Targeted Next-Generation Sequencing of a 12.5 Mb Homozygous Region Reveals ANO10 Mutations in Patients with Autosomal-Recessive Cerebellar Ataxia

Vermeer, Sascha and Hoischen, Alexander and Meijer, Rowdy PP and Gilissen, Christian and Neveling, Kornelia and Wieskamp, Nienke and de Brouwer, Arjan and Koenig, Michel and Anheim, Mathieu and Assoum, Mirna and Drouot, Nathalie and Todorovic, Slobodanka and Milic-Rasic, Vedrana and Lochmüller, Hanns and Stevanin, Giovanni and Goizet, Cyril and David, Albert and Durr, Alexandra and Brice, Alexis and Kremer, Berry and van de Warrenburg, Bart PC and Schijvenaars, Mascha MVAP and Heister, Angelien and Kwint, Michael and Arts, Peer and van der Wijst, Jenny and Veltman, Joris and Kamsteeg, Erik-Jan and Scheffer, Hans and Knoers, Nine (2010) Targeted Next-Generation Sequencing of a 12.5 Mb Homozygous Region Reveals ANO10 Mutations in Patients with Autosomal-Recessive Cerebellar Ataxia. American Journal of Human Genetics, 87 (6). p. 813. ISSN 0002-9297 (FP7- 223143)

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Abstract

Autosomal-recessive cerebellar ataxias comprise a clinically and genetically heterogeneous group of neurodegenerative disorders. In contrast to their dominant counterparts, unraveling the molecular background of these ataxias has proven to be more complicated and the currently known mutations provide incomplete coverage for genotyping of patients. By combining SNP array-based linkage analysis and targeted resequencing of relevant sequences in the linkage interval with the use of next-generation sequencing technology, we identified a mutation in a gene and have shown its association with autosomal-recessive cerebellar ataxia. In a Dutch consanguineous family with three affected siblings a homozygous 12.5 Mb region on chromosome 3 was targeted by array-based sequence capture. Prioritization of all detected sequence variants led to four candidate genes, one of which contained a variant with a high base pair conservation score (phyloP score: 5.26). This variant was a leucine-to-arginine substitution in the DUF 590 domain of a 16K transmembrane protein, a putative calcium-activated chloride channel encoded by anoctamin 10 (ANO10). The analysis of ANO10 by Sanger sequencing revealed three additional mutations: a homozygous mutation (c.1150_1151del [p.Leu384fs]) in a Serbian family and a compoundheterozygous splice-site mutation (c.1476þ1G>T) and a frameshift mutation (c.1604del [p.Leu535X]) in a French family. This illustrates the power of using initial homozygosity mapping with next-generation sequencing technology to identify genes involved in autosomalrecessive diseases. Moreover, identifying a putative calcium-dependent chloride channel involved in cerebellar ataxia adds another pathway to the list of pathophysiological mechanisms that may cause cerebellar ataxia.

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