目的:ARL3(ADP-核糖基化因子样3)变异导致常染色体显性视网膜色素变性(RP)或常染色体隐性Joubert综合征。我们发现了一个具有视锥营养不良(RCD)的家族,并证实它与ARL3基因中的复合杂合变体有关。方法:进行眼科检查,包括光学相干断层扫描和视网膜电图(ERG)。使用定制设计的小组对先证者进行靶向下一代测序(NGS)。在家庭成员中进行Sanger测序和共分离。体外研究了由变体介导的蛋白质结构的变化。通过环己酰亚胺追踪测定和共免疫沉淀(Co-IP)测定评估ARL3蛋白稳定性及其与RP2蛋白的相互作用。结果:18岁男性先证者右眼视力为0.25,左眼视力为0.20,而他的非血亲父母和妹妹是正常的。先证者有刚果民盟的迹象,包括夜视,外围场损耗,视网膜中的骨针沉积,并减少ERG响应。父亲,50岁,两只眼睛的视力均为1.0。与先证者不同,他表现为迟发性和轻度锥杆营养不良(CRD),包括黄斑萎缩,中央阴囊,适光ERG反应的适度减少。所有家庭成员都没有听力异常,精神发育不良或步态不稳定。我们鉴定了两个新的复合杂合变体(c.91A>G,p.T31A;c.358G>T,p.C118F)在先证者的ARL3中,而他的父亲只有变异c.91A>G。生物信息学分析表明,两种变体的氨基酸位置在物种之间高度保守。模拟工具预测变体是有害的。蛋白质结构分析显示这两种变体具有改变蛋白质结构的潜力。根据ACMG指南,这两个变种可能是致病的。此外,ARL3突变使ARL3蛋白不稳定,突变c.358G>T破坏了HEK293T细胞中ARL3和RP2之间的相互作用。结论:我们发现ARL3中的新型复合杂合变体与常染色体隐性遗传RCD的早期发作有关,而c.91A>Galong可能与显性CRD的晚发作有关。ARL3中的两个变体可能是通过使ARL3蛋白不稳定并损害其与RP2蛋白的相互作用而引起的。
Purpose:
ARL3 (ADP-ribosylation factor-like 3) variants cause autosomal dominant retinitis pigmentosa (RP) or autosomal recessive Joubert syndrome. We found a family with rod-cone dystrophy (RCD) and verified it was associated with compound heterozygous variants in
ARL3 gene. Methods: Ophthalmic examinations including optical coherence tomography and electroretinogram (ERG) were performed. Targeted next generation sequencing (NGS) was performed for the proband using a custom designed panel. Sanger sequencing and co-segregation were conducted in the family members. Changes of protein structure mediated by the variants were studied in vitro. ARL3 protein stability and its interaction with RP2 protein were assessed by cycloheximide chase assay and co-immunoprecipitation (Co-IP) assay. Results: Visual acuity of the 18-year-old male proband was 0.25 in the right and 0.20 in the left eye, while his non-consanguineous parents and sister was normal. The proband showed signs of RCD, including nyctalopia, peripheral field loss, bone-spicule deposits in the retina, and reduced ERG responses. The father, aged 50 years old, showed visual acuity of 1.0 in both eyes. Unlike the proband, he presented late onset and mild cone-rod dystrophy (CRD), including macular atrophy, central scotomata, moderate reduction in photopic ERG responses. None of all the family members had hearing abnormality, mental dysplasia or gait instability. We identified two novel compound heterozygous variants (c.91A>G, p.T31A; c.353G>T, p.C118F) in ARL3 in the proband, while his father only had variant c.91A>G. Bioinformatics analysis indicated amino acid positions of the two variants are highly conserved among species. The in silico tools predicted the variants to be harmful. Protein structure analysis showed the two variants had potential to alter the protein structure. Based on the ACMG guidelines, the two variants were likely pathogenic. In addition, the
ARL3 mutations destabilized
ARL3 protein, and the mutation c.353G>T disrupted the interaction between
ARL3 and RP2 in HEK293T cells. Conclusions: We showed novel compound heterozygous variants in ARL3 were associated with early onset of autosomal recessive RCD, while c.91A>G along may be associated with a late onset of dominant CRD. The two variants in
ARL3 could be causative by destabilizing
ARL3 protein and impairing its interaction with RP2 protein.