Loss-of-Function Mutations in RSPH1 Cause Primary Ciliary Dyskinesia with Central-Complex and Radial-Spoke Defects
2013; Elsevier BV; Volume: 93; Issue: 3 Linguagem: Inglês
10.1016/j.ajhg.2013.07.013
ISSN1537-6605
AutoresEsther Kott, Marie Legendre, Bruno Copin, Jean‐François Papon, Florence Dastot‐Le Moal, Guy Montantin, Philippe Duquesnoy, William Piterboth, Daniel Amram, Laurence Bassinet, J. Beucher, Nicole Beydon, Eric Deneuville, Véronique Houdouin, Hubert Journel, J. Just, Nadia Nathan, Aline Tamalet, Nathalie Collot, Ludovic Jeanson, Morgane Le Gouez, Benoit Vallette, Anne-Marie Vojtek, Ralph Epaud, A. Coste, Annick Clément, Bruno Housset, Bruno Louis, Estelle Escudier, Serge Amselem,
Tópico(s)Genetic and Kidney Cyst Diseases
ResumoPrimary ciliary dyskinesia (PCD) is a rare autosomal-recessive respiratory disorder resulting from defects of motile cilia. Various axonemal ultrastructural phenotypes have been observed, including one with so-called central-complex (CC) defects, whose molecular basis remains unexplained in most cases. To identify genes involved in this phenotype, whose diagnosis can be particularly difficult to establish, we combined homozygosity mapping and whole-exome sequencing in a consanguineous individual with CC defects. This identified a nonsense mutation in RSPH1, a gene whose ortholog in Chlamydomonas reinhardtii encodes a radial-spoke (RS)-head protein and is mainly expressed in respiratory and testis cells. Subsequent analyses of RSPH1 identified biallelic mutations in 10 of 48 independent families affected by CC defects. These mutations include splicing defects, as demonstrated by the study of RSPH1 transcripts obtained from airway cells of affected individuals. Wild-type RSPH1 localizes within cilia of airway cells, but we were unable to detect it in an individual with RSPH1 loss-of-function mutations. High-speed-videomicroscopy analyses revealed the coexistence of different ciliary beating patterns—cilia with a normal beat frequency but abnormal motion alongside immotile cilia or cilia with a slowed beat frequency—in each individual. This study shows that this gene is mutated in 20.8% of individuals with CC defects, whose diagnosis could now be improved by molecular screening. RSPH1 mutations thus appear as a major etiology for this PCD phenotype, which in fact includes RS defects, thereby unveiling the importance of RSPH1 in the proper building of CCs and RSs in humans. Primary ciliary dyskinesia (PCD) is a rare autosomal-recessive respiratory disorder resulting from defects of motile cilia. Various axonemal ultrastructural phenotypes have been observed, including one with so-called central-complex (CC) defects, whose molecular basis remains unexplained in most cases. To identify genes involved in this phenotype, whose diagnosis can be particularly difficult to establish, we combined homozygosity mapping and whole-exome sequencing in a consanguineous individual with CC defects. This identified a nonsense mutation in RSPH1, a gene whose ortholog in Chlamydomonas reinhardtii encodes a radial-spoke (RS)-head protein and is mainly expressed in respiratory and testis cells. Subsequent analyses of RSPH1 identified biallelic mutations in 10 of 48 independent families affected by CC defects. These mutations include splicing defects, as demonstrated by the study of RSPH1 transcripts obtained from airway cells of affected individuals. Wild-type RSPH1 localizes within cilia of airway cells, but we were unable to detect it in an individual with RSPH1 loss-of-function mutations. High-speed-videomicroscopy analyses revealed the coexistence of different ciliary beating patterns—cilia with a normal beat frequency but abnormal motion alongside immotile cilia or cilia with a slowed beat frequency—in each individual. This study shows that this gene is mutated in 20.8% of individuals with CC defects, whose diagnosis could now be improved by molecular screening. RSPH1 mutations thus appear as a major etiology for this PCD phenotype, which in fact includes RS defects, thereby unveiling the importance of RSPH1 in the proper building of CCs and RSs in humans. Primary ciliary dyskinesia (PCD [MIM 244400]) is a rare genetic disease usually transmitted in an autosomal-recessive fashion and affects 1 in 15,000–30,000 individuals.1Afzelius B.A. A human syndrome caused by immotile cilia.Science. 