Microsatellite instability and promoter methylation as possible causes of NF1 gene inactivation in neurofibromas

Abstract

Neurofibromatosis type 1 (NF1) is a frequent hereditary disorder. One of the characteristic features of this disease is the development of neurofibromas. Since the NF1 gene is supposed to be a tumour suppressor gene, these neurofibromas should develop upon inactivation of both NF1 alleles. So far, mutation and deletion have been found to be involved in NF1 gene inactivation. However, these inactivating mechanisms explain the development of only a limited fraction of analysed neurofibromas. In this study, we investigated microsatellite instability (MSI) and promoter methylation as potential contributors to NF1 gene inactivation. As site-specific methylation in the NF1 promoter inhibits binding of transcription factors Sp1 and CREB, we studied the methylation status of their binding sites in particular. We analysed 20 neurofibromas and three neurofibrosarcomas, but did not find evidence for microsatellite instability or NF1 promoter methylation in any of the tumours. Thus, our data suggest that both microsatellite instability and promoter methylation are unlikely to be the major causes of NF1 gene inactivation in these tumours.

Exploring the somatic NF1 mutational spectrum associated with NF1 cutaneous neurofibromas

“…17,18 Indeed, three of the somatic NF1 mutations characterised here (p.R816X, p.R304X, p.L1569X) have previously been identified as somatic lesions (two of these three mutations are CpG located and therefore compatible with a mechanism of methylation-mediated deamination of 5-methylcytosine) in both benign and malignant tumours. [41][42][43][44][45][46] A total of seven novel somatic NF1 missense mutations were identified during the course of this study. To ascertain their functional/pathological relevance, we used a series of bioinformatic tools Abbreviations: Del, deletion; fs, frameshift; Ins, insertion; LOH, loss of heterozygosity; MLPA, multiplex ligation-dependent probe amplification; n/a, not applicable (RefSeq: NM_000267); NF1, neurofibromatosis type-1; UTR, untranslated region.…”

“…Tumour-associated microsatellite instability was not analysed here, as our previous studies failed to detect significant levels of microsatellite instability in these cutaneous neurofibromas. 44 Although LOH within the NF1 gene region was observed in 25 cutaneous neurofibromas, we sought for the remaining intragenic somatic lesions to ascertain whether there might be a relationship between the type of somatic NF1 mutation found in a given tumour and the type of germline NF1 mutation in the same patient. Analysis of the somatic NF1 mutation data (Table 1) indicated that, with the exception of six tumours derived from three unrelated patients (T49.2/T49.3; T133.2/T137; and T150.2/T180.1), all individual cutaneous neurofibromas derived from the same patient were found to harbour independent somatic NF1 mutations, consistent with the findings of previous studies.…”

Characterization of functional elements in the neurofibromatosis (NF1) proximal promoter region

“…In contrast, only two studies have addressed DNA methylation in the context of the NF1 gene (Horan et al, 2000;Luijten et al, 2000). In particular, Horan et al (2000) tested a limited number of NF1-related neurofibromas and peripheral blood cells, and observed tumorspecific cytosine methylation at only a small number of cytosines upstream of the transcription start site.…”

Quest: A New Approach to Molecular Staging of Tumors

“…In other neurofibromas, somatic mutations appear to destabilize NF1 mRNA (66). Transcriptional silencing via hypermethylation of promoter regions, a prominent mechanism of inactivating other tumor suppressor genes (72), has not been detected in neurofibroma tissue (73,74). As expected from the mutational and LOH analyses, some neurofibromas had no detectable NFl transcripts or neurofibromin (75,76).…”