REPORT VAMP1 Mutation Causes Dominant Hereditary Spastic Ataxia in Newfoundland Families Cynthia V. Bourassa, 1,2 Inge A. Meijer, 1,2 Nancy D. Merner, 1,2 Kanwal K. Grewal, 3 Mark G. Stefanelli, 3 Kathleen Hodgkinson, 4 Elizabeth J. Ives, 3 William Pryse-Phillips, 5 Mandar Jog, 6 Kym Boycott, 7 David A. Grimes, 8 Sharan Goobie, 9 Richard Leckey, 10 Patrick A. Dion, 1,2 and Guy A. Rouleau 1,2,11, * Our group previously described and mapped to chromosomal region 12p13 a form of dominantly inherited hereditary spastic ataxia (HSA) in three large Newfoundland (Canada) families. This report identifies vesicle-associated membrane protein 1 (VAMP1), which encodes a critical protein for synaptic exocytosis, as the responsible gene. In total, 50 affected individuals from these families and three independent probands from Ontario (Canada) share the disease phenotype together with a disruptive VAMP1 mutation that affects a critical donor site for the splicing of VAMP1 isoforms. This mutation leads to the loss of the only VAMP1 isoform (VAMP1A) expressed in the nervous system, thus highlighting an association between the well-studied VAMP1 and a neurological disorder. Given the variable phenotype seen in the affected individuals examined here, we believe that VAMP1 should be tested for mutations in patients with either ataxia or spastic paraplegia. Spastic ataxia 1 (SPAX1 [MIM 108600]) is a rare neurode- generative disorder characterized by lower-limb spasticity and ataxia in the form of head jerks, ocular movement abnormalities, dysphagia, dysarthria, and gait distur- bance 1,2 starting at the age of 10–20 years. Other clinical features are supranuclear gaze palsy, hyperreflexia, hyper- tonicity, dystonia, pes cavus, mild ptosis, and decreased vibration sense in the lower limbs. Symptom severity varies between individuals, but neither life span nor cogni- tion is affected. 1,2 This disease resembles both spinocere- bellar ataxias (SCAs [MIM 164400]) and hereditary spastic paraplegias (HSPs [MIM 182600]) but is part of a distinct group: hereditary spastic ataxias (HSAs). Previous reports describing the three Newfoundland (Canada) families affected by SPAX1 had indicated the existence of an ances- trally shared haplotype located in chromosomal region 12p13. 1,2 A single neurologist (M.G.S.) diagnosed most of the family members (Figures S2, S3, and S4A, available online), and blood was collected for DNA analyses. More recently, Dr. Jog ascertained an additional family (Figure S4B) (with a Newfoundland origin [Figure S1]), and other neurologists (K.B., D.A.G., and S.G.) identified three single Ontario cases, all of whom presented with a similar SPAX1 phenotype. The collection of samples and genetic studies were approved by the relevant ethic committees, and informed consent was obtained from all subjects. A mutation screening panel, composed of two affected individuals with the ancestral disease haplotype from two different families 2 (V-3 in Figure S2 and V-11 in Figure S3) and an unaffected control, was established. All coding exons and flanking intron regions of the 53 genes and predicted genes located in the 1.9 Mb disease haplo- type were amplified by PCR. The PCR conditions and primer sets are available upon request. PCR products were sequenced at the McGill University and Ge ´nome Que ´bec Innovation Centre (Montre ´al, Canada), and the sequences were analyzed with Mutation Surveyor v.3.0 (SoftGenetics). We searched for rare variants (those with less than 1% allele frequency) that were exclusively present in the two affected individuals of the mutation screening panel. Variants that were previously reported in dbSNP were excluded from further analysis. We determined the allele frequency of the remaining interesting variants by screening 169 Newfoundland population controls who were previously collected for a regional colorectal cancer study. 3 A single variant of interest was observed in vesicle-asso- ciated membrane protein 1 (VAMP1 [MIM 185880]) at posi- tion chr12: g.6574054T>G(Figure 1A). VAMP1 has three annotated isoforms, VAMP1A (RefSeq accession number NM_014231.3), VAMP1B (RefSeq NM_016830.2), and VAMP1D (RefSeq NM_199245.1), that only differ by their last exon (Figure 1B). The primers amplifying VAMP1 are given in Table S1. The mutation in VAMP1A and VAMP1B is c.340þ2T>G, and the VAMP1D mutation is c.342T>G (p.Ser114Arg). In order to predict the impact of the vari- ants, we used in silico programs (SIFT, 4 PolyPhen, 5 and 1 The Centre of Excellence in Neurosciences, Centre Hospitalier de l’Universite ´ de Montre ´al Research Center, Universite ´ de Montre ´al, Montre ´al, QC H2L 2W5, Canada; 2 Department of Medicine, Universite ´ de Montre ´al, Montre ´al, QC H2L 2W5, Canada; 3 Discipline of Medical Genetics and Division of Neurology, Faculty of Medicine, Health Sciences Center, Memorial University, St. John’s, NL A1B 3V6, Canada; 4 Department of Clinical Epidemiology, Faculty of Medicine, Memorial University, St. John’s, NL A1B 3V6, Canada; 5 Department of Medicine, Faculty of Medicine, Memorial University, St. John’s, NL A1B 3V6, Canada; 6 Department of Clinical Neurological Sciences, London Health Sciences Centre-University Hospital, University of Western Ontario, London, ON N6A 5A5, Canada; 7 Department of Genetics, Children’s Hospital of Eastern Ontario, Ottawa, ON K1H 8L1, Canada; 8 Department of Neurology, Ottawa Hospital, Ottawa, ON K1Y 4E9, Canada; 9 Medical Genetics Program of Southwestern Ontario Children’s Hospital, London Health Sciences Centre-University Hospital, University of Western Ontario, London, ON N6A 5A5, Canada; 10 Department of Medicine, Faculty of Medicine, Dalhousie University, Halifax, NS B3H 2Y9, Canada; 11 Sainte-Justine Hospital, Montre ´al, QC H3T 1C5, Canada *Correspondence: guy.rouleau@umontreal.ca http://dx.doi.org/10.1016/j.ajhg.2012.07.018. Ó2012 by The American Society of Human Genetics. All rights reserved. 548 The American Journal of Human Genetics 91, 548–552, September 7, 2012