Abstract
Single nucleotide polymorphisms (SNPs) are the most common form of genetic variation. We previously demonstrated that SNPs (rs1800734, rs749072, and rs13098279) in the MLH1 gene region are associated with MLH1 promoter island methylation, loss of MLH1 protein expression, and microsatellite instability (MSI) in colorectal cancer (CRC) patients. Recent studies have identified less CpG-dense “shore” regions flanking many CpG islands. These shores often exhibit distinct methylation profiles between different tissues and matched normal versus tumor cells of patients. To date, most epigenetic studies have focused on somatic methylation events occurring within solid tumors; less is known of the contributions of peripheral blood cell (PBC) methylation to processes such as aging and tumorigenesis. To address whether MLH1 methylation in PBCs is correlated with tumorigenesis we utilized the Illumina 450 K microarrays to measure methylation in PBC DNA of 846 healthy controls and 252 CRC patients from Ontario, Canada. Analysis of a region of chromosome 3p21 spanning the MLH1 locus in healthy controls revealed that a CpG island shore 1 kb upstream of the MLH1 gene exhibits different methylation profiles when stratified by SNP genotypes (rs1800734, rs749072, and rs13098279). Individuals with wild-type genotypes incur significantly higher PBC shore methylation than heterozygous or homozygous variant carriers (p<1.1×10−6; ANOVA). This trend is also seen in CRC cases (p<0.096; ANOVA). Shore methylation also decreases significantly with increasing age in cases and controls. This is the first study of its kind to integrate PBC methylation at a CpG island shore with SNP genotype status in CRC cases and controls. These results indicate that CpG island shore methylation in PBCs may be influenced by genotype as well as the normal aging process.
Highlights
Epigenetic mechanisms induce functionally relevant changes to the genome without changing the nucleotide sequence itself
We have demonstrated a link between these SNPs and MLH1 CpG island methylation in colorectal cancer (CRC) tumours, but the potential correlation of these three SNPs with MLH1 shore methylation has never been investigated, nor has it been analyzed in peripheral blood cells of normal healthy individuals
846 controls and 252 CRC cases from the Ontario Familial Colorectal Cancer Registry were successfully analyzed for methylation levels across the genome spanning 450,000 CpG sites
Summary
Epigenetic mechanisms induce functionally relevant changes to the genome without changing the nucleotide sequence itself. These mechanisms include DNA methylation, histone modifications and non-coding RNAs. Of these, DNA methylation is the most studied epigenetic mark, with clear links to a variety of diseases established. ALU, LINE-1 repeats) while methylation is low or non-existent in the promoter CpG islands of most genes. These methylation patterns reverse with increasing age, as well as in disease states, including cancer [1,2,3]. The vast majority of epigenetic studies have investigated methylation at CpG islands; the role of CpG island shore methylation is only just beginning to be understood
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