Duran et al. Cilia (2017) 6:7 DOI 10.1186/s13630-017-0051-y Cilia RESEARCH Open Access Mutations in IFT‑A satellite core component genes IFT43 and IFT121 produce short rib polydactyly syndrome with distinctive campomelia Ivan Duran1,9, S. Paige Taylor2, Wenjuan Zhang4, Jorge Martin1, Faisal Qureshi6, Suzanne M. Jacques6, Robert Wallerstein7, Ralph S. Lachman5, Deborah A. Nickerson8, Michael Bamshad8, Daniel H. Cohn1,4,5 and Deborah Krakow1,2,3,5* Abstract Background: Skeletal ciliopathies comprise a spectrum of ciliary malfunction disorders that have a profound efect on the skeleton. Most common among these disorders is short rib polydactyly syndrome (SRPS), a recessively inher- ited perinatal lethal condition characterized by a long narrow chest, markedly shortened long bones, polydactyly and, often, multi-organ system involvement. SRPS shows extensive locus heterogeneity with mutations in genes encoding proteins that participate in cilia formation and/or function. Results: Herein we describe mutations in IFT43, a satellite member of the retrograde IFT-A complex, that produce a form of SRPS with unusual bending of the ribs and appendicular bones. These newly described IFT43 mutations disrupted cilia formation, produced abnormalities in cartilage growth plate architecture thus contributing to altered endochondral ossifcation. We further show that the IFT43 SRPS phenotype is similar to SRPS resulting from mutations in the gene encoding IFT121 (WDR35), a direct interactor with IFT43. Conclusions: This study defnes a new IFT43-associated phenotype, identifying an additional locus for SRPS. The data demonstrate that IFT43 is essential for ciliogenesis and that the mutations disrupted the orderly proliferation and diferentiation of growth plate chondrocytes, resulting in a severe efect on endochondral ossifcation and miner- alization. Phenotypic similarities with SRPS cases resulting from mutations in the gene encoding the IFT43 direct interacting protein IFT121 suggests that similar mechanisms may be disrupted by defects in these two IFT-A satellite interactors. Keywords: Short rib polydactyly syndrome, SRPS, Cilia, Skeletal ciliopathy, IFT43, IFT121, Intrafagellar transport, IFT, IFT-A complex, Retrograde transport, Cartilage Background [1]. Te disorders exhibit overlapping clinical and radio- A spectrum of skeletal dysplasias results from muta- graphic fndings, but within this group there are distinct tions in genes that participate in primary cilia forma- phenotypes that include short rib polydactyly syndrome tion and/or function. Tese disorders are referred to as (SRPS), asphyxiating thoracic dystrophy (ATD) or Jeune ciliary disorders of the skeleton or skeletal ciliopathies syndrome, Sensenbrenner syndrome or cranioectodermal dysplasia (CED), Ellis-van Creveld syndrome (EVC), and Weyers acrofacial dysostosis [1]. Among these, the perina- *Correspondence: [email protected] tal lethal SRPS is the most severe and is characterized by 1 Department of Orthopaedic Surgery, David Gefen School of Medicine a long narrow thorax, markedly shortened long bones and at the University of California at Los Angeles, Los Angeles, CA 90095, USA Full list of author information is available at the end of the article frequent polydactyly. Extra-skeletal defects also occur in © The Author(s) 2017. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/ publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Duran et al. Cilia (2017) 6:7 Page 2 of 13 the neurologic, cardiac, gastrointestinal, and genitourinary core components of the complex, IFT122, IFT140, and systems. SRPS radiographic fndings include horizontal IFT144, and three satellite components, IFT43, IFT121, ribs, an abnormal acetabulum, small iliac bones, hypo- and IFT139 [29, 30], as shown in Fig. 3e. As characterized plastic or absent fbulae and disharmonious ossifcation in Chlamydomonas reinhardtii, IFT-A complex stability of the skeleton. Based primarily on radiographic fndings, appears to be dependent on an intact core, as loss of IFT122 SRPS has been further subdivided into type I/III (Saldino- resulted in decreased levels of the other core members, Noonan/Verma-Naumof), type II (Mohr-Majewski), and IFT140 and IFT144, undetectable levels of satellite mem- type IV (Beemer-Langer) [1]. Asphyxiating thoracic dys- bers IFT121 and IFT139, and abnormal cytosolic locali- trophy (ATD) is phenotypically similar to SRPS, but with zation of IFT43 [29]. By contrast, loss of satellite member less severe overall features, and about one-third of afected