1976; 193: 317-319Crossref PubMed Scopus (936) Google Scholar The disease results from functional and/or structural defects of motile cilia. These defects are responsible for impaired mucociliary transport, leading to recurrent respiratory-tract infections beginning in early childhood. Most male individuals have asthenospermia due to similar defects in their sperm flagella.2Afzelius B.A. The immotile-cilia syndrome: a microtubule-associated defect.CRC Crit. Rev. Biochem. 1985; 19: 63-87Crossref PubMed Scopus (122) Google Scholar Approximately 50% of persons with PCD display situs inversus, thereby defining Kartagener syndrome (MIM 244400).3Kartagener M. Zur Pathogenese der Bronchiektasien: Bronchiektasien bei Situs viscerum inversus.Beiträge zur Klinik der Tuberkulose. 1933; 83: 489-501Crossref Scopus (315) Google Scholar A number of genes playing a key role in the proper building of motile cilia have recently been involved in the pathogenesis of PCD.4Pennarun G. Escudier E. Chapelin C. Bridoux A.M. Cacheux V. Roger G. Clément A. Goossens M. Amselem S. Duriez B. Loss-of-function mutations in a human gene related to Chlamydomonas reinhardtii dynein IC78 result in primary ciliary dyskinesia.Am. J. Hum. Genet. 1999; 65: 1508-1519Abstract Full Text Full Text PDF PubMed Scopus (295) Google Scholar, 5Loges N.T. Olbrich H. Fenske L. Mussaffi H. Horvath J. Fliegauf M. Kuhl H. Baktai G. Peterffy E. Chodhari R. et al.DNAI2 mutations cause primary ciliary dyskinesia with defects in the outer dynein arm.Am. J. Hum. Genet. 2008; 83: 547-558Abstract Full Text Full Text PDF PubMed Scopus (188) Google Scholar, 6Olbrich H. Häffner K. Kispert A. Völkel A. Volz A. Sasmaz G. Reinhardt R. Hennig S. Lehrach H. Konietzko N. et al.Mutations in DNAH5 cause primary ciliary dyskinesia and randomization of left-right asymmetry.Nat. Genet. 2002; 30: 143-144Crossref PubMed Scopus (423) Google Scholar, 7Duriez B. Duquesnoy P. Escudier E. Bridoux A.-M. Escalier D. Rayet I. Marcos E. Vojtek A.-M. Bercher J.-F. Amselem S. A common variant in combination with a nonsense mutation in a member of the thioredoxin family causes primary ciliary dyskinesia.Proc. Natl. Acad. Sci. USA. 2007; 104: 3336-3341Crossref PubMed Scopus (151) Google Scholar, 8Mazor M. Alkrinawi S. Chalifa-Caspi V. Manor E. Sheffield V.C. Aviram M. Parvari R. Primary ciliary dyskinesia caused by homozygous mutation in DNAL1, encoding dynein light chain 1.Am. J. Hum. Genet. 2011; 88: 599-607Abstract Full Text Full Text PDF PubMed Scopus (96) Google Scholar, 9Onoufriadis A. Paff T. Antony D. Shoemark A. Micha D. Kuyt B. Schmidts M. Petridi S. Dankert-Roelse J.E. Haarman E.G. et al.UK10KSplice-site mutations in the axonemal outer dynein arm docking complex gene CCDC114 cause primary ciliary dyskinesia.Am. J. Hum. Genet. 2013; 92: 88-98Abstract Full Text Full Text PDF PubMed Scopus (130) Google Scholar, 10Knowles M.R. Leigh M.W. Ostrowski L.E. Huang L. Carson J.L. Hazucha M.J. Yin W. Berg J.S. Davis S.D. Dell S.D. et al.Genetic Disorders of Mucociliary Clearance ConsortiumExome sequencing identifies mutations in CCDC114 as a cause of primary ciliary dyskinesia.Am. J. Hum. Genet. 2013; 92: 99-106Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar, 11Duquesnoy P. Escudier E. Vincensini L. Freshour J. Bridoux A.