Exome Sequencing and Disease-Network Analysis of a Single Family Implicate a Mutation in KIF1A in Hereditary Spastic Paraparesis
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Downloaded from genome.cshlp.org on September 28, 2021 - Published by Cold Spring Harbor Laboratory Press Research Exome sequencing and disease-network analysis of a single family implicate a mutation in KIF1A in hereditary spastic paraparesis Yaniv Erlich,1,4,5 Simon Edvardson,2,4 Emily Hodges,3 Shamir Zenvirt,2 Pramod Thekkat,3 Avraham Shaag,2 Talya Dor,2 Gregory J. Hannon,3 and Orly Elpeleg2 1Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA; 2Monique and Jacques Roboh Department of Genetic Research, Department of Genetic and Metabolic Diseases, Hadassah, Hebrew University Medical Center, 91120 Jerusalem, Israel; 3Watson School of Biological Sciences, Howard Hughes Medical Institute, Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 11724, USA Whole exome sequencing has become a pivotal methodology for rapid and cost-effective detection of pathogenic vari- ations in Mendelian disorders. A major challenge of this approach is determining the causative mutation from a sub- stantial number of bystander variations that do not play any role in the disease etiology. Current strategies to analyze variations have mainly relied on genetic and functional arguments such as mode of inheritance, conservation, and loss of function prediction. Here, we demonstrate that disease-network analysis provides an additional layer of information to stratify variations even in the presence of incomplete sequencing coverage, a known limitation of exome sequencing. We studied a case of Hereditary Spastic Paraparesis (HSP) in a single inbred Palestinian family. HSP is a group of neuro- pathological disorders that are characterized by abnormal gait and spasticity of the lower limbs. Forty-five loci have been associated with HSP and lesions in 20 genes have been documented to induce the disorder. We used whole exome se- quencing and homozygosity mapping to create a list of possible candidates. After exhausting the genetic and functional arguments, we stratified the remaining candidates according to their similarity to the previously known disease genes. Our analysis implicated the causative mutation in the motor domain of KIF1A, a gene that has not yet associated with HSP, which functions in anterograde axonal transportation. Our strategy can be useful for a large class of disorders that are characterized by locus heterogeneity, particularly when studying disorders in single families. [Supplemental material is available for this article. The sequencing and genotyping data from this study have been submitted to the NCBI dbGaP (http://www.ncbi.nlm.nih.gov/gap) and at http://cancan.cshl.edu/hsp/.] Whole exome sequencing has ushered in a renaissance in identi- protein, either by analyzing biochemical and structural features fying pathogenic variations in monogenic diseases. The approach (Adzhubei et al. 2010) or by measuring the extent of multispecies enables a rapid and cost-effective detection from a small number of conservation at the variation site (Cooper et al. 2010). individuals, and has proved useful for a wide range of clinical Monogenic disorders in isolated inbred families cast a unique settings (Choi et al. 2009; Ng et al. 2009, 2010a, 2010b; Krawitz setting for identifying the causative variations. Homozygosity et al. 2010; Pierce et al. 2010; Walsh et al. 2010). A major challenge mapping can quickly identify regions that are identical by decent, of whole exome sequencing is determining the causative mutation refining the search area to regions that typically span several from a substantial number of bystander variations that do not play megabases (Lander and Botstein 1987). However, this setting a role in the disease etiology. The common strategy to filter out the brings several challenges. First, bystander variations inside the bystander variations is based on a systematic rejection of variations homozygous region are (almost) always homozygous; therefore, according to genetic and functional arguments, narrowing down they are not amenable for rejection based on mode of inheritance the candidate list until the causative variation is isolated. One class (Choi et al. 2009; Walsh et al. 2010). Second, the disease-harboring of genetic arguments that has been widely used is rejection of region is identical between the affected siblings and sequencing variations that are not shared between multiple cases or that do not multiple cases from the same family adds minimal information follow the assumed mode of inheritance (Ng et al. 2009, 2010b; beyond the data from the homozygosity mapping. Third, inbred Krawitz et al. 2010). Another class of genetic arguments asserts that unions are more prevalent in non-Western societies, such as North harmful mutations must be rare due to purifying selection and Africa, the Middle East, and Central Asia (Bittles 2001). These accordingly reject variations that have been catalogued in dbSNP ethnic groups tend to be less represented in variation repositories or 1000 