Hum Genet (2015) 134:815–822 DOI 10.1007/s00439-015-1561-6
ORIGINAL INVESTIGATION
ARL6IP6, a susceptibility locus for ischemic stroke, is mutated in a patient with syndromic Cutis Marmorata Telangiectatica Congenita Iman S. Abumansour1 · Hadia Hijazi1 · Anas Alazmi1 · Fatma Alzahrani1 · Fahad A. Bashiri2 · Hamdy Hassan3 · Mohammed Alhaddab4 · Fowzan S. Alkuraya1,5
Received: 27 February 2015 / Accepted: 27 April 2015 / Published online: 10 May 2015 © Springer-Verlag Berlin Heidelberg 2015
Abstract Cutis Marmorata Telangiectatica Congenita (CMTC) is a congenital localized or generalized vascular anomaly, usually sporadic in occurrence. It can be associated with other cutaneous or systemic manifestations. About 300 cases have been reported. The molecular etiology remains largely unknown. The main purpose of this study is to delineate the molecular basis for a syndromic CMTC phenotype in a consanguineous Saudi family. Clinical phenotyping including detailed neurological imaging, followed by autozygosity mapping and trio whole exome sequencing (WES) are also studied. We have identified a homozygous truncating mutation in ARL6IP6 as the likely cause of a syndromic form of CMTC associated with major dysmorphism, developmental delay, transient ischemic attacks and cerebral vascular malformations. This gene
I. S. Abumansour and H. Hijazi have contributed equally to this work. * Fowzan S. Alkuraya
[email protected] 1
Developmental Genetics Unit, Department of Genetics, King Faisal Specialist Hospital and Research Center, MBC‑03 PO BOX 3354, Riyadh 11211, Saudi Arabia
2
Division of Neurology, Department of Pediatrics, King Khalid University Hospital and College of Medicine, King Saud University, Riyadh, Saudi Arabia
3
Department of Radiology, King Khalid University Hospital and College of Medicine, King Saud University, Riyadh, Saudi Arabia
4
Department of Dermatology, King Khalid University Hospital and College of Medicine, King Saud University, Riyadh, Saudi Arabia
5
Department of Anatomy and Cell Biology, College of Medicine, Alfaisal University, Riyadh, Saudi Arabia
was previously implicated by genome wide association study (GWAS) as a susceptibility locus to ischemic stroke in young adults. We identify ARL6IP6 as a novel candidate gene for a syndromic form of CMTC. This suggests that ischemic stroke or transient ischemic attacks (TIA) may represent, at least in some cases, the mild end of a phenotypic spectrum that has at its severe end autosomal recessive CMTC. This finding contributes to a growing appreciation of the continuum of Mendelian and common complex diseases.
Introduction Diffuse temporary mottling appearance of skin when exposed to cold is a well-known phenomena in normal infants and many adults. It represents an exaggerated vasomotor response of skin capillaries giving rise to a reticulated marble-like appearance of skin, which is called Cutis Marmorata (CM). Congenital presence of persistent CM was first recognized in 1922 by Van Lohuizen and named as Cutis Marmorata Telangiectatica Congenita (CMTC) (Van Lohuizen 1922). Similar to CM, CMTC may be accentuated by cold. However, unlike CM, the cutaneous abnormalities do not usually resolve with rewarming and may be associated with telangiectasia, phlebectasia, skin atrophy and ulceration. Since the first description, more than 300 cases are reported in the literature (Amitai et al. 2000; Gerritsen et al. 2000). Some congenital anomalies are seen more frequently in association with CMTC with estimates ranging widely between 19 and 70 % in different series (Amitai et al. 2000; Gerritsen et al. 2000; Kienast and Hoeger 2009). Previously observed non-vasculocutaneous anomalies included body asymmetry, transverse limb defects, ocular anomalies mainly glaucoma, hypospadias,
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and cardiac defects. Neurological manifestations reported in literature were seizures, psychomotor retardation and intellectual disability. Recurrent strokes and stroke-like episodes were also reported, sometimes in association with a specific cerebral occlusive vasculopathy pattern known as Moyamoya disease (Van Schaik et al. 2015; Gruppo et al. 1998; Tsuruta et al. 1999). Despite the many cases reported in literature, the molecular cause and pathogenesis of CMTC have remained unknown with the exception of a few syndromes (see below). Potential mechanisms proposed to explain the usually sporadic occurrence of CMTC include autosomal dominant mutations with reduced penetrance or variable expressivity, multifactorial inheritance, or a lethal mutation surviving only by mosaicism. In this report, we describe the clinical and molecular characterization of a patient with a syndromic form of CMTC that led to the identification of biallelic loss-of-function mutation in ARL6IP6 as the likely cause.
