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Amyotrophic lateral sclerosis (ALS) is a fatal adult-onset neurodegenerative disease characterised by the degeneration of both upper and lower motor neurons. Although most cases occur as sporadic ALS (sALS), approximately 5%–10% of patients with ALS have a family history of ALS (fALS).1
More than 20 potential genes have been identified as ALS-related genes so far, and the function of these causative genes can be loosely grouped into three categories: RNA metabolism, cytoskeletal dynamics and protein homeostasis.1 However, genetic heterogeneity among different ethnic background exists.
The kinesin family member 5A (KIF5A), composed of an N-terminal motor domain, a coiled-coil stalk region and a C-terminal cargo binding domain, is a member of the kinesin superfamily proteins (KIFs) and is involved in the axonal transport.2 Recently, two independent studies found that splice site mutations in the C-terminal cargo binding domain of KIF5A can cause fALS in the Caucasian ALS population,3 4 and the functional studies elucidated that splice site mutations can cause exon skipping and lead to the loss of RNA expression, which suggest that haploinsufficiency is the most likely underlying molecular mechanism.3 4 In this context, we aimed to reveal the mutation frequency of the C-terminal cargo binding domain in KIF5A in a large cohort of Chinese patients with ALS.
A total of 960 Chinese patients with sALS and 37 patients with fALS diagnosed based on the El Escorial revised criteria for definite or probable ALS were enrolled. All patients were excluded for other known common ALS/FTD causative genes, including C9orf72, SOD1, TARDBP, FUS, PFN1, SQSTM1 and CHCHD10. The C-terminal cargo binding domain of KIF5A (exons 24–28) was directly sequenced by Tsingke Corporation (Chengdu, China) using the ABI 3100 automated DNA sequencing system. Then, variants discovered in patients with ALS were confirmed in 500 unrelated healthy controls (HCs). Written and signed informed consent was obtained from all the participants. The primer sequences and PCR condition are listed in online supplementary table 1.
The clinical data of patients with ALS and HCs are shown in online supplementary table 2. In total, five variants (including one synonymous variant) were identified in patients with ALS and were absent in the HCs. Detailed features of these variants are listed in table 1 and online supplementary figure 1.
Two missense mutations—p.V922I and p.F1023C (rs756348570)—were absent in our HCs and were predicted to be disease causative mutations by in silico prediction tools. Additionally, they are both located in the C-terminal cargo binding domain of KIF5A, which is a crucial domain in the pathogenesis of ALS, and the amino acids V and F are highly conserved among species (online supplementary figure 2). Therefore, the p.V922I and p.F1023C are concluded to have uncertain significance but tend to be pathogenic based on the ‘Standards and Guidelines for the Interpretation of Sequence Variants’.
The two variants in the intronic region—c.2993–3 C>T and c.3020+2 T>A—located adjacent to exon 27 were found in two patients with sALS who had a late age of onset, and they were both predicted to be splice site mutations. Moreover, these two mutations were absent in the HCs and in the public database. Therefore, both of them met the requirement for very strong evidence of pathogenicity and one strong evidence of pathogenicity, and were concluded to be pathogenic.
A synonymous variant c.2718C>T [p.Tyr906=] was identified in a male patient with sALS. This mutation did not change the amino acid, so we predicted it to be non-pathogenic.
Additionally, no mutation was found in the 37 patients with fALS.
KIF5A, belonging to the KIFs, is mainly expressed in the neurons. Missense mutations in the N-terminal globular motor domain and central stalk domain have been identified to be associated with autosomal dominant hereditary spastic paraplegia (HSP) and Charcot-Marie-Tooth disease type 2,5 while splice site mutations in the C-terminal binding domain of KIF5A were found in fALS.3 4
In our study, we identified four mutations in the C-terminal cargo binding domain of KIF5A among 960 Chinese patients with sALS. KIF5A mutation accounted for 0.41% (4/960) of Chinese patients with sALS.
Interestingly, missense mutation c.3068 T>G [p.F1023C] has been reported to be associated with autosomal dominant HSP by Illumina Clinical Services Laboratory previously (www.ncbi.nlm.nih.gov/clinvar/RCV000263091/). However, our patient carrying the p.F1023C mutation exhibited classical clinical and electrophysiological manifestations of ALS and died of respiratory failure at 7-month disease duration. Therefore, the p.F1023C mutation might be associated with both ALS and HSP; further studies are needed to ascertain the mechanism. Two splice site mutations found in the current study were also described in Caucasian ALS population,3 which suggests that these splice site mutations may be a common cause for ALS both in the Caucasian and Chinese populations. Among these patients carrying the C-terminal cargo binding domain in KIF5A reported in our study and previous studies,3 4 most of them showed a late age of onset, except three patients who had an age of onset younger than 40 years old. Interestingly, the patients carrying the C-terminal binding domain mutations in the Caucasian population exhibited a longer survival time than patients in our population. This discrepancy can be explained by the difference in the genetic background.
Unlike the previous studies in Caucasian population,3 4 we failed to identify any disease-causing mutations in the fALS cohort, which may be due to the small sample size of our fALS cohorts. Further studies with a larger sample size are needed. Additionally, further studies from different ethnic populations are also necessary.
This is the first and largest study exploring the mutation frequency and clinical features of the C-terminal cargo binding domain of the KIF5A in Chinese patients with ALS. KIF5A mutation accounted for 0.41% (4/960) of Chinese patients with sALS, which suggests that KIF5A is an uncommon cause of ALS in the Chinese population. However, our findings expanded the genotypic and phenotypic spectrum of the C-terminal cargo binding domain of KIF5A in ALS, and further provided evidence of the important role of C-terminal cargo binding domain in KIF5A in the pathogenesis of ALS.
XG and CL contributed equally.
Contributors XG, CL and H-FS designed the study. XG, CL and YC performed the experiments and data analysis. XG and CL wrote the manuscript and H-FS revised the manuscript. QW, BC, RO, XY, YH, LZ, HL, YW, WS, BZ and XC collected clinical information for all the participants.
Funding This work was supported by the National Science Fund of China (grant no 81571247) and the National Key Research and Development Program of China (grant no 2016YFC0901504).
Competing interests None declared.
Patient consent Not required.
Ethics approval The study was approved by the Ethics Committee of Sichuan University.
Provenance and peer review Not commissioned; externally peer reviewed.
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