Ex Vivo Splicing Assays of Mutations at Noncanonical Positions of Splice Sites in USHER Genes

被引:39
作者
Le Gueard-Mereuze, Sandie [1 ,2 ]
Vache, Christel [3 ]
Baux, David [3 ]
Faugere, Valerie [3 ]
Larrieu, Lise [3 ]
Abadie, Caroline [3 ]
Janecke, Andreas [4 ]
Claustres, Mireille [1 ,2 ,3 ]
Roux, Anne-Francoise [3 ]
Tuffery-Giraud, Sylvie [1 ,2 ]
机构
[1] Univ Montpellier I, UFR Med, Montpellier, France
[2] INSERM, U827, Montpellier, France
[3] CHU Montpellier, Hop Arnaud Villeneuve, Genet Mol Lab, Montpellier, France
[4] Innsbruck Med Univ, Dept Med Genet Mol & Clin Pharmacol, Div Clin Genet, Innsbruck, Austria
关键词
Usher genes; MYO7A; CDH23; USH1C; USH2A; splicing; minigenes; IN-SILICO ANALYSIS; SYNDROME TYPE-II; NONSYNDROMIC DEAFNESS; ANALYSIS REVEALS; USH2A MUTATIONS; RNA; PREDICTION; SPECTRUM; SEQUENCE; IDENTIFICATION;
D O I
10.1002/humu.21193
中图分类号
Q3 [遗传学];
学科分类号
071007 ; 090102 ;
摘要
Molecular diagnosis in Usher syndrome type 1 and 2 patients led to the identification of 21 sequence variations located in noncanonical positions of splice Sites in MYO7A, CDH23, USH1C, and USH2A genes. To establish experimentally the splicing pattern of these substitutions, whose impact on splicing is not always predictable by available softwares, ex vivo splicing assays were performed. The branch-point mapping strategy was also used to investigate further a putative branch-point mutation in USH2A intron 43. Aberrant splicing was demonstrated for 16 of the 21 (76.2%) tested sequence variations. The mutations resulted more frequently in activation of a nearby cryptic splice site or use of a de novo splice site than exon skipping (37.5%). This study allowed the reclassification as splicing mutations of one silent (c.7872G>A (p.Glu2624Glu) in CDH23) and four missense mutations (c.2993G>A (p.Arg998Lys) in USH2A, c.592G>A (p.Ala198Thr), c.3503G>C [p.Arg1168Pro], c.5944G>A (p.Gly1982Arg) in MYO7A), whereas it provided clues about a role in structure/function in four other cases: c.802G>A (p.Gly268Arg), c.653T>A (p.Val218Glu) (USH2A), and c.397C>T (p.His133Tyr), c.3502C>T (p.Arg1168Trp) (MYO7A). Our data provide insights into the contribution of splicing mutations in Usher genes and illustrate the need to define accurately their splicing outcome for diagnostic pin-poses. Hum Mutat 31:347-355, 2010. (C) 2010 Wiley-Liss, Inc.
引用
收藏
页码:347 / 355
页数:9
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