Subjects and methods Human subjects The patient (12DG0006) received full clinical genetics, neurological and dermatological evaluations, including laboratory and radiological tests. The patient, his unaffected siblings and parents were recruited using a KFSHRC IRBapproved protocol (RAC# 2080006) with informed written consent. Blood was collected in EDTA tubes for DNA extraction. Autozygosity mapping Determination of runs of homozygosity as surrogates of autozygosity was previously described (Alkuraya 2012). Briefly, genomewide SNP genotyping was performed on Axiom SNP Chip array followed by interrogation of the SNP genotypes for runs of homozygosity >2 Mb using AutoSNPa. Trio whole exome sequencing (WES) Exome capture was performed using the TruSeqExome Enrichment kit (Illumina, San Diego, CA) following the manufacturer’s protocol. Samples of the affected patient and his unaffected parents were prepared as an Illumina sequencing library, and in the second step, the sequencing libraries were enriched for the desired target using the IlluminaExome Enrichment protocol. The captured libraries were sequenced using IlluminaHiSeq 2000 Sequencer. The reads were mapped against UCSC hg19 (http://genome.
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ucsc.edu/) by BWA (http://bio-bwa.sourceforge.net/). The SNPs and Indels were detected by SAMTOOLS (http:// samtools.sourceforge.net/). We initially searched for de novo variants. When excluded, we considered homozygous coding/splicing variants within the autozygome, with a 1000 genomes allele frequency 97th ‰); height could not be obtained at this age, however all previous height measurements were within a range
+ +
Facial dysmorphism
High arch palate
+
Glaucoma
+ +
Macrocephaly
GDD
–
Megalencephaly (Macrocephaly + Generalized brain overgrowth)
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Systemic HTN
– – –
StenosingTendonitis
Short limbs/ Limb Deformities
Syndactyly
Telangiectases
CMTC (Livedo Reticularis)
Integumentary +
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Body or limb asymmetry
+
+
Hydronephrosis
Skeletal
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Kidney disease
Hypospadias +
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Pulmonary HTN
GU
+ (Regurgitation)
Tricuspid anomaly
Cardiac defect +
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Brain Abnormalities
CVS
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+
Strokes/TIA
Brain vascular anomaly
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Seizures
CNS
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Scleral discoloration
Eyes
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Teeth malocclusion
Mouth
Our case
System
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+ (Multicystic kidney disease)
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–
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+ (Atresia)
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+
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Ben Amitai et al. (2000)# Cohort of n = 85
Table 1 Summary of the clinical manifestations seen in our patient
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Case 1
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Case 3
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+ (Asymmetry of – hemispheres)
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Case 2
Torrelo et al. (2003, 2006)
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+
–
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–
–
–
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–
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+ (Regurgitation)
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–
+
–
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+
+
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–
–
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Hinek et al. (2008)
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+
+
–
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+
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+ (Renal hypoplasia)
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Kienast and Hoeger Van Schaik (2009) Cohort of n = 27 et al. (2015)
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Mirzaa et al. (2012)
Hum Genet (2015) 134:815–822 819
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(+) Observed; (–) not observed
Our case was compared to recent publications with comparable number of cases and clinically relevant reports
+ – – – Congenital hypothyroidism
+
–
– – – Nail abnormalities
Endocrine
# This study reported the clinical findings in 85 patients with CMTC compared to five large series of 89 patients reported prior their study. Any clinical finding observed in this cohort even if it was seen once is indicated as (+)
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+ – – – –
–
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Dot-like white lesions (hypopigemnted + vs. nevus anemicus)
– Mongolian spots
+
+ +
Hyperpigmentation
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+
–
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–
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–
– – – –
–
–
–
+
+ +
– – –
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+ + Nevous Falmmeus
–
Case 3 Case 1
Case 2
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Kienast and Hoeger Van Schaik (2009) Cohort of n = 27 et al. (2015) Hinek et al. (2008) Torrelo et al. (2003, 2006)
