Differential MSH2 promoter methylation in blood cells of Neurofibromatosis type 1 (NF1) patients

Abstract

Neurofibromatosis type 1 (NF1) is caused by NF1 gene mutations. The phenotype is highly variable, with 'modifiers' being discussed as potential determinants. Mismatch repair deficiency was shown to cause NF1 mutations, but constitutional mutation of mismatch repair genes was identified only once in a NF1 patient. We aimed to analyze whether DNA methylation of mismatch repair gene promoters, known to lead to transcriptional silencing, is associated with increased tumor load in NF1 defined by the number of cutaneous neurofibromas. Leukocyte DNA of 79 controls and 79 NF1 patients was investigated for methylation of mismatch repair genes MLH1, MSH2, MSH6, and PMS2 by methylation-specific PCR and pyrosequencing. MLH1, MSH6, and PMS2 promoters were not methylated. By contrast, we found promoter methylation of MSH2 with a higher rate of methylation in NF1 patients compared with controls. Furthermore, when comparing NF1 patients with a low vs those with a high number of cutaneous neurofibromas, MSH2 promoter methylation was significantly different. In patients with a high tumor burden, methylation of two (out of six) CpGs was enhanced. This finding was not confounded by age. In conclusion, enhanced methylation involving transcription start points of mismatch repair genes, such as MSH2 in NF1, has not been described so far. Methylation-induced variability of MSH2 gene expression may lead to variable mismatch repair capacity. Our results may point toward a role of MSH2 as a modifier for NF1, although the amount of DNA methylation and subsequent gene expression in other cell types of NF1 patients needs to be elucidated.

Neurofibromatosis type 1: from genotype to phenotype

“…However, apart from one report, no constitutional mutations in MMR genes have been detected in NF1 patients 44. A recent study examined whether increased tumour load in NF1 (higher number of cutaneous neurofibromas) was associated with methylation of MMR genes 45. Titze et al performed methylation-specific PCR and pyrosequencing of MMR gene promoters most frequently involved in human cancers ( MLH1 , MSH6 , PMS2 , and MSH2 ) in leukocytes of NF1 patients.…”

Comparative methylome analysis of benign and malignant peripheral nerve sheath tumors

“…So far, few studies have assessed the DNAm state of MPNSTs, and most of those have been restricted to the NF1 gene itself (Harder et al 2004) or selected CGIs and promoters (Gonzalez-Gomez et al 2003). Nevertheless, these studies have identified differential DNAm of several genes thought to be involved in MPNST development and progression, including CDKN2A and MSH2 Titze et al 2010). To provide a more comprehensive catalog of DNAm differences in MPNST, we performed unbiased methylome analysis of benign and malignant NF samples as well as controls.…”

Evidence of expression variation and allelic imbalance in Crohn's disease susceptibility genes NOD2 and ATG16L1 in human dendritic cells

“…Several studies have reported the association of DNA methylation of the NOD2 promoter with patient’s age and disease status (Fernandez et al, 2011; Kerkel et al, 2010). Studies on various diseases have shown that differential promoter specific DNA methylation plays a critical role in expression variation between two alleles (Gonsky et al, 2009; Grayson et al, 2005; Movassagh et al, 2010; Titze et al, 2009). Therefore, a possible mechanism for the variation in AI of NOD2 and ATG16L1 is that it results from differences in gene expression between alleles with differential DNA promoter-specific methylation.…”