DNA methylation impact on Fabry disease

Abstract

Background: Fabry disease (FD) is a rare X-linked disease caused by mutations in GLA gene with consequent lysosomal accumulation of globotriaosylceramide (Gb3). Women with FD often show highly heterogeneous symptoms that can manifest from mild to severe phenotype.

Main body: The phenotypic variability of the clinical manifestations in heterozygous women with FD mainly depends on the degree and direction of inactivation of the X chromosome. Classical approaches to measure XCI skewness might be not sufficient to explain disease manifestation in women. In addition to unbalanced XCI, allele-specific DNA methylation at promoter of GLA gene may influence the expression levels of the mutated allele, thus impacting the onset and the outcome of FD. In this regard, analyses of DNA methylation at GLA promoter, performed by approaches allowing distinction between mutated and non-mutated allele, may be much more informative. The aim of this review is to critically evaluate recent literature articles addressing the potential role of DNA methylation in the context of FD. Although up to date relatively few works have addressed this point, reviewing all pertinent studies may help to evaluate the importance of DNA methylation analysis in FD and to develop new research and technologies aimed to predict whether the carrier females will develop symptoms.

Conclusions: Relatively few studies have addressed the complexity of DNA methylation landscape in FD that remains poorly investigated. The hope for the future is that ad hoc and ultradeep methylation analyses of GLA gene will provide epigenetic signatures able to predict whether pre-symptomatic female carriers will develop symptoms thus helping timely interventions.

Fig. 1

Different possible methods to analyze allele-specific methylation at GLA gene. Schematic representation of GLA gene showing promoter, exons and introns positions (not in scale) and exemplificative positions of hypothetical point mutations. The methylation state at promoter region is the best candidate factor able to regulate GLA gene expression. In order to identify single molecules containing information necessary to distinguish the two alleles and at the same time methylation profiles, different approaches can be used according to specific cases. In the case in which the mutation occurs close to promoter region (e.g., in exon 1), it is possible to perform amplicon bisulfite sequencing (a) with amplicon length ranging between 200 and 600 bp and then distinguish alleles by bioinformatic analyses, as performed by Echevarria et al. [16]. If mutation is located at downstream exons (e.g., exon 5), it is possible to analyze allele-specific DNA methylation at a region surrounding mutations that may be not informative for regulation of the gene (b), unless in these patients a polymorphisms in the promoter region may be associated with the mutated allele (e.g., following other members of the same family) as shown in De Riso et al. [56]. Alternatively, Nanopore sequencing may analyze much longer regions allowing to match methylation status of the promoter with mutation present in any position of the gene on the same molecule (c). Advantages of using this method are that it is possible to perform methylation analyses without PCR amplification and bisulfite treatment. However, it may be necessary to isolate the whole GLA genomic region by in vitro CRISPR Cas9 system [58]