CFAP45 deficiency causes situs abnormalities and asthenospermia by disrupting an axonemal adenine nucleotide homeostasis module

Sandra Cindrić,Gerard W Dougherty,Petra Pennekamp,Hiroshi Hamada,Katsuyoshi Takaoka,Rim Hjeij,Zeev Perles,Thomas Kaiser,Claudius Werner,Friederike Stumme,Heike Olbrich,Bernd Dworniczak,Inga M Höben,Isabella Aprea,Judith A Klinkenbusch,Takahiro Ide,Stef J.f Letteboer ,Niki T Loges,Micha Aviram,Luisa Biebach,Yuan Ping Pang ,Kerstin Bartscherer,Ronald Roepman,Nicholas Katsanis,Johanna Raidt,Franziska Rabert,Tabea Nöthe-Menchen,Timo Strünker,Samuel Young,Yayoi Ikawa,Jana Gützlaff,Diana Bracht ,Erica E Davis,Huda Mussaffi,Hidetaka Shiratori,Wang Kyaw Twan,Karsten Boldt,Nicola Horn,Julia Wallmeier,Heymut Omran,Kavita Praveen,Yasin Memari,Katsura Minegishi,Israel Amirav,Katrin Junger,Petras P Dzeja,Marius Ueffing,Orly Elpeleg,Asaf Ta‐Shma ,Katsutoshi Mizuno,Jörg Große-Onnebrink

doi:10.1038/s41467-020-19113-0

Abstract

Axonemal dynein ATPases direct ciliary and flagellar beating via adenosine triphosphate (ATP) hydrolysis. The modulatory effect of adenosine monophosphate (AMP) and adenosine diphosphate (ADP) on flagellar beating is not fully understood. Here, we describe a deficiency of cilia and flagella associated protein 45 (CFAP45) in humans and mice that presents a motile ciliopathy featuring situs inversus totalis and asthenospermia. CFAP45-deficient cilia and flagella show normal morphology and axonemal ultrastructure. Proteomic profiling links CFAP45 to an axonemal module including dynein ATPases and adenylate kinase as well as CFAP52, whose mutations cause a similar ciliopathy. CFAP45 binds AMP in vitro, consistent with structural modelling that identifies an AMP-binding interface between CFAP45 and AK8. Microtubule sliding of dyskinetic sperm from Cfap45−/− mice is rescued with the addition of either AMP or ADP with ATP, compared to ATP alone. We propose that CFAP45 supports mammalian ciliary and flagellar beating via an adenine nucleotide homeostasis module.

Highlights

Axonemal dynein ATPases direct ciliary and flagellar beating via adenosine triphosphate (ATP) hydrolysis
We analyzed individual OP-28 II1 and identified compound heterozygous nonsense mutations (c.721C>T, p.Gln241* and c.907C>T, p.Arg303*, rs201144590) in cilia and flagella associated protein 45 (CFAP45; GenBank ID: NM_012337) (Fig. 1a), which we confirmed by Sanger sequencing to segregate in an autosomal recessive manner (Fig. 1b)
A homozygous frameshift CFAP45 mutation (c.1472_1477delAGAACCinsT, p.Gln491Leufs*5) was identified in individual TB-19 II1 that was prenatally diagnosed with left–right asymmetry (LRA) abnormalities including heart defect (Supplementary Fig. 1)

Summary

Introduction

Axonemal dynein ATPases direct ciliary and flagellar beating via adenosine triphosphate (ATP) hydrolysis. The coordinated beating of eukaryotic cilia and flagella is regulated by axonemal heavy chain dynein ATPases, which are motor proteins that generate force along microtubules (MTs) via adenosine triphosphate (ATP) hydrolysis[1,2,3]. It has been shown that adenosine monophosphate (AMP) incubated with ATP can mimic the modulatory effect of ADP alone on flagellar beat pattern[19] The mechanism underlying this effect is not fully understood but explained in part by adenylate kinase (AK), which reversibly catalyses the reaction ATP + AMP ↔ 2 ADP and maintains balanced nucleotide pools to support ciliary beating[20]. CFAP45 loss-of-function mutations in humans as well as CRISPR/Cas[9] ablation of Cfap[45] in mice cause LRA abnormalities including situs inversus totalis as well as asthenospermia, due to dyskinetic beating of embryonic nodal cilia and sperm flagella, respectively. This study advances the molecular framework of mammalian ciliary and flagellar beating and presents disruption of adenine nucleotide homeostasis as a pathomechanism underlying a human motile ciliopathy

Methods

Results

Conclusion