Genome-wide Analyses Identify KIF5A as a Novel ALS Gene

Abstract

To identify novel genes associated with ALS, we undertook two lines of investigation. We carried out a genome-wide association study comparing 20,806 ALS cases and 59,804 controls. Independently, we performed a rare variant burden analysis comparing 1,138 index familial ALS cases and 19,494 controls. Through both approaches, we identified kinesin family member 5A (KIF5A) as a novel gene associated with ALS. Interestingly, mutations predominantly in the N-terminal motor domain of KIF5A are causative for two neurodegenerative diseases: hereditary spastic paraplegia (SPG10) and Charcot-Marie-Tooth type 2 (CMT2). In contrast, ALS-associated mutations are primarily located at the C-terminal cargo-binding tail domain and patients harboring loss-of-function mutations displayed an extended survival relative to typical ALS cases. Taken together, these results broaden the phenotype spectrum resulting from mutations in KIF5A and strengthen the role of cytoskeletal defects in the pathogenesis of ALS.

Keywords: ALS; GWAS; KIF5A; WES; WGS; axonal transport; cargo.

Conflict of interest statement

Declaration of Interests: J.D.Be. is a consultant to Neuraltus Pharmaceuticals and Denali Therapeutics, and held a research fellow position funded by Voyager Therapeutics. M.C. has been a consultant for Eli Lilly and Company, Mitsubishi Tanabe Pharma America (MT Pharma America), Denali Therapeutics, Karyopharm Therapeutics and Cytokinetics. S.A.G. has served as a consultant and received research support from Cytokinetics. O.H. has received speaking honoraria from Novarits, Biogen Idec, Sanofi Aventis and Merck-Serono and has been a member of advisory panels for Biogen Idec, Allergen, Ono Pharmaceuticals, Novartis, Cytokinetics and Sanofi Aventis. O.H. serves as Editor-in-Chief of Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. L.H.v.d.B. serves on scientific advisory boards for the Prinses Beatrix Spierfonds, Thierry Latran Foundation, Biogen and Cytokinetics. Serves on the editorial board of Amyotrophic Lateral Sclerosis And Frontotemporal Degeneration and The Journal of Neurology, Neurosurgery, and Psychiatry. J.H.V. reports that his institute received consultancy fees from Vertex Pharmaceuticals. A.C. serves on scientific advisory boards for Biogen Idec, Cytokinetics, Italfarmaco, and Neuraltus. P.M.A. serve on advisory board panels for Biogen and Orphazyme. V.S. serves as a consultant for Cytokinetics.

Copyright © 2018 Elsevier Inc. All rights reserved.

Figures

Figure 1. Identification of association between KIF5A locus and ALS risk through GWAS

(A) Manhattan plot showing P values from the discovery set GWAS. Analysis of a combined set of 20,806 cases and 59,804 controls is shown. The dashed red line denotes the threshold for genome-wide significance after multiple test correction (P < 5.0×10−8). Five previously reported ALS associated loci are labeled in grey and one novel loci, containing theKIF5A gene, is labeled in black. (B) Regional association plot of the KIF5A locus. Recombination rates are from HapMap phase 2 European ancestry samples. The R2 pattern is based on the rs113247976 SNP using 85 European ancestry samples (CEU) from the November 2010 release of the 1000 Genomes Project dataset. R2 of the p.Pro986Leu (rs113247976) with additional SNPs achieving genome-wide significance was 0.544 (rs117027576), 0.544 (rs118082508), 0.741 (rs116900480), and 0.347 (rs142321490).

Figure 2. Discovery and replication for the association of the KIF5A p.Pro986Leu (rs113247976) variant with ALS

Analysis of the p.Pro986Leu (rs113247976) variant within each of the described cohorts is shown. Allelic association for all subcohorts were analyzed by logistic regression followed by a fixed-effects meta-analysis. The Forest plot (right) displays the distribution of OR estimates across study cohorts with the vertical dotted line denoting the OR estimated under the meta-analysis.

Figure 3. Identification of association between KIF5A and ALS risk through rare variant burden analysis of exome sequencing

Manhattan plot showing gene-level P values from an exome-wide rare variant burden analysis. Analyses of 1,138 index FALS cases versus 19,494 controls were restricted to rare LOF variants (splice altering/nonsense, MAF < 0.001). A minimum of 3 LOF gene variants were required for analysis. The dashed red line denotes the threshold for exome-wide significance after correction for 11,472 genes (4.36×10−6). Previously reported (grey) and novel (black) genes exhibiting a significant excess of rare LOF variants in patients are shown.

Figure 4. ALS associated loss of function variants of KIF5A disrupt C-terminal sequence by inducing skipping of exon 27

(A) Single nucleotide variants (SNVs) within KIF5A identified in ALS patients are clustered at the 5′ and 3′ splice junctions of exon 27. The consensus splice sequence is shown. (B) ALS associated SNVs are predicted to induce skipping of exon 27 and result in an aberrant mRNA transcript. (C) The skipping of exon 27 of KIF5A yields an out-of-frame and extended disrupted C-terminal peptide sequence. The amino acids in red signify the divergence from the normal protein. (D) RT-PCR was performed using RNA derived from ALS patients with the indicated LOF variant or without (controls) using primers to either amplify both wild-type (155 bp) and mutant (127 bp) splice forms or specifically the mutant splice form (80 bp, right panel). The arrow represents the position of the mutant specific product. The tick marks represent 200 bp (upper) and 100 bp (lower) markers.

Figure 5. KIF5A ALS mutations show distinct localization from missense mutations previously associated with SPG10 and CMT2

Causative mutations for SPG10 and CMT2 described within the literature (Crimella et al., 2011; Jennings et al., 2017; Liu et al., 2014; Reid et al., 2002) and ALS associated mutations identified within this study are shown. As illustrated, mutations causative for SPG10/CMT2 are predominantly missense changes located in the N-terminal motor domain. In contrast, ALS mutations are primarily located at the C-terminal motor domain and are LOF.