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ORIGINAL ARTICLE
Novel IFT122 mutation associated with impaired
ciliogenesis and cranioectodermal dysplasia
Anas M. Alazami1,a, Mohammed Zain Seidahmed2,a, Fatema Alzahrani1, Adam O. Mohammed3 &
Fowzan S. Alkuraya1,4
1
Department
Department
3
Department
4
Department
2
of
of
of
of
Genetics, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
Pediatrics, Security Forces Hospital, Riyadh, Saudi Arabia
Pediatrics, National Guard Hospital, Dammam, Saudi Arabia
Anatomy and Cell Biology, College of Medicine, Alfaisal University, Riyadh, Saudi Arabia
Keywords
Ciliopathy, craniosynostosis, intraflagellar
transport.
Correspondence
Fowzan S. Alkuraya, Developmental Genetics
Unit, King Faisal Specialist Hospital and
Research Center, MBC-03 PO BOX 3354,
Riyadh 11211, Saudi Arabia.
Tel: +966 1 442 7875;
Fax: +966 1 442 4585;
E-mail: [email protected]
Abstract
Cranioectodermal dysplasia (CED) is a very rare autosomal recessive disorder
characterized by a recognizable craniofacial profile in addition to ectodermal
manifestations involving the skin, hair, and teeth. Four genes are known to be
mutated in this disorder, all involved in the ciliary intraflagellar transport confirming that CED is a ciliopathy. In a multiplex consanguineous family with
typical CED features in addition to intellectual disability and severe cutis laxa,
we used autozygosity-guided candidate gene analysis to identify a novel homozygous mutation in IFT122, and demonstrated impaired ciliogenesis in patient
fibroblasts. This report on IFT122 broadens the phenotype of CED and expands
its allelic heterogeneity.
Funding Information
This study was funded in part by a KACST
grant 09-MED941-20 (FSA).
a
These two authors have contributed equally
to this work.
Received: 24 August 2013; Revised: 2
October 2013; Accepted: 4 October 2013
Molecular Genetics & Genomic Medicine
2014; 2(2): 103–106
doi: 10.1002/mgg3.44
Brief Report
Cranioectodermal dysplasia (CED) is a skeletal dysplasia
characterized by typical craniofacial features in the form of
dolichocephaly, sagittal craniosynostosis, and facial dysmorphism (frontal bossing, epicanthic folds, flat nose with
anteverted nares, and everted lower lip), and skeletal anomalies in the form of narrow thorax and short extremities, in
addition to ectodermal dysplastic features in the form of
thin sparse scalp hair and micro/hypodontia (Levin et al.
1977). Since its first description in 1975 (Sensenbrenner
et al. 1975), less than 50 cases have been reported indicating the rarity of this syndrome. The subsequent expansion
of the phenotype to include corpus callosal dysgenesis,
hepatic fibrosis, nephrophthisis, and retinitis pigmentosa
made it likely that CED is a ciliopathy (Konstantinidou
et al. 2009). Indeed, genetic studies confirmed this hypothesis by identifying mutations in four genes all encoding
components of the ciliary intraflagellar transport complexA (IFT122, WDR35, C14ORF179, and WDR19) (Gilissen
et al. 2010; Walczak-Sztulpa et al. 2010; Arts et al. 2011;
Bredrup et al. 2011). Not unlike other ciliopathy disease
genes, mutations in some of these genes have been observed
to cause overlapping ciliopathy phenotypes such as the
finding of WDR35 mutations in short rib-polydactyly syndrome (Mill et al. 2011). Thus, additional reports of mutations in these genes will be critical to our understanding of
the spectrum of resulting phenotypes.
ª 2013 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.
This is an open access article under the terms of the Creative Commons Attribution License, which permits use,
distribution and reproduction in any medium, provided the original work is properly cited.
103
Novel IFT122 Mutation
A. M. Alazami et al.
A
B
C
Figure 1. The family reported in this study. (A) Pedigree of the family with the index case boxed in red. Facial and hand profiles for the two
male patients are given in (B) and (C). Note the typical facial features (prominent forehead, depressed nasal bridge, anteverted nares, and everted
lower lip) and typical hand features (brachdactyly, single interphalangeal crease for some fingers and clinodactyly). Please note the redundant
palm skin in the lower panels, consistent with cutis laxa.
