Aim: Our previous study showed that methylation of the mutL homolog 1 (MLH1) promoter may spread upstream from the Alu elements in intron 1. In this study, we investigated if the Alu methylation could also spread downstream.
Materials and methods: Two colorectal cancer cell lines (RKO, SW48), and four colorectal and three gastric carcinomas [all Microsatellite Instability (MSI)-positive] were selected as cases. Normal colorectal and gastric mucosa and human peripheral blood, and one MSI-negative colorectal cancer case served as controls. After extraction of DNA, bisulfite genomic sequencing was used to analyze the methylation level of exon 2 and the adjacent part of intron 2 of the MLH1 gene.
Results: Exon 2 and the partial intron 2 exhibited a high level of methylation in controls. In contrast, demethylation in these regions was seen in gastrointestinal cancer.
Conclusion: Exon 2 methylation is not likely to influence MLH1 gene expression. It seems that de-methylation of exon 2 and intron 2, in combination with intron 1, is associated with methylation spreading of the MLH1 promoter. The region around exon 2 could possibly bind proteins that regulate methylation, and therefore affect gene expression.