REGULATION OF MOTILE CILIA ASSEMBLY IN LUNG DISEASE

 描述(由申请者提供)：呼吸道清除障碍是慢性呼吸道疾病的特征。纤毛遗传疾病的研究以及纤毛功能障碍可能导致慢性阻塞性肺疾病(COPD)和哮喘的报道表明，完整的粘液纤毛器对于清除纤毛是必不可少的。广泛收集的纤毛基因和蛋白质有助于快速确定运动性纤毛功能障碍综合征的遗传病因，称为原发性纤毛运动障碍(PCD)。然而，目前还没有针对这种疾病的特效疗法。 进展为支气管扩张症。在这里，我们解决一组关键问题，以更全面地了解活动纤毛的组装和调节。以PCD中突变的新基因为指导，我们发现了对运动纤毛组装至关重要的特定调控步骤，并提供了促进纤毛生成的治疗干预的前景。我们的初步数据确定了一条运动纤毛发生的途径，该途径(I)开始于由HEATR2控制的细胞质中运动蛋白的预组装的最早点，(Ii)在功能上与由CCDC11调控的蛋白质从细胞质运输到基本体和运动纤毛的网络相连，以及(Iii)需要CCDC39和CCDC40蛋白来建立运动纤毛中运动和微管的适当组织。我们发现，这些分子连接链中的每一个突变或缺失都会导致运动纤毛的生物发生失败，并发挥预期的功能。我们还发现了在通路的每个点上的容易处理的机制，我们展示了被新的抑制基因的基因破坏来修复纤毛缺陷的改变。我们的初步和拟议研究利用了原代多纤毛人类呼吸道上皮细胞、遗传缺陷的斑马鱼品系和 藻类衣藻，共同提供体外和活体分析和纤毛组装途径的快速遗传操作。利用这些策略，我们将从以下目的探讨动力蛋白马达组装的组装、运输和活性：(1)鉴定动力蛋白马达蛋白复合体运送所需的预组装途径；(2)鉴定马达蛋白从基底向纤毛运输的调节；(3)鉴定能够恢复缺乏马达蛋白对接和组装因子的细胞功能的基因。我们认为，对这些通路的操纵将为设计运动纤毛缺陷的分子疗法提供可能性。

 DESCRIPTION (provided by applicant): Failure of airway clearance is a characteristic feature of chronic airway disorders. Evidence that an intact mucociliary apparatus is essential for clearance derives from studies of genetic disease of cilia as well as reports that cilia dysfunctio may contribute to chronic obstructive pulmonary disease (COPD) and asthma. Comprehensive collections of ciliary genes and proteins have contributed to the rapid pace of progress in the identification of genetic etiologies of the syndrome of motile cilia dysfunction, called primary ciliary dyskinesia (PCD). However, there are no specific therapies for this disease that ultimately progresses to bronchiectasis. Here, we address a key set of questions to gain a more complete understanding of the assembly and regulation of motile cilia. Using novel genes mutated in PCD as a guide, we have uncovered specific regulatory steps that are critical for the assembly of motile cilia and offer the prospect of therapeutic intervention to augment ciliogenesis. Our preliminary data define a pathway for motile ciliogenesis that (i) commences at the earliest point of motor protein preassembly in the cytoplasm controlled by HEATR2, (ii) is functionally linked to a network that traffics protein from the cytoplasm to the basal bodies and motile cilia regulated by CCDC11, and (iii) requires the proteins CCDC39 and CCDC40 to establish the proper organization of motors and microtubules within the motile cilium. We show that mutation or depletion of each of these molecular linchpins results in unsuccessful motile cilia biogenesis and function as expected. We also uncover tractable mechanisms at each point of the pathway that we show are modified by the genetic disruption of novel suppressor genes to remediate cilia defects. Our preliminary and proposed studies take advantage of complementary models of primary multiciliated human airway epithelial cells, genetically deficient strains of zebrafish and the alga Chlamydomonas, which together provide in vitro and in vivo analysis and rapid genetic manipulation of cilia assembly pathways. Using these strategies, we will probe the assembly, trafficking and activity of dynein motor assembly in the following aims: (1) Identify the preassembly pathways required for dynein motor protein complex delivery; (2) Identify the regulation of motor protein trafficking from basal bodies to cilia; (3) Identify genes that can restore function to cells lacking docking and assembly factors for the motor proteins. We propose that manipulation of these pathways will offer possibilities of devising molecular therapies for defects in motile cilia.