Immunofluorescence Analysis and Diagnosis of Primary Ciliary Dyskinesia with Radial Spoke Defects
Immunofluorescence Analysis and Diagnosis of Primary Ciliary Dyskinesia with Radial Spoke Defects
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DOI：
10.1165/rcmb.2014-0483oc
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发表时间：
2015-10-01
影响因子：
6.4
通讯作者：
Omran, Heymut
中科院分区：
文献类型：
--
作者：
Frommer, Adrien;Hjeij, Rim;Omran, Heymut
原发性纤毛运动障碍(PCD)是一种遗传异质性的隐性疾病，由纤毛搏动相关基因的几个明显缺陷引起，导致粘液纤毛清除缺陷，通常与左右身体不对称的随机化有关。由于缺乏粗大的超微结构缺陷和无内翻部位，由放射头缺陷引起的PCD患者很难诊断。到目前为止，在人类放射状辐条基因(RSPH)中发现的大多数突变都是功能丧失突变，错义突变很少被描述。我们研究了不同的RSPH9、RSPH4A和RSPH1突变对RS复合体组装的影响，以提高PCD的诊断水平。我们报告了21例PCD患者(16个家系)在RSPH9、RSPH4A和RSPH1上有双等位基因突变，其中包括7个新的含有错义变异的突变，并对人类呼吸道纤毛进行了高分辨率免疫荧光分析。错义突变是伴有RS缺陷的PCD中常见的遗传缺陷。由于RSPH4A突变导致RSPH4A的缺失导致RS头部组件RSPH1和RSPH9的轴丝组装缺陷。RSPH1突变体纤毛不能组装RSPH9，而RSPH9突变导致RSPH9轴丝缺失，但不影响其他头部蛋白RSPH1和RSPH4A的组装。有趣的是，我们的结果在携带功能丧失突变、错义变异或一个氨基酸缺失的个体中是相同的。免疫荧光分析可以提高对功能丧失突变和错义突变患者的PCD的诊断。RSPH4A是RS头蛋白的核心蛋白。
Primary ciliary dyskinesia (PCD) is a genetically heterogeneous recessive disorder caused by several distinct defects in genes responsible for ciliary beating, leading to defective mucociliary clearance often associated with randomization of left/right body asymmetry. Individuals with PCD caused by defective radial spoke (RS) heads are difficult to diagnose owing to lack of gross ultrastructural defects and absence of situs inversus. Thus far, most mutations identified in human radial spoke genes (RSPH) are loss-of-function mutations, and missense variants have been rarely described. We studied the consequences of different RSPH9, RSPH4A, and RSPH1 mutations on the assembly of the RS complex to improve diagnostics in PCD. We report 21 individuals with PCD (16 families) with biallelic mutations in RSPH9, RSPH4A, and RSPH1, including seven novel mutations comprising missense variants, and performed high-resolution immunofluorescence analysis of human respiratory cilia. Missense variants are frequent genetic defects in PCD with RS defects. Absence of RSPH4A due to mutations in RSPH4A results in deficient axonemal assembly of the RS head components RSPH1 and RSPH9. RSPH1 mutant cilia, lacking RSPH1, fail to assemble RSPH9, whereas RSPH9 mutations result in axonemal absence of RSPH9, but do not affect the assembly of the other head proteins, RSPH1 and RSPH4A. Interestingly, our results were identical in individuals carrying loss-of-function mutations, missense variants, or one amino acid deletion. Immunofluorescence analysis can improve diagnosis of PCD in patients with loss-of-function mutations as well as missense variants. RSPH4A is the core protein of the RS head.