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Sci Rep. 2019 Sep 17;9(1):13463. doi: 10.1038/s41598-019-49952-x.

The polymorphic variant rs1800734 influences methylation acquisition and allele-specific TFAP4 binding in the MLH1 promoter leading to differential mRNA expression.

Author information

1
Cancer Gene Regulation Group, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford, OX3 7BN, UK.
2
Anatomic Pathology Unit, Department of Medicine and Surgery and Research Center of Hereditary and Familial Tumors, University of Insubria, Varese, 21100, Italy.
3
Institute of Structural and Molecular Biology, Department of, Biological Sciences, Birkbeck, London, UK.
4
Ludwig Institute for Cancer Research Ltd, University of Oxford, Nuffield Department of Clinical Medicine, Old Road Campus Research Building, Roosevelt Drive, Oxford, OX3 7DQ, UK.
5
Fundación Pública Galega de Medicina Xenómica, Grupo de Medicina Xenómica, IDIS, Santiago de Compostela, Spain.
6
Cancer Genetics and Evolution Laboratory, Institute of Cancer and Genomic Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK.
7
Intestinal Stem Cell Biology Group, Wellcome Trust Centre for Human Genetics, Oxford University, Roosevelt Drive, Oxford, OX3 7BN, UK.
8
Gastrointestinal Cancer Genetics Laboratory, Institute of Cancer and Genomic Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK.
9
Oxford Institute of Radiation Oncology, University of Oxford, Old Road Campus Research Building, Roosevelt Drive, Oxford, OX3 7DQ, UK.
10
Cancer Gene Regulation Group, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford, OX3 7BN, UK. lewisa@well.ox.ac.uk.
11
Division of Biosciences, Department of Life Sciences, Brunel University London, Old Road Campus Research Building, Roosevelt Drive, Uxbridge, UB8 3PN, UK. lewisa@well.ox.ac.uk.

Abstract

Expression of the mismatch repair gene MutL homolog 1 (MLH1) is silenced in a clinically important subgroup of sporadic colorectal cancers. These cancers exhibit hypermutability with microsatellite instability (MSI) and differ from microsatellite-stable (MSS) colorectal cancers in both prognosis and response to therapies. Loss of MLH1 is usually due to epigenetic silencing with associated promoter methylation; coding somatic mutations rarely occur. Here we use the presence of a colorectal cancer (CRC) risk variant (rs1800734) within the MLH1 promoter to investigate the poorly understood mechanisms of MLH1 promoter methylation and loss of expression. We confirm the association of rs1800734 with MSI+ but not MSS cancer risk in our own data and by meta-analysis. Using sensitive allele-specific detection methods, we demonstrate that MLH1 is the target gene for rs1800734 mediated cancer risk. In normal colon tissue, small allele-specific differences exist only in MLH1 promoter methylation, but not gene expression. In contrast, allele-specific differences in both MLH1 methylation and expression are present in MSI+ cancers. We show that MLH1 transcriptional repression is dependent on DNA methylation and can be reversed by a methylation inhibitor. The rs1800734 allele influences the rate of methylation loss and amount of re-expression. The transcription factor TFAP4 binds to the rs1800734 region but with much weaker binding to the risk than the protective allele. TFAP4 binding is absent on both alleles when promoter methylation is present. Thus we propose that TFAP4 binding shields the protective rs1800734 allele of the MLH1 promoter from BRAF induced DNA methylation more effectively than the risk allele.

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