Significantly Elevated FMR1 mRNA and Mosaicism for Methylated Premutation and Full Mutation Alleles in Two Brothers with Autism Features Referred for Fragile X Testing.

Michael Field,David Francis,Emma K Baker,Chriselle Hickerton,Dean G Phelan,Solange M Aliaga,David E Godler,Howard Slater,Marta Arpone,Elizabeth E Palmer,David J Amor,Ling Ling,Lesley Bretherton,Carolyn Rogers,Tracy Dudding-Byth

doi:10.3390/ijms20163907

Abstract

Although fragile X syndrome (FXS) is caused by a hypermethylated full mutation (FM) expansion with ≥200 cytosine-guanine-guanine (CGG) repeats, and a decrease in FMR1 mRNA and its protein (FMRP), incomplete silencing has been associated with more severe autism features in FXS males. This study reports on brothers (B1 and B2), aged 5 and 2 years, with autistic features and language delay, but a higher non-verbal IQ in comparison to typical FXS. CGG sizing using AmplideX PCR only identified premutation (PM: 55–199 CGGs) alleles in blood. Similarly, follow-up in B1 only revealed PM alleles in saliva and skin fibroblasts; whereas, an FM expansion was detected in both saliva and buccal DNA of B2. While Southern blot analysis of blood detected an unmethylated FM, methylation analysis with a more sensitive methodology showed that B1 had partially methylated PM alleles in blood and fibroblasts, which were completely unmethylated in buccal and saliva cells. In contrast, B2 was partially methylated in all tested tissues. Moreover, both brothers had FMR1 mRNA ~5 fold higher values than those of controls, FXS and PM cohorts. In conclusion, the presence of unmethylated FM and/or PM in both brothers may lead to an overexpression of toxic expanded mRNA in some cells, which may contribute to neurodevelopmental problems, including elevated autism features.

Highlights

Fragile X syndrome (FXS) is a leading single-gene cause of inherited intellectual disability (ID), with a prevalence of up to 1 in 4000 [1]
This study describes two young brothers with expanded fragile X mental retardation 1 (FMR1) alleles, who were ‘higher functioning’ based on intellectual functioning assessments compared to a typical fragile X syndrome (FXS) cohort reported in previous studies [6]
Based on methylation-sensitive Southern blot analysis of blood DNA, both brothers have been described as having unmethylated FM (UFM) alleles approaching 200 CGGs in blood

Summary

Introduction

Fragile X syndrome (FXS) is a leading single-gene cause of inherited intellectual disability (ID), with a prevalence of up to 1 in 4000 [1]. The vast majority (~90%) of males affected by FXS, and ~50% of females, show autism spectrum disorder (ASD) features, including speech perseveration, compulsions, echolalia, repetitive behaviors, poor eye contact, and deficits in social communication [2,3,4,5]. Many of these features are shared with idiopathic ASD [6]. The primary molecular cause of FXS is abnormal regulation of the fragile X mental retardation 1 (FMR1) gene due to the presence of ≥200 cytosine-guanine-guanine (CGG) repeats, termed full mutation (FM), within the FMR1 promoter (reviewed in Kraan et al [7]). Loss of FMR1 mRNA in turn leads to depletion of the fragile X mental retardation protein (FMRP) [13,14,15,16], which is the primary cause of FXS symptomology, as FMRP has a critical function in synaptic plasticity and normal brain development [17,18,19,20,21]

Methods

Results

Conclusion