Deciphering Planar Cell Polarity (PCP) Pathway In A Congenital Scoliosis Cohort

BackgroundCongenital scoliosis(CS)is defined as a three-dimensional deformity over 10 degrees of lateral curvature caused by vertebral malformation.It occurs about 0.5?1/1000 in live births.CS is characterized by various types of vertebral malformations;however,it's frequently involved in multi-systems disorders,such as neural tube defects(NTDs)and congenital heart disease.It is difficult to make an early diagnosis before progression,and troublesome to treat this kind of patient,both of which have a great impact on individuals,families and society.The pathogenesis of CS is complex.CS is believed to be etiologically caused by multifactorial factors.A multitude of molecular signaling pathways and diverse factors are suggested to participate in the pathogenesis.Until now,it remains unclear how these pathways and factors contribute to CS.However,genetic factors have been largely demonstrated as one of the most important etiologies recently.Planar cell polarity(PCP)pathway,a non-canonical WNT pathway,regulates planar cell polarity,and it may be a pathogenic pathway for CS.VANGL1 and VANGL2 are transmembrane receptors in PCP pathway.Scoliosis has been observed in some of NTDs patients caused by VANGL mutations and Vangl2Lp/Lp mice show vertebral malformations.VANGL1 and VANGL2 might be causative genes of CS.It is widely accepted that whole exome sequencing(WES)is able to identify causative genes.However,there are no studies regarding the evaluation of pathogenesis of PCP pathway in CS.ObjectiveTo evaluate the pathogenesis of PCP pathway in CS.MethodsWe conducted WES in 464 subjects with CS in DISCO(Deciphering Disorders Involving Scoliosis and COmorbidities)cohort and 616 subjects without soliosis,followed by bioinformatics analyses and manual curation.All the causative variants were validated by Sanger sequencing.We also investigated the correlation between VANGL1/2 and phenotype of CS.Moreover,the expression and stability of protein were validated by in vitro functional tests in HEK393T cells.ResultsSeven rare(MAF<1%)missense variants(Arg214Trp,GIu281Gly,Asn313Ser,Arg334Trp,Arg443Trp,Val491lle,Arg517His)were found in VANGL1 in 13 patients.Six rare missense variants(Arg35Gln,Val99Ala,Leu226Phe,Val297lle,Ala299Val,Arg482Cys)were identified in VANGL2 in six patients.What's more,three patients were found to be affected by two PCP genes,VANGL1 and ROR2,fitting a digenic model.In the control group,two VANGL1 missense variants were identified,including three individuals with Arg214Trp and two individuals with Asn313Ser respectively.The incidence of neural tube defects in CS patients with VANGL1 and VANGL2 mutations was significantly higher than that in the non-mutant group through genotype and phenotype association analysis(OR = 3.05,P = 0.026).Furthermore,the number of vertebral malformations in the group with VANGL1 and VANGL2 mutations were significantly correlated with pathogenesis of variants(P = 0.022).The in vitro functional tests demonstrated that the expression of VANGL1(GIu281Gly,Arg443Trp)and VANGL2(Leu226Phe,Ala299Val)proteins were significantly decreased and that VANGL1(Glu281Gly,Arg443Trp)proteins were unstable,and could be easily degraded by proteasome.ConclusionsThe PCP pathway may contribute a lot to CS.VANGL1 and VANGL2 may be candidate genes for CS.The phenotype of CS with VANGL1 and VANGL2 mutations has both commonalities and characteristics.This study provides further evidence supporting the role of VANGL1/2 in CS.However,the pathogenesis of VANGL1/2 in deciphering the genetic complexity of CS is still unclear and further study is required.