Cilia are small microtubule-based structures found on the surface of most mammalian cells, which have key sensory and sometimes motile functions. Primary ciliary dyskinesia (PCD) is a type of ciliopathy caused by defects in motile cilia. The genetic basis of PCD is only partially understood. Studying a cohort of 11 human patients with PCD, we find thatde novomutations inTUBB4B, a beta tubulin isotype, cause three distinct classes of ciliopathic disease.In vivostudies in mice show thatTubb4bplays a specific role in cilia, building centrioles and axonemes in multiciliated cells. Examining the effects of specific TUBB4B variants in cells and in mice, we further demonstrate that distinctTUBB4Bmutations differentially affect microtubule dynamics and cilia formation in a dominant negative manner. Finally, structure-function studies reveal that different TUBB4B mutations disrupt distinct tubulin interfaces. Importantly, these molecular differences correlate with disease features. We show that tubulin heterodimer-impairing TUBB4B variants underlie nonsyndromic PCD, whilst additional renal and sensorineural ciliopathic features in a syndromic PCD subtype arise from microtubule lumenal interface-impaired TUBB4B variants. These findings suggest that specific tubulin isotypes have distinct and non-redundant subcellular functions, and demonstrate that human tubulinopathies can be drivers of ciliopathic syndromes.

Diagnostic testing for primary ciliary dyskinesia (PCD) started in 2013 in Palestine. We aimed to describe the diagnostic, genetic, and clinical spectrum of the Palestinian PCD population.Individuals with symptoms suggestive of PCD were opportunistically considered for diagnostic testing: nasal nitric oxide (nNO) measurement, transmission electron microscopy (TEM), and/or PCD genetic panel or whole exome testing. Clinical characteristics of those with a positive diagnosis were collected close to testing including FEV1GLI z-scores, and BMI z-scores.Sixty-eight individuals had a definite positive PCD diagnosis, 31 confirmed by genetic and TEM results, 23 by TEM results alone, and 14 by genetic variants alone. Forty-five individuals from 40 families had seventeen clinically actionable variants, and 4 had variants of unknown significance in 14 PCD-genes. CCDC39, DNAH11, andDNAAF11were the most commonly mutated genes. 100% of variants were homozygous. Patients had median age of 11.2 years at diagnosis, were highly consanguineous (93%) and 100% of Arabic descent. Clinical features included persistent wet cough (99%), neonatal respiratory distress (84%), and situs inversus(43%). Lung function at diagnosis was already impaired (FEV1z-score median −1.90 (−5.0 to 1.32)) and growth was mostly within the normal range (z-score mean= −0.36 (−3.03 to 2.57). 19% individuals had finger clubbing.Despite limited local resources, detailed geno- and phenotyping forms the basis of one of the largest national PCD populations globally. There was notable familial homozygosity within the context of significant population heterogeneity.

Primary ciliary dyskinesia (PCD) is a rare genetic condition characterized by chronic respiratory tract infections and in some cases laterality defects and infertility. The symptoms of PCD are caused by malfunction of motile cilia, hair-like organelles protruding out of the cell that are responsible for removal of mucus from the airways and organizing internal organ positioning during embryonic development. PCD is caused by mutations in genes coding for structural or assembly proteins in motile cilia. Thus far mutations in over 50 genes have been identified and these variants explain around 70% of all known cases. Population specific genetics underlying PCD has been reported, thus highlighting the importance of characterizing gene variants in different populations for development of gene-based diagnostics. In this study, we identified a recurrent loss-of-function mutation c.198_200delinsCC in CFAP300 causing lack of the protein product. PCD patients homozygous for the identified CFAP300 mutation have immotile airway epithelial cilia associated with missing dynein arms in their ciliary axonemes. Furthermore, using super resolution microscopy we demonstrate that CFAP300 is transported along cilia in normal human airway epithelial cells suggesting a role for CFAP300 in dynein complex transport in addition to preassembly in the cytoplasm. Our results highlight the importance of CFAP300 in dynein arm assembly and improve diagnostics of PCD in Finland.