Molecular characterization of outer dynein arm defects in Primary Ciliary Dyskinesia (PCD)

原发性纤毛运动障碍（primaryciliarydyskinesia，PCD）是一种遗传异质性疾病，其特征是慢性气道疾病、随机化的左右身体不对称和由于运动纤毛/鞭毛功能缺陷导致的男性不育。这种疾病通常会导致永久性的肺损伤，并可能发展为呼吸衰竭。目前的诊断主要依赖于电子显微镜显示的超微结构缺陷，这往往揭示了外动力蛋白臂（ODA）的缺陷。我们证明了编码ODA重链的DNAH 5突变导致了一半的ODA缺陷病例。通过使用高分辨率免疫荧光成像（IF），我们在ODA缺陷患者中发现了两种不同类型的鼻气道细胞DNAH 5错误定位，为PCD引入了一种新的诊断工具。在这里，我们想从遗传学上描述一个大型国际PCD患者队列中已知PCD基因DNAH 5和DNAI 1突变的存在。为了分析人ODA的组成和产生，我们将产生针对其他尚未表征的ODA重链的抗体，并应用蛋白质技术。为了提高对PCD致病机制和诊断的认识，我们将把遗传学结果与临床、超微结构和IF结果联系起来。为了评估新的治疗选择，我们将进行体外纤毛发生和应用药物基因治疗突变的呼吸细胞。

Primary ciliary dyskinesia (PCD) is a genetically heterogeneous disorder characterized bychronic airway disease, randomization of left-right body asymmetry and male infertility due todefects of motile cilia/flagella function. The disease often results in permanent lung damageand can progress to respiratory failure. Currently diagnosis mainly relies on demonstration ofultrastructural defects by electron microscopy, which often reveals outer dynein arm (ODA)deficiency. We demonstrated that mutations in DNAH5, encoding an ODA heavy chain, areresponsible for half of all ODA defect cases. With the use of high-resolution immunofluorescenceimaging (IF) we identified in patients with ODA defects two distinct types of DNAH5mislocalization in nasal airway cells, introducing a novel diagnostic tool in PCD. Here, wewant to genetically characterize a large international cohort of PCD patients for presence ofmutations in both of the known PCD genes, DNAH5 and DNAI1. To analyze composition andgeneration of human ODAs we will generate antibodies directed against other yet uncharacterizedODA heavy chains and apply protein techniques. To improve understanding ofthe disease causing mecanisms and diagnosis in PCD we will correlate genetic results withclinical, ultrastructural and IF findings. To evaluate novel therapeutic options we will performin vitro ciliogenesis and apply pharmaco-gene therapy in mutant respiratory cells.