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

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 compound-heterozygous splice-site mutation (c.1476+1G>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 autosomal-recessive 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.

A Coding Variant of ANO10, Affecting Volume Regulation of Macrophages, Is Associated with Borrelia Seropositivity

“…Notably, mutations in ANO10 were found to cause spinocerebellar ataxia (25,26), which is also a reported phenotype of neuroborreliosis (43). Similar to R263H, these mutations also inhibited I Hypo in our present report.…”

“…Interestingly, both variants, located on chromosomes 3 and 16, happen to be in some context with spinocerebellar ataxia (25,26). The SNP on chromosome 3, rs41289586, represents the missense variant ANO10-R263H, encoded by the gene ANO10.…”

“…This may, however, be a bias of the small number of individuals included there ( (23,24). Notably, also mutations in ANO10 were reported to be causative for spinocerebellar ataxia (25,26). One of the two genome-wide significant SNPs, rs17850869, is a synonymous coding variant of zinc finger protein 821, encoded by the ZNF821 gene on chromo-We investigated, but did not find, an association of either SNP with antibodies against several other bacterial infections (Helicobacter pylori, Mycoplasma pneumoniae, Chlamydia pneumoniae, Chlamydia trachomatis).…”

Exposure of phosphatidylserine on the cell surface

“…and Savarese et al 130 for TMEM16E, from Suzuki et al 40 and Castoldi et al 93 for TMEM16F, and Vermeer et al 131 and Balreira et al 132 for TMEM16K Figure 3 The molecular mechanism for PtdSer exposure in cells with high Ca 2+ -concentration. The flippase comprised of P4-ATPase (ATP11A or ATP11C) and CDC50A, and a Ca 2+ -dependent scramblase (TMEM16F) are schematically shown.…”

Functional Swapping between Transmembrane Proteins TMEM16A and TMEM16F

“…In addition, abnormalities (mutations and overexpression) in TMEM16 family members are implicated in a number of human diseases (6). For example, genetic mutations in TMEM16C, -16E, -16F, and -16K are associated with craniocervical dystonia (14), musculoskeletal disorder (15,16), bleeding disorder (9), and cerebellar ataxia (17), respectively. TMEM16A is overexpressed in human gastrointestinal stromal tumors and in head and neck squamous carcinoma; TMEM16G is overexpressed in prostate cancer (18,19).…”