Table 1. Clinical features.
Clinical feature
Patient 1
Patient 2
Patient 3
Age
Frequent chest infection
Sagittal craniosynostosis
Dolichocephaly
Epicanthal folds
Broad nasal bridge
Anteverted nares
Everted lower lip
Micro/Hypodontia
Narrow thorax
Short limbs
Brachydactyly
Clinodactyly
Joint hypermobility
Cutis laxa
Osteoporosis
Retinal dystrophy
Nephronophthisis
Congenital heart disease
Frequent chest infection
Others
22 years
Female
+
+
+
+
+
9 years
Male
+
2 years
Male
+
+
+
+
+
+
+
+
+
+
+
+
?
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
ESRF
+
ID
+
ID
+
ESRF, end stage renal failure; ID, intellectual disability; ?, unknown.
104
In this report, we describe a multiplex family containing three affected siblings born to healthy first cousin
Saudi parents (Fig. 1A). In addition to the classical
features of CED (Table 1, Fig. 1B and C), they all had
markedly lax skin with joint laxity fulfilling the clinical
definition of cutis laxa, but none had evidence of retinal
involvement and only the oldest patient developed
end-stage renal failure. In addition, the two older siblings
have confirmed intellectual disability with intelligence
quotient of 70, a feature that has not been reported
in CED. Given the consanguinity of the parents and
the genetic heterogeneity of CED, we pursued autozygome-guided candidate gene sequencing essentially as
described before (Alkuraya 2012). The autozygome of
the three siblings overlapped on just two genomic
regions (chr3:102718000-136160000, and chr11:1339500027766000, hg19 build) (Fig. 2A). IFT122 was the only
known CED disease gene mapping to either of the two
regions. Direct sequencing of IFT122 revealed a novel
homozygous missense mutation (c.1868G>T, p.G623V;
NM_052985.2) (Fig. 2B). The affected amino acid residue
is highly conserved across species (Fig. 2D), is associated
with high in silico pathogenicity scores (PolyPhen 1.0 and
ª 2013 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.
Novel IFT122 Mutation
A. M. Alazami et al.
Figure 2. A missense mutation in IFT122 causes cranioectodermal dysplasia in the reported family. (A) Homozygosity Mapper reveals two regions
of autozygosity which are shared by the three patients genomewide. The IFT122 locus is indicated with an arrow. (B) Sequence chromatogram of
one control individual and one patient, with the site of mutation denoted by an asterisk. (C) Schematic of the IFT122 protein indicating the
position of WD40 domains. The mutation reported here is located with a green arrow, while all previously published mutations are given below
the schematic (red arrowheads). (D) Protein alignment data reveal that the affected amino acid residue is highly conserved across species, down
to moss and trichoplax.
SIFT 0.0) and was absent from 374 Saudi control chromosomes.
In order to confirm the pathogenicity of this mutation,
we cultured fibroblasts from the foreskin of the younger
brother following circumcision and proceeded with
stress-induced ciliogenesis assay, essentially as described
before (Shaheen et al. 2012). In addition to observing a
marked reduction in ciliated fibroblasts, existing cilia in
patient fibroblasts were also smaller compared to control
age-matched fibroblasts (Fig. 3A and B). These reductions
in ciliary frequency and length were confirmed to be
highly significant (Fig. 3C and D). Thus, it appears that
G623V is associated with a similar ciliogenesis defect to
the ones reported in the original description of IFT122 as
a novel CED disease gene (Walczak-Sztulpa et al. 2010).
The molecular confirmation of CED in this expands the
allelic heterogeneity of IFT122 in CED (Tsurusaki et al.
2013). In addition, the remarkable cutis laxa phenotype
in this family supports one previous report of CED with
cutis laxa (Fry et al. 2009) thus confirming that this is a
bona fide phenotypic aspect of the disease albeit at low
frequency. Finally, this is the first instance of confirmed
intellectual disability in CED, which suggests that intellectual disability may be a low-frequency feature of this
disorder.
Acknowledgments
We thank the family for their enthusiastic participation.
We also thank the Sequencing and Genotyping Core
Facilities at KFSH&RC for their expert technical assistance.
Conflict of Interest
None declared.
