Sequencing of neurofilament genes identified NEFH Ser787Arg as a novel risk variant of sporadic amyotrophic lateral sclerosis in Chinese subjects

Abstract

Background Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease with neuronal cell inclusions composed of neurofilaments and other abnormal aggregative proteins as pathological hallmarks. Approximately 90% of patients have sporadic cases (sALS), and at least 4 genes, i.e. C9orf72, SOD1, FUS and TARDBP, have been identified as the main causative genes, while many others have been proposed as potential risk genes. However, these mutations could explain only ~ 10% of sALS cases. The neurofilament polypeptides encoded by NEFH, NEFM, and NEFL are promising protein biomarkers for ALS and other degenerative diseases. However, whether the genetic variants of these genes were associated with ALS remain ambiguous. Methods Here, we used PCR-Sanger to sequence the exons of these three genes in a cohort of 371 sALS patients and 711 healthy controls (Phase I) and validated the risk variant in another 300 sALS patients and 1076 controls (Phase II). Results A total of 92 variants were identified, including 36 rare heterozygous variants in NEFH, 27 in NEFM, and 16 in NEFL, and only rs568759161 (p.Ser787Arg) in NEFH reached nominal statistical power (P = 0.02 at Phase I, P = 0.009 at Phase II) in the case–control comparison. Together, the Phase I and II studies showed the significantly higher frequency of the variant in cases (9/1342, 0.67%) than in controls (2/3574, 0.07%) (OR 12.06; 95% CI 2.60–55.88; P = 0.0003). No variants passed multiple testing in the discovery cohort, but rs568759161 was associated with ALS in a replication cohort. Conclusions Our results confirmed that NEFH Ser787Arg is a novel sALS risk variant in Chinese subjects, but NEFM and NEFL were not associated with sALS. These data may have implications for genetic counselling and for understanding the pathogenesis of sALS.

Intronic NEFH variant is associated with reduced risk for sporadic ALS and later age of disease onset

“…Once validated, it would be interesting to know if the protective effects of the NEFH TTTA variant could reduce the influence of pathogenic variants such as C9orf72, and this should be considered in future studies . Similarly, the DNA samples were not screened for previously reported NEFH exonic variants, considered to be a rare cause of sALS, occurring in ~ 1% of sporadic cases 21 – 27 . Secondly, end-point survival data was only available for the smaller Duke sALS cohort.…”

“…On a genetic level, mutations and polymorphisms in all three NEFH protein domains (head, rod and tail) have been reported to be associated with ALS 21 – 27 , although studies have typically focused on exonic variants. In particular, large insertions or deletions in the tail region of NEFH have been shown to impact phosphorylation sites critical for its interaction and assembly with other smaller neurofilament proteins 21 – 23 , 28 .…”

“…In particular, large insertions or deletions in the tail region of NEFH have been shown to impact phosphorylation sites critical for its interaction and assembly with other smaller neurofilament proteins 21 – 23 , 28 . However, many studies report variants in NEFH as a rare cause of ALS, occurring in only ~ 1% of sporadic cases 21 – 27 . Despite huge advances in the field of ALS genetics, there is still an urgent need for the characterization of genetic markers that may help to underpin disease heterogeneity and phenotypic variation, particularly in sporadic ALS (sALS) patients 3 , 29 , 30 .…”

Intronic NEFH variant is associated with reduced risk for sporadic ALS and later age of disease onset

“…On a genetic level, mutations and polymorphisms in all three NEFH protein domains (head, rod and tail) have been reported to be associated with ALS [21][22][23][24][25][26][27] , although studies have typically focused on exonic variants. In particular, large insertions or deletions in the tail region of NEFH have been shown to impact phosphorylation sites critical for its interaction and assembly with other smaller neuro lament proteins [21][22][23]28 .…”

“…In particular, large insertions or deletions in the tail region of NEFH have been shown to impact phosphorylation sites critical for its interaction and assembly with other smaller neuro lament proteins [21][22][23]28 . However, many studies report variants in NEFH as a rare cause of ALS, occurring in only ~ 1% of sporadic cases [21][22][23][24][25][26][27] . Despite huge advances in the eld of ALS genetics, there is still an urgent need for the characterization of genetic markers that may help to underpin disease heterogeneity and phenotypic variation, particularly in sALS patients 3,29,30 .…”