Genomes, as these are assumed to be relatively common. (Carlson et al. 2003; Via et al. 2010), reducing the power of elim- Functional arguments focus on the impact of the variation on the inating common polymorphisms. In this study, we present disease-network analysis as an ad- ditional layer of arguments to stratify variations. Accumulating 4 These authors contributed equally to this work. lines of evidence have shown that genes that are associated with 5 Corresponding author. phenotypically close disorders are prone to have similar molecular E-mail [email protected]. Article published online before print. Article, supplemental material, and pub- signatures (Goh et al. 2007; Feldman et al. 2008; Wu et al. 2008). lication date are at http://www.genome.org/cgi/doi/10.1101/gr.117143.110. These include similar expression profiles, participation in the same 21:000–000 Ó 2011 by Cold Spring Harbor Laboratory Press; ISSN 1088-9051/11; www.genome.org Genome Research 1 www.genome.org Downloaded from genome.cshlp.org on September 28, 2021 - Published by Cold Spring Harbor Laboratory Press Erlich et al. signaling or metabolic pathways, or sharing similar protein do- score of 2. No weakness, wasting, decreased vibration sense, or mains. Thus, it is appealing to reject variations by comparing the pinprick sensation was noted, and there were no sphincter distur- molecular signature of their harboring genes to the signatures of bances or cerebellar signs. Brain MRI in all patients was normal. closely related disease genes. EMG and NCV in II-4 and II-5 were normal. Unlike most of the rejection arguments above, disease-based Based on these results, the patients were diagnosed with arguments do not require sequencing information per se. This a pure type of Hereditary Spastic Paraparesis (HSP). HSP is a group provides a means to exclude positions that were not covered or of genetic disorders resulting in axonal degradation of the corti- miscalled, a common caveat in whole exome sequencing (Ng et al. cospinal tract (Reid 2003; Dion et al. 2009). They are characterized 2010a). Disease-network analysis has been mainly used for gene by a progressive lower-extremity spastic weakness, hypertonic prioritizing as the first step of gene-centric studies, when se- urinary bladder disturbance, mild diminution of lower-extremity quencing was a limiting factor (Ropers 2007). One drawback of this vibration sensation and, occasionally, of joint position sensation. approach is the requirement to prioritize a large number of genes, As of September 2010, 45 HSP loci and 20 HSP-related genes have which increases the amount of false positives. In our approach, the been identified (Supplemental Table 1), showing autosomal dom- analysis is executed at the last stage on a minimal list of genes, inant, autosomal recessive, and X-linked inheritance patterns increasing the specificity of the method. (Dion et al. 2009). HSPs are subdivided into pure and complicated forms, where the complicated forms involve additional patholo- Results gies, such as mental retardation, ocular signs, and skin abnormal- ities. However, some loci are associated with both pure and com- Description of the affected family plicated forms of HSPs (Reid 2003; Dion et al. 2009). We evaluated the performance of our strategy in a case of three Rejecting involvement of known HSP genes brothers aged 20 (Patient II-4), 15 (Patient II-5), and 14 (Patient II-7) or X-linked disorders years, who presented to our clinic with a chief complaint of abnormal gait. The parents were of Muslim Palestinian origin and We determined the complete exonic sequences of patient II-5 and denied consanguinity; however, both originated from the same his parents, using array-based hybrid selection and Illumina se- village. They and the other children were reportedly healthy at 16 quencing (see Methods), and we genotyped the whole genome of to 23 yr old (Fig. 1). The perinatal course and early development of Patient II-4 and Patient II-5, using GeneChip Human Mapping all patients were uneventful and walking appeared at around one 250K Nsp Array of Affymetrix, as previously described (Edvardson year of age. Stiff legs and a slowly progressive gait disturbance be- et al. 2007). came evident at two years, but both participated successfully in We first evaluated the possibility that the disorder is caused by sport activities throughout childhood. Further aggravation of known HSP genes. Patient II-4 and II-5 shared 610 Mbase (23%) of symptoms was noted at 10–13 yr of age and the patients could no identical autosomal segments, and 75 Mbase (50%) of identical longer play soccer or walk long distances. Lower limb sensation, segments on the X chromosome, as expected from full siblings. sphincters, upper limbs, and intellectual functions were intact The shared autosomal segments contained four known HSP genes: throughout childhood. ATL1, GJC2, HSPD1, and SPG20. The sequencing coverage was 99% Patient II-4, when first examined at age 20, was unable to run. for the coding regions of these genes. We could not find any ho- His gait was stiff with equinos-gait, scissoring, and crouching.