-M. Coste A. Deschildre A. de Blic J. Legendre M. Montantin G. et al.Loss-of-function mutations in the human ortholog of Chlamydomonas reinhardtii ODA7 disrupt dynein arm assembly and cause primary ciliary dyskinesia.Am. J. Hum. Genet. 2009; 85: 890-896Abstract Full Text Full Text PDF PubMed Scopus (127) Google Scholar, 12Loges N.T. Olbrich H. Becker-Heck A. Häffner K. Heer A. Reinhard C. Schmidts M. Kispert A. Zariwala M.A. Leigh M.W. et al.Deletions and point mutations of LRRC50 cause primary ciliary dyskinesia due to dynein arm defects.Am. J. Hum. Genet. 2009; 85: 883-889Abstract Full Text Full Text PDF PubMed Scopus (174) Google Scholar, 13Omran H. Kobayashi D. Olbrich H. Tsukahara T. Loges N.T. Hagiwara H. Zhang Q. Leblond G. O’Toole E. Hara C. et al.Ktu/PF13 is required for cytoplasmic pre-assembly of axonemal dyneins.Nature. 2008; 456: 611-616Crossref PubMed Scopus (283) Google Scholar, 14Mitchison H.M. Schmidts M. Loges N.T. Freshour J. Dritsoula A. Hirst R.A. O’Callaghan C. Blau H. Al Dabbagh M. Olbrich H. et al.Mutations in axonemal dynein assembly factor DNAAF3 cause primary ciliary dyskinesia.Nat. Genet. 2012; 44 (S1–S2): 381-389Crossref PubMed Scopus (190) Google Scholar, 15Kott E. Duquesnoy P. Copin B. Legendre M. Dastot-Le Moal F. Montantin G. Jeanson L. Tamalet A. Papon J.-F. Siffroi J.-P. et al.Loss-of-function mutations in LRRC6, a gene essential for proper axonemal assembly of inner and outer dynein arms, cause primary ciliary dyskinesia.Am. J. Hum. Genet. 2012; 91: 958-964Abstract Full Text Full Text PDF PubMed Scopus (135) Google Scholar, 16Horani A. Druley T.E. Zariwala M.A. Patel A.C. Levinson B.T. Van Arendonk L.G. Thornton K.C. Giacalone J.C. Albee A.J. Wilson K.S. et al.Whole-exome capture and sequencing identifies HEATR2 mutation as a cause of primary ciliary dyskinesia.Am. J. Hum. Genet. 2012; 91: 685-693Abstract Full Text Full Text PDF PubMed Scopus (126) Google Scholar, 17Panizzi J.R. Becker-Heck A. Castleman V.H. Al-Mutairi D.A. Liu Y. Loges N.T. Pathak N. Austin-Tse C. Sheridan E. Schmidts M. et al.CCDC103 mutations cause primary ciliary dyskinesia by disrupting assembly of ciliary dynein arms.Nat. Genet. 2012; 44: 714-719Crossref PubMed Scopus (182) Google Scholar, 18Merveille A.-C. Davis E.E. Becker-Heck A. Legendre M. Amirav I. Bataille G. Belmont J. Beydon N. Billen F. Clément A. et al.CCDC39 is required for assembly of inner dynein arms and the dynein regulatory complex and for normal ciliary motility in humans and dogs.Nat. Genet. 2011; 43: 72-78Crossref PubMed Scopus (251) Google Scholar, 19Becker-Heck A. Zohn I.E. Okabe N. Pollock A. Lenhart K.B. Sullivan-Brown J. McSheene J. Loges N.T. Olbrich H. Haeffner K. et al.The coiled-coil domain containing protein CCDC40 is essential for motile cilia function and left-right axis formation.Nat. Genet. 2011; 43: 79-84Crossref PubMed Scopus (223) Google Scholar, 20Bartoloni L. Blouin J.-L. Pan Y. Gehrig C. Maiti A.K. Scamuffa N. Rossier C. Jorissen M. Armengot M. Meeks M. et al.Mutations in the DNAH11 (axonemal heavy chain dynein type 11) gene cause one form of situs inversus totalis and most likely primary ciliary dyskinesia.Proc. Natl. Acad. Sci. USA. 2002; 99: 10282-10286Crossref PubMed Scopus (268) Google Scholar, 21Wirschell M. Olbrich H. Werner C. Tritschler D. Bower R. Sale W.S. Loges N.T. Pennekamp P. Lindberg S. Stenram U. et al.The nexin-dynein regulatory complex subunit DRC1 is essential for motile cilia function in algae and humans.Nat. Genet. 2013; 45: 262-268Crossref PubMed Scopus (150) Google Scholar, 22Castleman V.H. Romio L. Chodhari R. Hirst R.A. de Castro S.C.P. Parker K.A. Ybot-Gonzalez P. Emes R.D. Wilson S.W. Wallis C. et al.Mutations in radial spoke head protein genes RSPH9 and RSPH4A cause primary ciliary dyskinesia with central-microtubular-pair abnormalities.Am. J. Hum. Genet. 