Structural and functional characterization of DNAH5 variants in a Portuguese family with primary ciliary dyskinesia

Purpose Primary ciliary dyskinesia (PCD) is a rare genetic heterogeneous disorder mainly characterized by impaired mucociliar clearance and chronic respiratory symptoms. Although DNAH5 is commonly implicated in PCD, several DNAH5 variants remain unclassified. Methods The proband was studied by high-speed videomicroscopy, transmission electron microscopy, whole exome sequencing and immunofluorescence. Protein structural analysis was performed through structural models obtained by X-ray crystallography and cryo-electron microscopy. The family was studied by Sanger sequencing of the variants, high-speed videomicroscopy and immunofluorescence. Results The patient carry, in heterozygosity, the DNAH5 c.5290 T > C p.(Ser1764Pro) missense variant of uncertain significance and the pathogenic truncated variant DNAH5 c.4237C > T p.(Gln1413*). He was clinically diagnosed in adulthood with PCD, confirmed following nasal nitric oxide measurement, high-speed videomicroscopy and transmission electron microscopy, all of which revealed hallmark PCD defects, with decreased nasal nitric oxide levels (30 nl/min) and ciliary beating frequency (0.66 Hz), a dyskinetic ciliary beating pattern (62.5% total immotility) and a class-1 ultrastructure. Immunofluorescence analysis demonstrated reduced DNAH5 expression, and protein structural models predicted that the variant of uncertain significance causes an unstable protein. Family analyses confirmed a trans-inheritance and uncovered a brother with a similar PCD phenotype, the same two variants and similar reduced DNAH5 expression. Conclusions Results support a pathogenic role for the c.5290 T > C p.(Ser1764Pro) variant and elucidate the effects of the other variant. These results underscore the importance of integrating clinical, ultrastructural, molecular and protein expression analyses to clarify and contribute to PCD diagnosis, besides now serving as potential markers for diagnostics and targeted therapies.