ª 2013 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.
105
Novel IFT122 Mutation
A. M. Alazami et al.
Figure 3. Primary cilia in patient cells exhibit reduced frequency and length as compared with control cells. Primary cilia from serum-starved
primary control (A) and patient (B) fibroblasts, stained with anti-acetylated tubulin (green) and counterstained with 4’,6-diamidino-2-phenylindole
(blue). (C) Patient cells show significantly decreased ciliogenesis versus control cells (P < 0.0001, Fisher’s exact test). All cells within a total of six
fields, representing two independent experiments, were counted for each cell line. Error bars represent the standard error of the mean. (D)
Patient cells show significantly decreased ciliary length versus control cells (P < 0.002, unpaired t-test). Error bars represent the standard error of
the mean.
References
Alkuraya, F. S. 2012. Discovery of rare homozygous mutations
from studies of consanguineous pedigrees. Curr. Protoc.
Hum. Genet. Chapter 6:Unit6 12.
Arts, H. H., E. M. Bongers, D. A. Mans, S. E. van Beersum,
M. M. Oud, E. Bolat, et al. 2011. C14ORF179 encoding
IFT43 is mutated in Sensenbrenner syndrome. J. Med.
Genet. 48:390–395.
Bredrup, C., S. Saunier, M. M. Oud, T. Fiskerstrand, A.
Hoischen, D. Brackman, et al. 2011. Ciliopathies with
skeletal anomalies and renal insufficiency due to
mutations in the IFT-A gene WDR19. Am. J. Hum. Genet.
89:634–643.
Fry, A. E., C. Klingenberg, J. Matthes, K. Heimdal, R. C.
Hennekam, and D. T. Pilz. 2009. Connective tissue
involvement in two patients with features of cranioectodermal
dysplasia. Am. J. Med. Genet. A 149A:2212–2215.
Gilissen, C., H. H. Arts, A. Hoischen, L. Spruijt, D. A. Mans,
P. Arts, et al. 2010. Exome sequencing identifies WDR35
variants involved in Sensenbrenner syndrome. Am. J. Hum.
Genet. 87:418–423.
Konstantinidou, A. E., H. Fryssira, S. Sifakis, C. Karadimas,
P. Kaminopetros, G. Agrogiannis, et al. 2009.
Cranioectodermal dysplasia: a probable ciliopathy. Am.
J. Med. Genet. A 149A:2206–2211.
106
Levin, L. S., J. C. S. Perrin, L. Ose, J. P. Dorst, J. D. Miller,
and V. A. McKusick. 1977. A heritable syndrome of
craniosynostosis, short thin hair, dental abnormalities, and
short limbs: cranioectodermal dysplasia. J. Pediatr.
90:55–61.
Mill, P., P. J. Lockhart, E. Fitzpatrick, H. S. Mountford,
E. A. Hall, M. A. Reijns, et al. 2011. Human and mouse
mutations in WDR35 cause short-rib polydactyly syndromes
due to abnormal ciliogenesis. Am. J. Hum. Genet. 88:508–
515.
Sensenbrenner, J. A., J. P. Dorst, and R. P. Owens. 1975. New
syndrome of skeletal, dental and hair anomalies. Birth
Defects Orig. Artic. Ser. 11:372–379.
Shaheen, R., E. Faqeih, H. E. Shamseldin, R. R. Noche, A.
Sunker, M. J. Alshammari, et al. 2012. POC1A truncation
mutation causes a ciliopathy in humans characterized by
primordial dwarfism. Am. J. Hum. Genet. 91:330–336.
Tsurusaki, Y., R. Yonezawa, M. Furuya, G. Nishimura, R.
Pooh, M. Nakashima, et al. 2013. Whole exome sequencing
revealed biallelic IFT122 mutations in a family with CED1
and recurrent pregnancy loss. Clin. Genet. [Epub ahead of
print].
Walczak-Sztulpa, J., J. Eggenschwiler, D. Osborn, D. A. Brown, F.
Emma, C. Klingenberg, et al. 2010. Cranioectodermal
Dysplasia, Sensenbrenner syndrome, is a ciliopathy caused by
mutations in the IFT122 gene. Am. J. Hum. Genet. 86:949–956.
ª 2013 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.