2009; 84: 197-209Abstract Full Text Full Text PDF PubMed Scopus (243) Google Scholar, 23Olbrich H. Schmidts M. Werner C. Onoufriadis A. Loges N.T. Raidt J. Banki N.F. Shoemark A. Burgoyne T. Al Turki S. et al.UK10K ConsortiumRecessive HYDIN mutations cause primary ciliary dyskinesia without randomization of left-right body asymmetry.Am. J. Hum. Genet. 2012; 91: 672-684Abstract Full Text Full Text PDF PubMed Scopus (209) Google Scholar However, in spite of these significant results, the disease remains unexplained in more than half of affected individuals from our PCD cohort. Cilia are evolutionarily conserved organelles that protrude from the surface of most eukaryotic cells. Their axoneme consists of nine peripheral microtubule doublets either surrounding a central pair of microtubules (i.e., “9+2” pattern) or not (i.e., “9+0” pattern). The 9+0 cilia are immotile, except in the embryonic node, a structure involved in the early establishment of the left-right asymmetry.24Nonaka S. Yoshiba S. Watanabe D. Ikeuchi S. Goto T. Marshall W.F. Hamada H. De novo formation of left-right asymmetry by posterior tilt of nodal cilia.PLoS Biol. 2005; 3: e268Crossref PubMed Scopus (221) Google Scholar The axoneme of motile cilia is characterized by the presence of inner dynein arms (IDAs) and outer dynein arms (ODAs), which are multiprotein ATPase complexes that are attached to the peripheral doublets and are essential for normal ciliary and flagellar movements.25Satir P. Christensen S.T. Overview of structure and function of mammalian cilia.Annu. Rev. Physiol. 2007; 69: 377-400Crossref PubMed Scopus (745) Google Scholar Radial spokes (RSs), which are present in 9+2 cilia, are T-shaped structures linking each peripheral microtubule doublet to the two central microtubules (C1 and C2), which are surrounded by a central sheath and participate in the formation of the central complex (CC),26Escalier D. Jouannet P. David G. Abnormalities of the ciliary axonemal complex in children: An ultrastructural and cinetic study in a series of 34 cases.Biol. Cell. 1982; 44: 271-282Google Scholar, 27Papon J.F. Coste A. Roudot-Thoraval F. Boucherat M. Roger G. Tamalet A. Vojtek A.M. Amselem S. Escudier E. A 20-year experience of electron microscopy in the diagnosis of primary ciliary dyskinesia.Eur. Respir. J. 2010; 35: 1057-1063Crossref PubMed Scopus (115) Google Scholar also called the central-pair complex.28Heuser T. Dymek E.E. Lin J. Smith E.F. Nicastro D. The CSC connects three major axonemal complexes involved in dynein regulation.Mol. Biol. Cell. 2012; 23: 3143-3155Crossref PubMed Scopus (59) Google Scholar, 29Carbajal-González B.I. Heuser T. Fu X. Lin J. Smith B.W. Mitchell D.R. Nicastro D. Conserved structural motifs in the central pair complex of eukaryotic flagella.Cytoskeleton (Hoboken). 2013; 70: 101-120Crossref PubMed Scopus (62) Google Scholar RSs and CCs are believed to serve as sensors that control beating, especially the waveform, both in motile cilia and in flagella.30Smith E.F. Yang P. The radial spokes and central apparatus: mechano-chemical transducers that regulate flagellar motility.Cell Motil. Cytoskeleton. 2004; 57: 8-17Crossref PubMed Scopus (213) Google Scholar According to data obtained in the flagellated chlorophyte Chlamydomonas reinhardtii, premature RSs are assembled into the cytoplasm and are dimerized along the length of cilia;31Diener D.R. Yang P. Geimer S. Cole D.G. Sale W.S. Rosenbaum J.L. Sequential assembly of flagellar radial spokes.Cytoskeleton (Hoboken). 