A comprehensive functional analysis on the pathogenesis of novel TSPAN12 and NDP variants in familial exudative vitreoretinopathy
A comprehensive functional analysis on the pathogenesis of novel TSPAN12 and NDP variants in familial exudative vitreoretinopathy
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DOI：
10.1111/cge.14273
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发表时间：
2022-12
影响因子：
3.5
通讯作者：
Rulian Zhao;Erkuan Dai;Shi-yuan Wang;Xiang Zhang;Yunqi He;Li Peng;Peiquan Zhao;Zhenglin Yang-Zhenglin-Yan
中科院分区：
文献类型：
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作者：
Rulian Zhao;Erkuan Dai;Shi-yuan Wang;Xiang Zhang;Yunqi He;Li Peng;Peiquan Zhao;Zhenglin Yang-Zhenglin-Yan
家族性渗出性玻璃体视网膜病变(FEVR)是一种遗传性致盲疾病；然而，已知的FEVR相关变异只占大约50%的病例。目前，大多数已报道的变异体的发病机制还没有得到很好的研究，我们的目标是从FEVR相关基因中寻找新的变异体，并进行全面的功能分析，以揭示导致FEVR的变异体的发病机制。利用靶向基因面板和Sanger测序，我们在TSPAN12和NDP中鉴定了6个新的和3个已知的变异体。双荧光素酶报告基因分析和免疫印迹分析表明，这些突变体对去甲肾上腺素/β-连环蛋白途径有明显的抑制作用。结构分析和免疫共沉淀表明，在Norrin/β-Catenin途径中，错义变体和结合伙伴之间的相互作用受到损害。免疫荧光和亚细胞蛋白提取显示异常的亚细胞转运。此外，TSPAN12的过表达通过增强突变体Norrin与FZD4或β的结合亲和力，成功地增强了Norrin/LRP5信号转导活性。总之，这些观察结果扩大了FEVR相关变异的范围，用于FEVR的遗传咨询和产前诊断，并为FEVR的治疗提供了一种潜在的治疗策略。
Familial exudative vitreoretinopathy (FEVR) is an inherited blinding disorder; however, the known FEVR‐associated variants account for approximately only 50% cases. Currently, the pathogenesis of most reported variants is not well studied, we aim to identify novel variants from FEVR‐associated genes and perform a comprehensive functional analysis to uncover the pathogenesis of variants that cause FEVR. Using targeted gene panel and Sanger sequencing, we identified six novel and three known variants in TSPAN12 and NDP. These variants were demonstrated to cause significant inhibition of Norrin/β‐catenin pathway by dual‐luciferase reporter assay and western blot analysis. Structural analysis and co‐immunoprecipitation revealed compromised interactions between missense variants and binding partners in the Norrin/β‐catenin pathway. Immunofluorescence and subcellular protein extraction were performed to reveal the abnormal subcellular trafficking. Additionally, over‐expression of TSPAN12 successfully enhanced the Norrin/β‐catenin signaling activity by strengthening the binding affinity of mutant Norrin with FZD4 or LRP5. Together, these observations expanded the spectrum of FEVR‐associated variants for the genetic counseling and prenatal diagnosis of FEVR, as well providing a potential therapeutic strategy for the treatment of FEVR.