Discussion
In this study, two compound heterozygous mutations of DNAAF2 were identified in two siblings with similar characteristic manifestations of PCD including recurrent pneumonia with atelectasis mainly in right middle and left lingular lobe, early bronchiectasis, sinusitis, otitis media, significant reduction in nNO level and complete loss of ODAs and IDAs of bronchial cilia found by the TEM. One of the mutations is a nonsense mutation of NM_018139: c.156C>A (p.Y52*) which has already been reported in a Chinese family and proved to be pathogenic[5]. Another mutation is a frameshift mutation (NM_018139: c.177_178insA (p.E60Rfs*3) , which is a novel mutation.
DNAAF2 is located on chromosome 14q21.3 and consists of three exons encoding cDNAs [6]. The mutation p.E60Rfs*3 locates in exon1 and near another mutation p.Y52*. Exon1 is consists of 621 amino acids, and the mutation of E60Rfs*3 leads to early transcription termination after the 63 amino acids (Fig.6). The p.E60Rfs*3 mutation has been predicted to be pathogenic with the evidences of PVS1+PM2+PM3+PP4 according to the standards and guidelines of American College of Medical Genetics and Genomics (ACMG). Multiple software, such as SIFT, Polyphen‑2 and Mutation Taster, also predict the p.E60Rfs*3 mutation to be pathogenic. In addition, none of cystic fibrosis (CF) or primary immune deficiency (PID) related genes which may also lead to recurrent pneumonia, atelectasis and bronchiectasis were not detected. Based on the clinical manifestations and hereditary mode of compound heterozygous mutations from both parents, it’s reasonable to highly suspected that the novel p.E60Rfs*3 mutation of DNAAF2 is a disease causing mutation and is responsible for the pathogenesis of PCD.
The movements of cilia can help to clear airway mucous, cerebrospinal fluid flow along the brain ventricular system, support the transport of the oocyte to the uterus, and male germ cell move along the female reproductive tract [6]. The normal structure of the cilia consists of the conserved basic 9+2 microtubule-based axonemes and other several functional modules[7]. Members of multi-subunit motor protein complexes, ODAs and IDAs are responsible for the generation and regulation of ciliary beating. Dynein arms are ATPase-based protein-complexes, via ATP hydrolyzed by dynein heavy chain, the paired microtubules slide relative to each other, leading to cilia curve over the deformed microtubules[8-9]. During the process, IDA causes the curvature of cilia, ODA accelerates active sliding of the outer microtubules, leading to effective propulsion produced by the cilia effectively paddling forward and reverting paddling backward. Appropriate rheological properties and the functional structure of cilia are the basis that maintain mucociliary clearance which is essential to the respiratory tract clearance defense[5].
Approximately 40 genes have been reported to be associated with PCD, including DNAH5, DNAH11, DNAI1, DNAI2, DNAL1, TXNDC3, DNAAF1, and DNAAF2 et al. [1, 6, 10-14]. In 2008, two heterozygous mutations c.C23A [pS8X] and c.1214-1215insACGATACCTGCGTGGC [p.G406Rfs89X mutations of DNAAF2 were first identified in two consanguineous families with PCD presented with recurrent respiratory tract infections, laterality defects and impaired fertility[6]. DNAAF2 works as a facilitator of dynein pre-assembly. The disfunction of DNAAF2 is associated with specific defects in the interaction of intermediate and heavy chains in the cytoplasm which leads to complete or partial loss of ODAs and IDAs and loss of motility[6]. After that, only 15 PCD cases have been reported caused by DNAFF2 with 19 mutations. The cases of DNAAF2 mutation are summarized in Tab.1[5-6, 15-27]. As for Chinese children, DNAFF2 gene is rare. As Guan et al reported reported in 81 Chinese children, genes with the highest incidence of mutations that caused PCD were DNAH11, followed by HYDIN, DNAH5, CCDC39, DNAH1 and CCNO, no DNAFF2 gene mutation was detected[28]. In Chinese adult patients, Sun et.at have identified two heterozygous mutations c.C156A [p.Y52X] and c.C26A [p.S9X] in the DNAAF2 gene which lead to the defect of outer dynein arms and inner dynein arms resulting in PCD with the manifestation of male infertility[5]. Lu C et.at detected another two heterozygous mutations c.491T>C [p.L164P] and c.822del [p.A275Profs*10] in two females presents with the bronchiectasis, sinusitis, situs inversus, and infertility, besides, the one with c.491T>C mutation had scoliosis[27].
Consequently, we made an early diagnosis of PCD in two siblings with a novel mutation of the rare DNAFF2 gene. The early stagy of PCD may be easily misdiagnosed as common pneumonia or sinusitis. When a child presented with recurrent pneumonia or atelectasis accompanied with recurrent sinusitis or otitis media, PCD should be considered. The characteristic HRCT manifestations including atelectasis in the right middle and left lingular lobe, thickened bronchial wall, and mild bronchiectasis can provide a diagnostic clue for PCD. Besides, nNO and TEM of bronchial cilia are useful for the further diagnosis , and genetic tests can help to confirm the diagnosis.
In conclusion, our study suggests that the p.E60Rfs*3 mutation of DNAAF2 gene is a novel disease causing mutation of PCD. This study enriches our knowledge of clinical manifestations and genetic information of PCD caused by DNAAF2.
Acknowledgements We are grateful to the patients and their family for
their collaboration.
Funding Medical Science and Technology Project of Henan Province (2018020621); Demonstration application of remote collaborative service network for complex genetic diseases (SJPT-03-01).
Conflicts of interest The authors declare that they have no competeing conflicts.