2011; 68: 389-400Crossref PubMed Scopus (40) Google Scholar each RS dimer is composed of at least 23 proteins,32Yang P. Diener D.R. Yang C. Kohno T. Pazour G.J. Dienes J.M. Agrin N.S. King S.M. Sale W.S. Kamiya R. et al.Radial spoke proteins of Chlamydomonas flagella.J. Cell Sci. 2006; 119: 1165-1174Crossref PubMed Scopus (174) Google Scholar five of which belong to the RS head (the horizontal bar of the “T”), which interacts with the CC. In most individuals with PCD, the axonemal defect concerns the dynein arms and is related to mutations in different genes (DNAI14Pennarun G. Escudier E. Chapelin C. Bridoux A.M. Cacheux V. Roger G. Clément A. Goossens M. Amselem S. Duriez B. Loss-of-function mutations in a human gene related to Chlamydomonas reinhardtii dynein IC78 result in primary ciliary dyskinesia.Am. J. Hum. Genet. 1999; 65: 1508-1519Abstract Full Text Full Text PDF PubMed Scopus (295) Google Scholar [MIM 604366], DNAI25Loges N.T. Olbrich H. Fenske L. Mussaffi H. Horvath J. Fliegauf M. Kuhl H. Baktai G. Peterffy E. Chodhari R. et al.DNAI2 mutations cause primary ciliary dyskinesia with defects in the outer dynein arm.Am. J. Hum. Genet. 2008; 83: 547-558Abstract Full Text Full Text PDF PubMed Scopus (188) Google Scholar [MIM 605483], DNAH56Olbrich H. Häffner K. Kispert A. Völkel A. Volz A. Sasmaz G. Reinhardt R. Hennig S. Lehrach H. Konietzko N. et al.Mutations in DNAH5 cause primary ciliary dyskinesia and randomization of left-right asymmetry.Nat. Genet. 2002; 30: 143-144Crossref PubMed Scopus (423) Google Scholar [MIM 603335], NME87Duriez B. Duquesnoy P. Escudier E. Bridoux A.-M. Escalier D. Rayet I. Marcos E. Vojtek A.-M. Bercher J.-F. Amselem S. A common variant in combination with a nonsense mutation in a member of the thioredoxin family causes primary ciliary dyskinesia.Proc. Natl. Acad. Sci. USA. 2007; 104: 3336-3341Crossref PubMed Scopus (151) Google Scholar [MIM 607421], DNAL18Mazor M. Alkrinawi S. Chalifa-Caspi V. Manor E. Sheffield V.C. Aviram M. Parvari R. Primary ciliary dyskinesia caused by homozygous mutation in DNAL1, encoding dynein light chain 1.Am. J. Hum. Genet. 2011; 88: 599-607Abstract Full Text Full Text PDF PubMed Scopus (96) Google Scholar [MIM 610062], CCDC1149Onoufriadis A. Paff T. Antony D. Shoemark A. Micha D. Kuyt B. Schmidts M. Petridi S. Dankert-Roelse J.E. Haarman E.G. et al.UK10KSplice-site mutations in the axonemal outer dynein arm docking complex gene CCDC114 cause primary ciliary dyskinesia.Am. J. Hum. Genet. 2013; 92: 88-98Abstract Full Text Full Text PDF PubMed Scopus (130) Google Scholar, 10Knowles M.R. Leigh M.W. Ostrowski L.E. Huang L. Carson J.L. Hazucha M.J. Yin W. Berg J.S. Davis S.D. Dell S.D. et al.Genetic Disorders of Mucociliary Clearance ConsortiumExome sequencing identifies mutations in CCDC114 as a cause of primary ciliary dyskinesia.Am. J. Hum. Genet. 2013; 92: 99-106Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar [MIM 615038], DNAAF111Duquesnoy P. Escudier E. Vincensini L. Freshour J. Bridoux A.-M. Coste A. Deschildre A. de Blic J. Legendre M. Montantin G. et al.Loss-of-function mutations in the human ortholog of Chlamydomonas reinhardtii ODA7 disrupt dynein arm assembly and cause primary ciliary dyskinesia.Am. J. Hum. Genet. 2009; 85: 890-896Abstract Full Text Full Text PDF PubMed Scopus (127) Google Scholar, 12Loges N.T. Olbrich H. Becker-Heck A. Häffner K. Heer A. Reinhard C. Schmidts M. Kispert A. Zariwala M.A. Leigh M.W. et al.Deletions and point mutations of LRRC50 cause primary ciliary dyskinesia due to dynein arm defects.Am. J. Hum. Genet. 2009; 85: 883-889Abstract Full Text Full Text PDF PubMed Scopus (174) Google Scholar [MIM 613190], DNAAF213Omran H. Kobayashi D. Olbrich H. Tsukahara T. Loges N.T. Hagiwara H. Zhang Q. Leblond G. O’Toole E. Hara C. et al.Ktu/PF13 is required for cytoplasmic pre-assembly of axonemal dyneins.Nature. 2008; 456: 611-616Crossref PubMed Scopus (283) Google Scholar [MIM 612517], DNAAF314Mitchison H.M. Schmidts M. Loges N.T. Freshour J. Dritsoula A. Hirst R.A. O’Callaghan C. Blau H. Al Dabbagh M. Olbrich H. et al.Mutations in axonemal dynein assembly factor DNAAF3 cause primary ciliary dyskinesia.Nat. Genet. 