IFT121 did not destabilize the IFT-A core complex, but did individuals survive the neonatal period [2]. result in dissociation of IFT43 from the IFT-A core and its Mutations in a variety of ciliary genes have been associ- abnormal localization to the cytosol. Tus, incorporation of ated with SRPS/ATD phenotypic spectrum, refecting the IFT43 into the IFT-A complex requires both an intact core diferent components of ciliary function including those are and the presence of IFT121 [31]. involved in anterograde and retrograde intrafagellar trans- In a cohort of skeletal ciliopathy patients, we identifed port, IFT172 [OMIM 607386], IFT80 [OMIM 611177], IFT43 mutations in two genetically independent cases of IFT52 [OMIM 617094], IFT81 [OMIM 605489], IFT140 SRPS. Te clinical and radiographic phenotype was con- [OMIM 614620], IFT144/WDR19 [OMIM 608151], sistent with a diagnosis of SRPS type II, but there were DYNC2H1 [OMIM 603297], DYNC2LI1 [OMIM 617083], additional unusual abnormalities that included bent ribs WDR34 [OMIM 613363], WDR60 [OMIM 615462], and long bones, and under mineralization of the calva- TCTEX1D2, IFT121/WDR35 [OMIM 613602], TTC21B rium. A very similar radiographic phenotype was identi- [OMIM 612014], and those that have centrosomal roles, fed in two cases of SRPS resulting from IFT121 (WDR35) KIAA0586 [OMIM 610178], CEP120 [OMIM 613446], mutations. Similar to the fndings in C. reinhardtii, cells ICK [OMIM 612325], INTU [OMIM 610621], and NEK1 derived from patients with IFT43 mutations showed that [OMIM 604588] [3–22]. Mutations in several of the same loss of functional IFT43 did not impact the stability of the genes that produce SRPS and ATD also result in CED, a IFT-A core complex; however, mutations in the IFT43 related nonlethal autosomal recessive disorder primarily satellite complex partner IFT121 destabilized IFT43. Te distinguished from ATD by craniofacial and ectodermal data thus identify additional genetic and phenotypic het- abnormalities that include craniosynostosis, dolichoce- erogeneity in SRPS and demonstrate that the hierarchy of phaly, telecanthus, low set ears, hypo/microdontia, abnor- IFT-A complex formation is evolutionarily conserved. mal nails and hair, and a high incidence of development of renal symptoms [23]. Mutations producing either SRPS/ Methods ATD spectrum and/or CED have been identifed in mem- Collection of human samples, ethics, consent, bers of the ciliary intrafagellar transport-A (IFT-A) com- and permissions plex (Table 1): IFT144/WDR19 [OMIM 608151], IFT122/ Under an approved University of California at Los Ange- WDR140/WDR10 [OMIM 218330], IFT121/WDR35 les human subjects protocol, informed consent was [OMIM 613610], and IFT43 [OMIM 614099] that has thus obtained from the participants and clinical information, far only been associated with CED [23–27]. Te IFT-A radiographs, blood and tissue samples were obtained. All complex [27, 28], together with the dynein 2 motor com- experimental protocols and methods were carried out in plex, mediates retrograde transport in cilia. Tere are three accordance with institutional guidelines and regulations Table 1 Ciliopathies caused by mutations in IFT-A genes IFT-A gene/syndrome ATD or Jeune syndrome SRPS CED Other ciliopathies IFT122 CED [26] IFT140 Jeune syndrome [12, 39] Mainzer-Saldino [12, 39]a IFT144/WDR34 Jeune syndrome [6, 40] CED [6] Senior-Loken Syndrome [6, 41, 42] IFT139/TTC21B Jeune syndrome [17] Nephronophthisis 12 [17] IFT121/WDR35 SRPS [13] CED [25] IFT43 SRPS present report CED [27] a Now considered the same disorder Jeune syndrome, SRPS, short rib polydactyly syndrome, CED, cranioectodermal dysplasia, ATD, asphyxiating thoracic dysplasia Duran et al. Cilia (2017) 6:7 Page 3 of 13 and approved by the UCLA Institutional Review Board performed after starving cells in 0.5% serum for 48 h and Biosafety Committees. prior to cell collection. Exome analysis Western blot analysis DNA was isolated and submitted to the University of Each antibody was tested in at least three experimental Washington Center for Mendelian Genomics for library replicates from independent cultures. For the analyses, preparation and exome sequencing in a cohort of skel- there were two independent cases with either muta- etal ciliopathy cases. Te samples were barcoded, cap- tions in IFT43 or IFT121 mutations. In each western tured using the NimbleGen SeqCap EZ Exome Library blot analysis, there were at least three experimental v2.0 probe library targeting 36.5 Mb of genome, and replicates with technical replicates.