIC revealed an increase in H2AFX expression with worsening prognosis, despite the decrease in gene dosage. Furthermore, we found that H2AFX copy number decreases while H2AFX mRNA expression increases during the progression from luminal A to luminal B which are the most common breast cancer subtypes.ConclusionAnalysis of primary breast tumour genomic datasets revealed that H2AFX frequently has reduced copy number but is almost never fully deleted, mutated, or amplified. One copy is lost usually as a result of a major deletion in the 11q chromosome arm. Paradoxically, H2AFX expression is frequently increased suggesting that an independent mechanism for regulation exists to drive up-regulation. We are now bringing together H2AFX copy number, mRNA expression and protein data to understand whether H2AFX expression responds to challenges to genome integrity in breast cancer.