2012; 44 (S1–S2): 381-389Crossref PubMed Scopus (190) Google Scholar [MIM 614566], LRRC615Kott E. Duquesnoy P. Copin B. Legendre M. Dastot-Le Moal F. Montantin G. Jeanson L. Tamalet A. Papon J.-F. Siffroi J.-P. et al.Loss-of-function mutations in LRRC6, a gene essential for proper axonemal assembly of inner and outer dynein arms, cause primary ciliary dyskinesia.Am. J. Hum. Genet. 2012; 91: 958-964Abstract Full Text Full Text PDF PubMed Scopus (135) Google Scholar [MIM 614930], HEATR216Horani A. Druley T.E. Zariwala M.A. Patel A.C. Levinson B.T. Van Arendonk L.G. Thornton K.C. Giacalone J.C. Albee A.J. Wilson K.S. et al.Whole-exome capture and sequencing identifies HEATR2 mutation as a cause of primary ciliary dyskinesia.Am. J. Hum. Genet. 2012; 91: 685-693Abstract Full Text Full Text PDF PubMed Scopus (126) Google Scholar [MIM 614864], CCDC10317Panizzi J.R. Becker-Heck A. Castleman V.H. Al-Mutairi D.A. Liu Y. Loges N.T. Pathak N. Austin-Tse C. Sheridan E. Schmidts M. et al.CCDC103 mutations cause primary ciliary dyskinesia by disrupting assembly of ciliary dynein arms.Nat. Genet. 2012; 44: 714-719Crossref PubMed Scopus (182) Google Scholar [MIM 614677], CCDC3918Merveille A.-C. Davis E.E. Becker-Heck A. Legendre M. Amirav I. Bataille G. Belmont J. Beydon N. Billen F. Clément A. et al.CCDC39 is required for assembly of inner dynein arms and the dynein regulatory complex and for normal ciliary motility in humans and dogs.Nat. Genet. 2011; 43: 72-78Crossref PubMed Scopus (251) Google Scholar [MIM 613798], and CCDC4019Becker-Heck A. Zohn I.E. Okabe N. Pollock A. Lenhart K.B. Sullivan-Brown J. McSheene J. Loges N.T. Olbrich H. Haeffner K. et al.The coiled-coil domain containing protein CCDC40 is essential for motile cilia function and left-right axis formation.Nat. Genet. 2011; 43: 79-84Crossref PubMed Scopus (223) Google Scholar [MIM 613799]). Some individuals have no detectable structural axonemal defect; mutations in DNAH1120Bartoloni L. Blouin J.-L. Pan Y. Gehrig C. Maiti A.K. Scamuffa N. Rossier C. Jorissen M. Armengot M. Meeks M. et al.Mutations in the DNAH11 (axonemal heavy chain dynein type 11) gene cause one form of situs inversus totalis and most likely primary ciliary dyskinesia.Proc. Natl. Acad. Sci. USA. 2002; 99: 10282-10286Crossref PubMed Scopus (268) Google Scholar (MIM 603339) or DRC121Wirschell M. Olbrich H. Werner C. Tritschler D. Bower R. Sale W.S. Loges N.T. Pennekamp P. Lindberg S. Stenram U. et al.The nexin-dynein regulatory complex subunit DRC1 is essential for motile cilia function in algae and humans.Nat. Genet. 2013; 45: 262-268Crossref PubMed Scopus (150) Google Scholar (MIM 615288) have been found in this phenotype. As for the PCD phenotype characterized by CC defects, it was present in 15% of individuals in our cohort.27Papon J.F. Coste A. Roudot-Thoraval F. Boucherat M. Roger G. Tamalet A. Vojtek A.M. Amselem S. Escudier E. A 20-year experience of electron microscopy in the diagnosis of primary ciliary dyskinesia.Eur. Respir. J. 2010; 35: 1057-1063Crossref PubMed Scopus (115) Google Scholar, 33Vallet C. Escudier E. Roudot-Thoraval F. Blanchon S. Fauroux B. Beydon N. Boulé M. Vojtek A.M. Amselem S. Clément A. Tamalet A. Primary ciliary dyskinesia presentation in 60 children according to ciliary ultrastructure.Eur. J. Pediatr. 2013; 172: 1053-1060Crossref PubMed Scopus (22) Google Scholar The molecular basis of this particular phenotype, which is also characterized by the fact that affected individuals never have laterality defects, remains unexplained in the majority of cases. So far, three genes have been involved in this phenotype. Two of them encode RS-head proteins: RSPH4A (MIM 612647), which has been found to be mutated in nine families,22Castleman V.H. Romio L. Chodhari R. Hirst R.A. de Castro S.C.P. Parker K.A. Ybot-Gonzalez P. Emes R.D. Wilson S.W. Wallis C. et al.Mutations in radial spoke head protein genes RSPH9 and RSPH4A cause primary ciliary dyskinesia with central-microtubular-pair abnormalities.Am. J. Hum. Genet. 