Non-USH2A mutations in USH2 patients

CHU Montpellier, Laboratoire de Génétique Moléculaire, Montpellier, France.
Human Mutation (Impact Factor: 5.14). 03/2012; 33(3):504-10. DOI: 10.1002/humu.22004
Source: PubMed


We have systematically analyzed the two known minor genes involved in Usher syndrome type 2, DFNB31 and GPR98, for mutations in a cohort of 31 patients not linked to USH2A. PDZD7, an Usher syndrome type 2 (USH2) related gene, was analyzed when indicated. We found that mutations in GPR98 contribute significantly to USH2. We report 17 mutations in 10 individuals, doubling the number of GPR98 mutations reported to date. In contrast to mutations in usherin, the mutational spectrum of GPR98 predominantly results in a truncated protein product. This is true even when the mutation affects splicing, and we have incorporated a splicing reporter minigene assay to show this, where appropriate. Only two mutations were found which we believe to be genuine missense changes. Discrepancy in the mutational spectrum between GPR98 and USH2A is discussed. Only two patients were found with mutations in DFNB31, showing that mutations of this gene contribute to only a very small extent to USH2. Close examination of the clinical details, where available, for patients in whom no mutation was found in USH2A, GPR98, or DFNB31, showed that most of them had atypical features. In effect, these three genes account for the vast majority of USH2 patients and their analysis provide a robust pathway for routine molecular diagnosis.

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Available from: Thomas Besnard
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    • "Patients reach legal blindness at a median age of 58 years [Sandberg et al., 2008]. Mutations in one of three known genes (USH2A, GPR98 [ADGRV1] and DFNB31) can be responsible for USH2, but USH2A (MIM #608400) is by far the most frequently involved and accounts for up to 82% of cases [Dreyer et al., 2008; Garcia-Garcia et al., 2011; Besnard et al., 2012; Le Quesne Stabej et al., 2012]. "
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    ABSTRACT: Deep intronic mutations leading to pseudoexon (PE) insertions are underestimated and most of these splicing alterations have been identified by transcript analysis, for instance, the first deep intronic mutation in USH2A, the gene most frequently involved in Usher syndrome type II (USH2). Unfortunately, analyzing USH2A transcripts is challenging and for 1.8%-19% of USH2 individuals carrying a single USH2A recessive mutation, a second mutation is yet to be identified. We have developed and validated a DNA next-generation sequencing approach to identify deep intronic variants in USH2A and evaluated their consequences on splicing. Three distinct novel deep intronic mutations have been identified. All were predicted to affect splicing and resulted in the insertion of PEs, as shown by minigene assays. We present a new and attractive strategy to identify deep intronic mutations, when RNA analyses are not possible. Moreover, the bioinformatics pipeline developed is independent of the gene size, implying the possible application of this approach to any disease-linked gene. Finally, an antisense morpholino oligonucleotide tested in vitro for its ability to restore splicing caused by the c.9959-4159A>G mutation provided high inhibition rates, which are indicative of its potential for molecular therapy.
    Full-text · Article · Oct 2015 · Human Mutation
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    • "Mutations in three genes, USH2A [13], GPR98 [14], and DFNB31/WHRN [15] have been identified as disease-causing for USH2. USH2A is the most frequently mutated gene in USH2 patients and mutations in USH2A alone account for 85% of USH2 while mutations in GPR98 account for 6% of USH2 in the French and other Caucasian populations [16], [17]. Mutations in one gene CLRN1 are found causative for USH3 [18]. "
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    ABSTRACT: Background Usher syndrome (USH) is a genetically heterogeneous condition with ten disease-causing genes. The spectrum of genes and mutations causing USH in the Lebanese and Middle Eastern populations has not been described. Consequently, diagnostic approaches designed to screen for previously reported mutations were unlikely to identify the mutations in 11 unrelated families, eight of Lebanese and three of Middle Eastern origins. In addition, six of the ten USH genes consist of more than 20 exons, each, which made mutational analysis by Sanger sequencing of PCR-amplified exons from genomic DNA tedious and costly. The study was aimed at the identification of USH causing genes and mutations in 11 unrelated families with USH type I or II. Methods Whole exome sequencing followed by expanded familial validation by Sanger sequencing. Results We identified disease-causing mutations in all the analyzed patients in four USH genes, MYO7A, USH2A, GPR98 and CDH23. Eleven of the mutations were novel and protein truncating, including a complex rearrangement in GPR98. Conclusion Our data highlight the genetic diversity of Usher syndrome in the Lebanese population and the time and cost-effectiveness of whole exome sequencing approach for mutation analysis of genetically heterogeneous conditions caused by large genes.
    Full-text · Article · Sep 2014 · PLoS ONE
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    • "Furthermore, large rearrangements have been described in MYO7A, CDH23, GPR98, USH2A and, particularly, in PCDH15, and their detection requires array-CGH studies and/or multiplex ligation-dependent probe amplification (see USHbases). Taken together, these strategies allow a reliable diagnosis for Usher patients with a mutation detection rate of about 90% for USH1 and USH2 patients (Roux et al. 2011; Besnard et al. 2012). A genotyping microarray commercially available (Cremers et al. 2006) allows rapid screening for hundreds of previously identified variations in nine USH genes (Vozzi et al. 2011), but its application in clinical diagnosis is hampered by a very low detection rate as most USHcausing DNA alterations are private or restricted to one or two families (see USHBases). "
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    ABSTRACT: We show that massively parallel targeted sequencing of 19 genes provides a new and reliable strategy for molecular diagnosis of Usher syndrome (USH) and nonsyndromic deafness, particularly appropriate for these disorders characterized by a high clinical and genetic heterogeneity and a complex structure of several of the genes involved. A series of 71 patients including Usher patients previously screened by Sanger sequencing plus newly referred patients was studied. Ninety-eight percent of the variants previously identified by Sanger sequencing were found by next-generation sequencing (NGS). NGS proved to be efficient as it offers analysis of all relevant genes which is laborious to reach with Sanger sequencing. Among the 13 newly referred Usher patients, both mutations in the same gene were identified in 77% of cases (10 patients) and one candidate pathogenic variant in two additional patients. This work can be considered as pilot for implementing NGS for genetically heterogeneous diseases in clinical service.
    Full-text · Article · Jan 2014
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