HYDIN variants cause primary ciliary dyskinesia in the Finnish population

Abstract

Introduction: Primary ciliary dyskinesia (PCD) is a rare genetic disorder 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. Thus far, disease causing variants 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 highlighting the importance of characterizing gene variants in different populations for development of gene-based diagnostics and management.

Methods: Whole exome sequencing was used to identify disease causing variants in Finnish PCD cohort. The effect of the identified HYDIN variants on cilia structure and function was confirmed by high-speed video analysis, immunofluorescence and electron tomography.

Results: In this study, we identified three Finnish PCD patients carrying homozygous loss-of-function variants and one patient with compound heterozygous variants within HYDIN. The functional studies showed defects in the axonemal central pair complex. All patients had clinical PCD symptoms including chronic wet cough and recurrent airway infections, associated with mostly static airway cilia.

Conclusion: Our results are consistent with the previously identified important role of HYDIN in the axonemal central pair complex and improve specific diagnostics of PCD in different national populations.

Keywords: HYDIN; PCD; axoneme; diagnostics; motile cilia; variant.

Figure 1

Stop gain variants in HYDIN cause PCD in Finnish patients S6‐S8. (A) HYDIN and SPEF2 form a complex in the axonemal central pair projection. (B) Stop gain variant (c.1797 C > G) in HYDIN exon 14 was identified in three Finnish PCD patients and confirmed by Sanger sequencing. (C) The c.1797 C > G variant results in a predicted truncated HYDIN protein product as visualized by the AlphaFold protein structure prediction tool (https://alphafold.ebi.ac.uk/). PCD, primary ciliary dyskinesia.

Figure 2

Compound heterozygous variants result in decreased HYDIN expression in a PCD patient S10. (A) Sanger sequencing of patient RNA confirmed heterozygosity of the identified pathogenic HYDIN variants. (B) HYDIN transcript expression was decreased in the patient with HYDIN variants c.12899 G > C and c.13801delG compared to control. GAPDH was used as a loading control. PCD, primary ciliary dyskinesia.

Figure 3

The axonemal central pair projection is affected in the PCD patient S10 with compound heterozygous HYDIN variants. (A) TEM showed apparently normal microtubular structure and the presence of dynein arms and radial spokes in the patient's axonemal cross sections. (B) SPEF2 staining was depleted in cilia of the patient's nasal brushing samples. (C) Electron tomography showed a lack of the SPEF2/HYDIN complex in the patient's central pair projection. (D) 2D image averaging using PCD Detect identified the lack of the SPEF2/HYDIN complex in the patient's central pair. PCD, primary ciliary dyskinesia.

Figure 4

Immunofluoresence confirmed the presence of the main axonemal structures and Sanger sequencing the inheritance pattern for one HYDIN variant in patient S10. (A) Outer dynein arm protein DNAH5 and radial spoke protein RSPH4A protein localization in PCD patient with c.12899 G > C and c.13801delG variants was comparable to the control. (B) SPEF2, DNAH5 and RSPH4A were present in the airway cilia of the patient's carrier mother. (C) Sanger sequencing of patients mother showed heterozygosity of variant c.12899 G > C and control homozygosity of variant c.13801delG confirming the inheritance of c.12899 G > C from the mother to the patient. PCD, primary ciliary dyskinesia.