Coding Variants Coupled With Rapid Modeling in Zebrafish Implicate Dynein Genes, dnaaf1 and zmynd10, as Adolescent Idiopathic Scoliosis Candidate Genes

Abstract

Adolescent idiopathic scoliosis (AIS) is the most common pediatric spine disorder affecting ∼3% of children worldwide. Human genetic studies suggest a complex polygenic disease model for AIS with large genetic and phenotypic heterogeneity. However, the overall genetic etiology of AIS remains poorly understood. To identify additional AIS susceptibility loci, we performed whole-exome sequencing (WES) on a cohort of 195 Southern Chinese AIS patients. Bioinformatics analysis identified 237 novel rare variants associated with AIS, located in 232 new susceptibility loci. Enrichment analysis of these variants revealed 10 gene families associated with our AIS cohort. We screened these gene families by comparing our candidate gene list with IS candidate genes in the Human Phenotype Ontology (HPO) database and previous reported studies. Two candidate gene families, axonemal dynein and axonemal dynein assembly factors, were retained for their associations with ciliary architecture and function. The damaging effects of candidate variants in dynein genes dnali1, dnah1, dnaaf, and zmynd10, as well as in one fibrillin-related gene tns1, were functionally analyzed in zebrafish using targeted CRISPR/Cas9 screening. Knockout of two candidate genes, dnaaf1 or zmynd10, recapitulated scoliosis in viable adult zebrafish. Altogether, our results suggest that the disruption of one or more dynein-associated factors may correlate with AIS susceptibility in the Southern Chinese population.

The orthopedic characterization of cfap298tm304 mutants validate zebrafish to faithfully model human AIS

“…While scoliosis occured rarely in quadrupedal animals, the longitudinal shape of zebrafish coupled with their ability to experience spinal loads in water make them naturally sensitive to three-dimensional spine deformities, which they even naturally develop with elderliness, highly similarly to humans 15 , 16 . Recent studies have demonstrated a link between spinal curves and cilia motility involved in cerebrospinal fluid (CSF) circulation 11 – 13 , 17 , 18 , as mutants defective in cilia motility develop AIS phenotypes. A cilium-linked physiopathology in AIS patients is highly suspected, as mutations related to cilium has been found in AIS cohorts (POC5, PAX1) and due to the correlation between CSF-flow and AIS illustrated by the higher prevalence of scoliosis in AIS patients 19 – 22 .…”

Identification of potential pathogenic genes causing occipitalization of the atlas

“…Cowden syndrome is a genetic condition characterized by a large head, hamartomatous lesions affecting derivatives of ectodermal, mesodermal, and endodermal layers, macrocephaly, facial trichilemmomas, and scoliosis [24,25]. The association of TNS1 with AO or KFS has not been previously reported, thus the present ndings may represent a novel disease association [26].…”

“…Cowden syndrome is a genetic condition characterized by a large head size, facial trichilemmomas, and scoliosis. Wang et al reported that the TNS1 mutation may result in adolescent scoliosis in 2020 [26]. Mutations in FRFR2 have been previously associated with Pfeiffer and Crouzon syndromes.…”

Turning the Curve Into Straight: Phenogenetics of the Spine Morphology and Coordinate Maintenance in the Zebrafish

“…In the recent years, several studies have focused on discovering the genetic variants associated with AIS, either by identifying mutations that segregate with the disease or polymorphic loci in affected population studies. Thus, genome-wide association studies (GWAS) and whole exome sequencing (WES) have greatly contributed to understanding the etiology of AIS by identifying single nucleotide polymorphisms (SNPs) and gene variants in several loci, respectively ( Gao et al, 2007 ; Takahashi et al, 2011 ; Kou et al, 2013 ; Baschal et al, 2014 ; Ogura et al, 2015 ; Patten et al, 2015 ; Sharma et al, 2015 ; Zhu et al, 2015 ; Zhu et al, 2017 ; Khanshour et al, 2018 ; Kou et al, 2019 ; Liu et al, 2019 ; Wang et al, 2020 ; Terhune et al, 2021b ). These efforts have been pivotal to characterize genotype-AIS associations and expand our understanding of the genetic regulation behind scoliosis in humans.…”

“…Therefore, and considering that humans are unsuitable for robust functional interrogation of the largely unknown functions of AIS-associated genes at large scale, there is a need for model systems that enable the generation of mutants to study genotype-phenotype interactions. Indeed, the function of AIS risk genes identified by GWAS and WES studies, including LBX1, DNAAF1, ZMYND10, MAPK7, KIF6, and PAX1, has successfully been studied in a vertebrate mutant system ( Figure 1A ) ( Adham et al, 2005 ; Guo et al, 2016 ; Gao et al, 2017 ; Konjikusic et al, 2018 ; Wang et al, 2020 ).…”