2009; 84: 197-209Abstract Full Text Full Text PDF PubMed Scopus (243) Google Scholar, 34Ziętkiewicz E. Bukowy-Bieryłło Z. Voelkel K. Klimek B. Dmeńska H. Pogorzelski A. Sulikowska-Rowińska A. Rutkiewicz E. Witt M. Mutations in radial spoke head genes and ultrastructural cilia defects in East-European cohort of primary ciliary dyskinesia patients.PLoS ONE. 2012; 7: e33667Crossref PubMed Scopus (43) Google Scholar and RSPH9 (MIM 612648), in which the same mutation has been identified in three Bedouin families;22Castleman V.H. Romio L. Chodhari R. Hirst R.A. de Castro S.C.P. Parker K.A. Ybot-Gonzalez P. Emes R.D. Wilson S.W. Wallis C. et al.Mutations in radial spoke head protein genes RSPH9 and RSPH4A cause primary ciliary dyskinesia with central-microtubular-pair abnormalities.Am. J. Hum. Genet. 2009; 84: 197-209Abstract Full Text Full Text PDF PubMed Scopus (243) Google Scholar, 35Alsaadi M.M. Gaunt T.R. Boustred C.R. Guthrie P.A.I. Liu X. Lenzi L. Rainbow L. Hall N. Alharbi K.K. Day I.N.M. From a single whole exome read to notions of clinical screening: primary ciliary dyskinesia and RSPH9 p.Lys268del in the Arabian Peninsula.Ann. Hum. Genet. 2012; 76: 211-220Crossref PubMed Scopus (18) Google Scholar the third gene, HYDIN (MIM 610812), has recently been involved in PCD with central-sheath defects.23Olbrich H. Schmidts M. Werner C. Onoufriadis A. Loges N.T. Raidt J. Banki N.F. Shoemark A. Burgoyne T. Al Turki S. et al.UK10K ConsortiumRecessive HYDIN mutations cause primary ciliary dyskinesia without randomization of left-right body asymmetry.Am. J. Hum. Genet. 2012; 91: 672-684Abstract Full Text Full Text PDF PubMed Scopus (209) Google Scholar We undertook this study with the aim of identifying additional molecular defects involved in PCD with CC defects by combining homozygosity mapping and whole-exome sequencing (WES). The current study was approved by the Ile-de-France ethics committee (CPP07729), and written informed consent was obtained from all individuals and/or their parents. Forty-eight unrelated families (containing 62 affected individuals) in our PCD cohort were affected by CC defects; none of them showed laterality defects. In all of them, the axonemal defect, which affected a variable proportion of cilia in each individual, was confirmed by transmission electronic microscopy (TEM), which showed the absence of one or both microtubules of the central pair (9+1 or 9+0 pattern), sometimes associated with the internalization of one peripheral doublet (8+1 pattern).27Papon J.F. Coste A. Roudot-Thoraval F. Boucherat M. Roger G. Tamalet A. Vojtek A.M. Amselem S. Escudier E. A 20-year experience of electron microscopy in the diagnosis of primary ciliary dyskinesia.Eur. Respir. J. 2010; 35: 1057-1063Crossref PubMed Scopus (115) Google Scholar, 33Vallet C. Escudier E. Roudot-Thoraval F. Blanchon S. Fauroux B. Beydon N. Boulé M. Vojtek A.M. Amselem S. Clément A. Tamalet A. Primary ciliary dyskinesia presentation in 60 children according to ciliary ultrastructure.Eur. J. Pediatr. 2013; 172: 1053-1060Crossref PubMed Scopus (22) Google Scholar, 36Tamalet A. Clement A. Roudot-Thoraval F. Desmarquest P. Roger G. Boulé M. Millepied M.C. Baculard T.A. Escudier E. Abnormal central complex is a marker of severity in the presence of partial ciliary defect.Pediatrics. 