FMR1 mRNA from full mutation alleles is associated with ABC-CFX scores in males with fragile X syndrome

Emma K Baker,Lorena Santa María,Jonathan Cohen,Ling Ling,Bianca Curotto,Minh Bui,Claudine Kraan,David J Amor,Michael Field,Isabel Salas,Marta Arpone,Alexandra Ure,Carolyn Rogers,Víctor Faundes ,César Trigo ,Angelica M Alliende ,Patricio Morales ,Matthew Hunter ,Lesley Bretherton,David E Godler

doi:10.1038/s41598-020-68465-6

Abstract

Fragile X syndrome (FXS) is caused by a hypermethylated full mutation (FM) expansion with ≥ 200 CGG repeats, and a decrease in FMR1 mRNA and its protein. However, incomplete silencing from FM alleles has been associated with more severe autism features in FXS males. This study compared scores on the Aberrant Behavior Checklist-Community-FXS version (ABC-CFX) in 62 males affected with FXS (3 to 32 years) stratified based on presence or absence of mosaicism and/or FMR1 mRNA silencing. Associations between ABC-CFX subscales and FMR1 mRNA levels, assessed using real-time PCR relative standard curve method, were also examined. The FXS group mosaic for premutation (PM: 55–199 CGGs) and FM alleles had lower irritability (p = 0.014) and inappropriate speech (p < 0.001) scores compared to males with only FM alleles and complete loss of FMR1 mRNA. The PM/FM mosaic group also showed lower inappropriate speech scores compared to the incomplete silencing (p = 0.002) group. Increased FMR1 mRNA levels were associated with greater irritability (p < 0.001), and lower health-related quality of life scores (p = 0.004), but only in the incomplete silencing FM-only group. The findings suggest that stratification based on CGG sizing and FMR1 mRNA levels may be warranted in future research and clinical trials utilising ABC-CFX subscales as outcome measures.

Highlights

Fragile X syndrome (FXS) is caused by a large trinucleotide CGG expansion (≥ 200 repeats), termed full mutation (FM), in the promoter region of the Fragile X Mental Retardation 1 (FMR1) gene[1]
Rare unmethylated FM (UFM) alleles in adult individuals have been associated with neurodegeneration observed as Fragile X-associated Tremor/Ataxia Syndrome (FXTAS), hypothesised to be related to “RNA gain of function” toxicity originating from UFM a lleles[6,7,8]
This study aimed to determine if maladaptive behaviours are increased, as measured by the Aberrant Behavior Checklist-Community fragile X version (ABC-CFX), in males affected with FXS with complete and incomplete silencing of FM alleles, as compared to males mosaic for PM and FM alleles

Summary

Introduction

Fragile X syndrome (FXS) is caused by a large trinucleotide CGG expansion (≥ 200 repeats), termed full mutation (FM), in the promoter region of the Fragile X Mental Retardation 1 (FMR1) gene[1]. There is evidence from independent s tudies[9,10,11], demonstrating that in the majority of FM males, FMR1 mRNA is not completely silenced (i.e., there is still residual transcription, even in the presence of FM alleles) These studies have reported that between 44 and 60% of FXS males express FMR1 mRNA4,10,11, with this incomplete silencing more recently associated with elevated ASD features in FM-only males, but not intellectual functioning deficits[11]. This suggests that two reciprocal mechanisms, RNA toxicity and FMRP deficiency, may contribute to overlapping aspects of FXS, ID and ASD features. Based on our previous study[11], it was hypothesised that FM-only males with incomplete silencing of FM alleles would have elevated scores on the ABC-CFX compared to males with complete silencing of FM alleles

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