Array-based FMR1 sequencing and deletion analysis in patients with a fragile X syndrome-like phenotype

Abstract

Background: Fragile X syndrome (FXS) is caused by loss of function mutations in the FMR1 gene. Trinucleotide CGG-repeat expansions, resulting in FMR1 gene silencing, are the most common mutations observed at this locus. Even though the repeat expansion mutation is a functional null mutation, few conventional mutations have been identified at this locus, largely due to the clinical laboratory focus on the repeat tract.

Methodology/principal findings: To more thoroughly evaluate the frequency of conventional mutations in FXS-like patients, we used an array-based method to sequence FMR1 in 51 unrelated males exhibiting several features characteristic of FXS but with normal CGG-repeat tracts of FMR1. One patient was identified with a deletion in FMR1, but none of the patients were found to have other conventional mutations.

Conclusions/significance: These data suggest that missense mutations in FMR1 are not a common cause of the FXS phenotype in patients who have normal-length CGG-repeat tracts. However, screening for small deletions of FMR1 may be of clinically utility.

Figure 1. Targeted resequencing of FMR1.

The horizontal axis is formed by intronic sequence, and the numbered vertical spokes represent the 17 exons of FMR1. Coding exonic sequence is shown in blue, while noncoding exonic sequence is shown in white. The black region upstream of exon 1 is the minimal promoter of FMR1. The grey bars represent the four LR-PCR amplicons used for sequencing. The green boxes represent the FMR1 regions sequenced with the custom resequencing array.

Figure 2. FMRP expression in control and fragile X tissues.

Western blot of lymphoblastoid cell lysate from a healthy control, a fragile X patient, and a patient harboring a novel deletion in the 5′UTR of FMR1. The protein eIF4e was assessed as a loading control.