DYNC2H1基因突变导致短肋胸廓发育不良III型或伴多指（趾）畸形发生机制的研究

Short-Rib Thoracic Dysplasia-3 with or without polydactyly (SRTD3) is a lethal skeletal disease characterized by severely shortened tubular bones and narrow chest. Almost all fetuses had a outcome of termination of pregnancy, which bring heavy mental stress to the couple and family. SRTD3 is caused by the mutations in DYNC2H1 gene, which encodes the heavy chain of the cytoplasmic dynein-2 motor required for trafficking cargo from the tip to the base of the cilium. DYNC2H1 gene plays very important role in development of chondrocytes and skeletal. In the previous work, we have identified the new DYNC2H1 gene compound heterozygous mutations in a SRTD3 family and the immunohistochemical results of the affected fetal long bone suggested the proliferation of chondrocytes were disorganized. Our study will focus the function of the new mutations using Realtime-PCR, Western-blot, IHC and immunofluorescence on the level of cells, model animals and tissue samples in order to discover the changes on gene expression, protein structure and function. Our study will clarify the pathogenesis and provide the further evidence of SRTD3 caused by the DYNC2H1 gene new compound heterozygous mutation detected in the previous work, the results will provide the theoretical basis for prenatal and preimplantation diagnosis to the mutation carriers.

短肋胸廓发育不良III型或伴多指（趾）（SRTD3）是一种罕见的致死性骨骼系统疾病，受累胎儿在孕中期即表现为严重的长骨短小及胸廓狭窄等畸形，并因此终止妊娠，通常给夫妻双方带来沉重的心理压力。该病由DYNC2H1基因突变导致。DYNC2H1基因编码动力蛋白-2的重链，负责相关信号蛋白从纤毛顶部到纤毛底端的运输，在软骨细胞及骨骼发育中起重要作用。课题组前期在一中国SRTD3家系患儿中筛查到DYNC2H1基因新的复合杂合突变，并且进一步观察到受累胎儿长骨软骨细胞增殖紊乱。本项目拟通过实时荧光定量PCR、Western-blot、免疫组化、免疫荧光等技术在细胞、动物及引产组织标本中对发现的新复合杂合突变进行研究，探讨其对DYNC2H1基因表达、蛋白结构及功能的影响。本研究将阐明新发现的DYNC2H1基因复合杂合突变导致SRTD3发生的分子机制，为携带者产前诊断及植入前诊断提供理论依据。

短肋胸廓发育不良III型或伴多指（趾）（SRTD3）是一种罕见的致死性骨骼系统疾病，呈常染色体隐性遗传，正常人类携带者不出现临床症状，但患者在胎儿期即表现为严重的长骨短小及胸廓狭窄等畸形。该病由DYNC2H1基因突变导致。本课题组前期收集了4例中国SRTD3家系信息，并且完成了包括引产胎儿影像学检查，基因检测等研究，并且在一SRTD3家系中筛查到尚未见报道的DYNC2H1基因新的复合杂合突变。由于引产胎儿RNA及蛋白降解严重以及Dync2h1基因质粒过大转染失败，因此无法对突变的蛋白功能进行检测。但我们成功构建了携带相应变异位点（c.10045C>T和 c.10163C>T）的C57BL/6J小鼠模型，以便研究突变的Dync2h1蛋白功能。虽然因疫情原因使后续实验拖延，但是本课题组成功对该家系实施了植入前单基因病检测（PGT-M），并获得了健康后代。同时，课题组也依靠本课题收集了其他的骨骼畸形家系样本，发表了多篇中英文文章，并以此参与了一项国家重点研发计划，并获得33万经费。因此，基本上完成了本课题的目标。