Abnormal function of EPHA2/p.R957P mutant in congenital cataract

AIM:To identify genetic defects in a Chinese family with congenital posterior polar cataracts and assess the pathogenicity.METHODS:A four-generation Chinese family affected with autosomal dominant congenital cataract was recruited.Nineteen individuals took part in this study including 5 affected and 14 unaffected individuals.Sanger sequencing targeted hot-spot regions of 27 congenital cataract-causing genes for variant discovery.The pathogenicity of the variant was evaluated by the guidelines of American College of Medical Genetics and InterVar software.Confocal microscopy was applied to detect the subcellular localization of fluorescence-labeled ephrin type-A receptor 2(EPHA2).Co-immunoprecipitation assay was implemented to estimate the interaction between EphA2 and other lens membrane proteins.The mRNA and protein expression were analyzed by reverse transcription-polymerase chain reaction(qRT-PCR)and Western blotting assay,respectively.The cell migration was analyzed by wound healing assay.Zebrafish model was generated by ectopic expression of human EPHA2/p.R957P mutant to demonstrate whether the mutant could cause lens opacity in vivo.RESULTS:A novel missense and pathogenic variant c.2870G>C was identified in the sterile alpha motif(SAM)domain of EPHA2.Functional studies demonstrated the variant’s impact:reduced EPHA2 protein expression,altered subcellular localization,and disrupted interactions with other lens membrane proteins.This mutant notably enhanced human lens epithelial cell migration,and induced a central cloudy region and roughness in zebrafish lenses with ectopic expression of human EPHA2/p.R957P mutant under differential interference contrast(DIC)optics.CONCLUSION:Novel pathogenic c.2870G>C variant of EPHA2 in a Chinese congenital cataract family contributes to disease pathogenesis.

A functional missense variant in ITIH3 affects protein expression and neurodevelopment and confers schizophrenia risk in the Han Chinese population

The Psychiatric Genomics Consortium(PGC)has recently identified 10 potential functional coding variants for schizophrenia.However,how these coding variants confer schizophrenia risk remains largely unknown.Here,we investigate the associations between eight potential functional coding variants identified by PGC and schizophrenia in a large Han Chinese sample(n=4022 cases and 9270 controls).Among the eight tested single nucelotide polymorphisms(SNPs),rs3617(a missense variant,p.K315 Q in the ITIH3 gene)showed genome-wide significant association with schizophrenia in the Han Chinese population(P=8.36×10-16),with the same risk allele as in PGC.Interestingly,rs3617 is located in a genomic region that is highly evolutionarily conserved,and its schizophrenia risk allele(C allele)was associated with lower ITIH3 mRNA and protein expression.Intriguingly,mouse neural stem cells stably overexpressing ITIH3 with different alleles of rs3617 exhibited significant differences in proliferation,migration,and differentiation,suggesting the impact of rs3617 on neurodevelopment.Subsequent transcriptome analysis found that the differentially expressed genes in neural stem cells stably overexpressing different alleles of rs3617 were significantly enriched in schizophrenia-related pathways,including cell adhesion,synapse assembly,MAPK and PI3 K-AKT pathways.Our study provides convergent lines of evidence suggesting that rs3617 in ITIH3 likely affects protein function and neurodevelopment and thereby confers risk of schizophrenia.

Eight novel mutations in the ABCD1 gene and clinical characteristics of 25 Chinese patients with X-linked adrenoleukodystrophy

Background:X-linked adrenoleukodystrophy(X-ALD)is a fatal neurodegenerative disease caused by mutations in the adenosine triphosphate-binding cassette D1(ABCD1)gene.This study aimed to retrospectively investigate the clinical characteristics of 25 patients with X-ALD including members of large pedigrees,to analyze ABCD1 gene mutations,the effect of gene novel variants on ALD protein(ALDP)structure and function,and to expand gene mutation spectrum of Chinese patients.Methods:Twenty-five male patients diagnosed with X-ALD were enrolled in this study.The clinical characteristics of the patients were retrospectively summarized by reviewing medical records or telephone consultation.ABCD1 gene mutations were analyzed.The pathogenicity of novel missense variants was analyzed using cobalt constraint-based multiple protein alignment tool,polymorphism phenotyping,sorting intolerant from tolerant,Align-Grantham variation and Grantham deviation,and Swiss-Program Database Viewer 4.04 software,respectively.Results:Childhood cerebral form ALD(CCALD)is the most common phenotype(64%)in the 25 patients with X-ALD.The progressive deterioration of neurological and cognitive functions is the main clinical feature.The demyelination of the brain white matter and elevated plasma very long chain fatty acids(VLCFAs)were found in all patients.Different phenotypes were also presented within family members of the patients.Twenty-two different mutations including 8 novel mutations in the ABCD1 gene were identifi ed in the 25 patients.Of the mutations,63.6%were missense mutations and 34.8%located in exon 1.The amino acid residues of three novel missense mutations in eight species were highly conserved,and were predicted to be"probably"damaging to ALDP function.The other five novel mutations were splice,nonsense,deletion or duplication mutations.Conclusions:CCALD is the most common phenotype(64%)in our patients with X-ALD.Eight novel mutations in the ABCD1 gene identifi ed are disease-causing mutations.Brain magnetic resonance imaging and plasma VLCFA determination should be performed for the patients who present with progressive deterioration of neurological development.