2001; 108: E86Crossref PubMed Scopus (38) Google Scholar Moreover, this ultrastructural phenotype is characterized by the fact that the RSs are present in cilia with a CC but absent from cilia with no CC. The ultrastructural defect was confirmed on repeated airway biopsies performed at different levels (i.e., nasal and bronchial sampling). By screening the genes already involved in this PCD phenotype, we identified mutations in 11 of these 48 families: RSPH4A mutations in seven families and RSPH9 mutations in four families (Table S1, available online). One individual (DCP940 from family DC564) of the 37 unrelated families (44 affected individuals) with no mutations in RSPH4A or RSPH9 was selected for WES. This female person, who was born to a consanguineous union, presented with a sinopulmonary syndrome beginning in early childhood, subfertility, and no situs inversus. She exhibited a very marked ultrastructural phenotype in which 60% of cilia showed CC defects (Figure 1 and Table 1); total ciliary immotility was observed in ciliated cells obtained by airway brushing. Her genomic DNA was first genotyped with the HumanCytoSNP-12 chip from Illumina, and the data were analyzed with GenomeStudio and CNVPartition 3.1.6 software (Illumina), as well as with an in-house script designed to identify homozygous regions. Three large homozygous regions spanning 13.8, 8.3, and 6 Mb, were identified on chromosomes 1, 21, and 8, respectively, and contained a total of 329 genes (Figure S1).Table 1Phenotypic Features of Individuals with Identified RSPH1 MutationsFamily (Origin)IndividualKnown ConsanguinityGenderAirway DiseaseFertilityNOaNasal NO was measured during apnea with the use of a chemiluminscence NO analyzer (NIOX Flex, Aerocrine, and Endono 8000, Seres). The mean value of the plateau was recorded. NO values above 100 nl/min were considered normal. (nl/min)Abnormal Cilia (TEM)bThe method used is described in Papon et al.27 (%)Allele 1Allele 2DC564 (North African)DCP940yesfemaleNRD, bronchiectasis, rhinosinusitis, otitissubfertilityNP60c.85G>T (p.Glu29∗)c.85G>T (p.Glu29∗)DC18 (European)DCP876nomaleCOPD, bronchiectasis, rhinosinusitisNRNP50c.85G>T (p.Glu29∗)c.366−3C>ADC129 (European)DCP781nomalebronchiectasis, rhinosinusitis, otitisasthenospermialowcNO value not available.58c.366−3C>Ac.407_410delAGTA (p.Lys136Metfs∗6)DC337 (European)DCP1057nofemaleCOPD, bronchiectasis, lobectomy, rhinosinusitis, otitissubfertilityNP23c.275−2A>C (p.Gly92Alafs∗10)c.727+5G>A (p.Ala244Valfs∗22)DC402 (European)DCP638nomalechronic bronchitis, rhinosinusitisNRNPNPc.275−2A>C (p.Gly92Alafs∗10)c.275−2A>C (p.Gly92Alafs∗10)DCP639nofemalechronic bronchitis, rhinosinusitisNRNP34c.275−2A>C (p.Gly92Alafs∗10)c.275−2A>C (p.Gly92Alafs∗10)DC455 (European)DCP729nofemaleNRD, bronchiectasis, lobectomy, rhinosinusitis, otitissubfertility8022c.85G>T (p.Glu29∗)c.308G>A (p.Gly103Asp)DC477 (North African)DCP775yesmalebronchiectasis, pneumonia, rhinosinusitis, otitisNR3734.5c.85G>T (p.Glu29∗)c.85G>T (p.Glu29∗)DC530 (European)DCP873yesmalebronchiectasis, rhinosinusitisNDNP70c.275−2A>C (p.Gly92Alafs∗10)c.275−2A>C (p.Gly92Alafs∗10)DC645 (European)DCP1153nomaleNRD, chronic bronchitis, otitisNRNP25.5c.275−2A>C (p.Gly92Alafs∗10)c.275−2A>C (p.Gly92Alafs∗10)DCP1154nomalebronchiectasis, otitisNR4519c.275−2A>C (p.Gly92Alafs∗10)c.275−2A>C (p.Gly92Alafs∗10)DC651 (European)DCP1064nofemaleNRD, COPD, bronchiectasis, rhinosinusitis, otitisNR50044c.85G>T (p.Glu29∗)c.366G>A (p.[Arg122Serfs∗22, p.=])Abbreviations are as follows: NO, nitric oxide; TEM, transmission electron microscopy; NRD, neonatal respiratory distress; NP, not performed; COPD, chronic obstructive pulmonary disease; NR, not relevant; and ND, not determined.a Nasal NO was measured during apnea with the use of a chemiluminscence NO analyzer (NIOX Flex, Aerocrine, and Endono 8000, Seres). The mean value of the plateau was recorded. NO values above 100 nl/min were considered normal.b The method used is described in Papon et al.27Papon J.F. Coste A. Roudot-Thoraval F. Boucherat M. Roger G. Tamalet A. Vojtek A.M. Amselem S. Escudier E. A 20-y
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