Abstract
Loss of function of the FMR1 gene leads to fragile X syndrome (FXS), the most common form of intellectual disability. The loss of FMR1 function is usually caused by epigenetic silencing of the FMR1 promoter leading to expansion and subsequent methylation of a CGG repeat in the 5′ untranslated region. Very few coding sequence variations have been experimentally characterized and shown to be causal to the disease. Here, we describe a novel FMR1 mutation and reveal an unexpected nuclear export function for the C-terminus of FMRP. We screened a cohort of patients with typical FXS symptoms who tested negative for CGG repeat expansion in the FMR1 locus. In one patient, we identified a guanine insertion in FMR1 exon 15. This mutation alters the open reading frame creating a short novel C-terminal sequence, followed by a stop codon. We find that this novel peptide encodes a functional nuclear localization signal (NLS) targeting the patient FMRP to the nucleolus in human cells. We also reveal an evolutionarily conserved nuclear export function associated with the endogenous C-terminus of FMRP. In vivo analyses in Drosophila demonstrate that a patient-mimetic mutation alters the localization and function of Dfmrp in neurons, leading to neomorphic neuronal phenotypes.
Highlights
Fragile X syndrome (FXS [MIM 300624]) is a highly prevalent, inherited disorder in humans causing intellectual disability accompanied by a spectrum of behavioral and physical abnormalities (Penagarikano et al, 2007)
We identified a guanine insertion in exon 15 [1457insG] (Supplementary Fig S1A and B). This G-insertion mutation alters the open reading frame to one that is not used by any of the alternative FMR1 isoforms (Ensembl Genome Browser), and is predicted to create a novel peptide sequence followed by a premature stop codon, which results in the truncation of the C-terminus of FMRP (Fig 1A)
We studied the expression of GFPFMR1wt+nuclear localization signal (NLS) in HEK293 cells treated with leptomycin B (LMB), a potent and specific inhibitor of nuclear export (Wolff et al, 1997)
Summary
Fragile X syndrome (FXS [MIM 300624]) is a highly prevalent, inherited disorder in humans causing intellectual disability accompanied by a spectrum of behavioral and physical abnormalities (Penagarikano et al, 2007). Sleep disorders and vulnerability to epileptic seizures have been reported in association with FXS (BerryKravis, 2002; Kronk et al, 2010). In the diagnosis of FXS, clinicians routinely screen for large expansions of the polymorphic CGG repeat elements in the FMR1 locus (Xq27.3 [MIM 309550]). In FXS, the naturally occurring CGG repeats are expanded to numbers above 200, which usually leads to hypermethylation of the repeat itself and the upstream FMR1 promoter (Fu et al, 1991; Oberle et al, 1991; Pieretti et al, 1991; Verkerk et al, 1991). The FMR1 is transcriptionally silenced, and no protein product (FMRP) is formed
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