Mutation of CFAP57, a protein required for the asymmetric targeting of a subset of inner dynein arms in Chlamydomonas, causes primary ciliary dyskinesia.

Ximena M Bustamante-Marin,Maimoona A Zariwala,Mihaela Stoyanova,Hailey Bowen,Susan K Dutcher,Leigh Anne Daniels,Amjad Horani,Lawrence E Ostrowski,Wu-Lin Charng,Mathieu Bottier,Wei-Ning Yin,Donald F Conrad,Michael R Knowles,Patrick R Sears

doi:10.1371/journal.pgen.1008691

Abstract

Primary ciliary dyskinesia (PCD) is characterized by chronic airway disease, reduced fertility, and randomization of the left/right body axis. It is caused by defects of motile cilia and sperm flagella. We screened a cohort of affected individuals that lack an obvious axonemal defect for pathogenic variants using whole exome capture, next generation sequencing, and bioinformatic analysis assuming an autosomal recessive trait. We identified one subject with an apparently homozygous nonsense variant [(c.1762C>T), p.(Arg588*)] in the uncharacterized CFAP57 gene. Interestingly, the variant results in the skipping of exon 11 (58 amino acids), which may be due to disruption of an exonic splicing enhancer. In normal human nasal epithelial cells, CFAP57 localizes throughout the ciliary axoneme. Nasal cells from the PCD patient express a shorter, mutant version of CFAP57 and the protein is not incorporated into the axoneme. The missing 58 amino acids include portions of WD repeats that may be important for loading onto the intraflagellar transport (IFT) complexes for transport or docking onto the axoneme. A reduced beat frequency and an alteration in ciliary waveform was observed. Knockdown of CFAP57 in human tracheobronchial epithelial cells (hTECs) recapitulates these findings. Phylogenetic analysis showed that CFAP57 is highly conserved in organisms that assemble motile cilia. CFAP57 is allelic with the BOP2/IDA8/FAP57 gene identified previously in Chlamydomonas reinhardtii. Two independent, insertional fap57 Chlamydomonas mutant strains show reduced swimming velocity and altered waveforms. Tandem mass tag (TMT) mass spectroscopy shows that FAP57 is missing, and the “g” inner dyneins (DHC7 and DHC3) and the “d” inner dynein (DHC2) are reduced, but the FAP57 paralog FBB7 is increased. Together, our data identify a homozygous variant in CFAP57 that causes PCD that is likely due to a defect in the inner dynein arm assembly process.

Highlights

Motile cilia are complex organelles that project from the surface of cells and are essential for propelling fluids in the airways, ventricles and fallopian tubes or for providing cell locomotion as in sperm or Chlamydomonas
Primary ciliary dyskinesia (PCD) is a rare genetic disease that affects the function of motile cilia
By applying whole exome sequencing and bioinformatic analysis, we identified a variant in an uncharacterized gene, CFAP57, in a subject previously diagnosed with PCD

Summary

Introduction

Motile cilia are complex organelles that project from the surface of cells and are essential for propelling fluids in the airways, ventricles and fallopian tubes or for providing cell locomotion as in sperm or Chlamydomonas. Genetic variants that cause PCD have been identified in over 40 genes [27,28,29,30,31,32,33,34,35,36,37], there are individuals with confirmed clinical features of PCD but normal axonemal structure as determined by transmission electron microscopy, and the genetic basis of disease remains unknown in about 30% of patients. This diagnostic dilemma is likely to require multiple techniques and approaches. With the continual improvement in sequencing techniques and bioinformatic analysis, whole exome sequencing provides a powerful tool to confirm the diagnosis of PCD in these difficult cases

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Conclusion