Ben Amitai et al. (2000)# Cohort of n = 85 Our case System
Table 1 continued
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Hum Genet (2015) 134:815–822 Mirzaa et al. (2012)
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of 25–50 ‰. Facial dysmorphic features included an asymmetrical face, frontal bossing and a midline groove of the forehead, depressed nasal bridge and bulbous tip of the nose, downslanting palpebral fissures and soft bulging that appeared in the infra-orbital area over the zygoma bilaterally. He had a wide mouth, high arched palate and malocclusion of teeth. There was gray discoloration of the sclera bilaterally (Fig. 5). His skin exam showed cutis marmorata over his whole body with reticular patchy telangiectasia involving the nose and cheeks bilaterally. There was a hyperpigmented lesion on the neck as described earlier as well as white punctate lesions on his abdomen which may represent either hypopigmentation or nevus anemicus (Fig. 4). Multiple Mongolian spots were observed as noted earlier. He did not have hemihypertrophy or body asymmetry clinically (Fig. 4). There was no evidence of aplasia cutis congenita. A normal karyotype (46,XY) and molecular karyotyping were also normal. Urine organic acids and tandem MS were unremarkable. His skeletal survey was normal. Abdominal ultrasound at age of 3 years and 2 months showed hepatomegaly with a liver span of 14 cm and mildly congested hepatic veins. There was no evidence of splenomegaly. There was bilateral grade I hydronephrosis but normal kidneys. Voiding cystourethrogram was normal. Sanger sequence analysis on extracted DNA from blood of AKT3, PIK3R2 and PIK3CA genes did not reveal any mutation. Identification of a novel loss of function variant in ARL6IP6 Given the sporadic occurrence of these anomalies in our patient, we considered the possibility of a candidate de novo variant from trio WES analysis but none was identified. The parental consanguinity raised the alternative possibility of an autosomal recessive cause. Under that assumption, two novel variants involving HDDC2 and ARL6IP6 remained after applying the filters described above. The HDDC2 variant (NM_016063;exon5: c.517G>C; p.G173R) was deemed unlikely to cause the patient’s phenotype as it appeared to be tribe-specific polymorphism with a carrier frequency of 3.3 % (3 carriers out of 90 individuals from the same tribe). The remaining ARL6IP6 variant (NM_152522; exon1:c.192G>A; p.W64X), on the other hand, is a nonsense mutation (stopgain) appeared to be truly novel as it was absent in public databases (1000 Genome, ExAC server, Exome Variant Server, ClinVar, and dbVar) as well as our in-house database of nearly 600 ethnically matched exomes. Sanger sequencing confirmed homozygosity for this mutation in the patient, heterozygosity in the parents and in the unaffected siblings.
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Fig. 5 The dysmorphic facial features in our patient in addition to the facial telangiectasia (a–c); the patient has high arched palate as shown in (d)
Discussion CMTC is a vascular anomaly that can be seen in several conditions such as Down syndrome, de Lange syndrome, homocystinuria, Divry–Van Bogaert syndrome, Adams– Oliver syndrome, and Klippel–Trénaunay syndrome (Kienast and Hoeger 2009). Our patient did not have clinical features suggestive for any of these conditions. It can be argued that his dermatological findings overlap with phacomatosis pigmentovascularis type V proposed by Torrelo and colleagues but his overall clinical picture is clearly distinct from those patients (Table 1) (Torrelo et al. 2003, 2006). The neurological involvement in CMTC has been studied extensively but usually in the context of macrocephaly-capillary malformation syndrome (MCAP), which our patient clearly does not have. Of particular interest are the cerebral vascular involvement and the various resulting clinical manifestations, which have seldom been reported in patients with CMTC outside the context of MCAP syndrome. One report described stroke-like episodes in a patient with CMTC and stenosis of the left internal carotid described by Van Schaik and colleagues (Van Schaik et al. 2015). Additionally, their patient had stenosis of the left renal artery and hypertension. Stenosis of the deep femoral artery and right common iliac artery has also been reported previously in other cases with CMTC (Van Schaik et al. 2015). These reports indicate that patients with CMTC may have deep blood vessel abnormalities in addition to their readily recognizable cutaneous vascular anomalies. MRA and the post-contrast images of brain in our patient revealed increased number of right MCA branches suggesting collaterals with irregular terminal branches as well as right temporal and parietal lobes enhancement that were consistent with pial angiomatosis (Fig. 5). No obvious stenosis was visualized. Clearly, the gene mutated in CMTC
must be playing an important role in the development of the vascular system, likely both large and small blood vessels. ARL6IP6 encodes ADP-ribosylation-like factor 6 interacting protein 6. The function of ARL6IP6 is currently unknown. The only known medical relevance for this gene comes from a study that showed an intragenic SNP (rs1986743) in ARL6IP6 to have reached genomewide significance in the search for genetic susceptibility factors for young patients with ischemic stroke (Cheng et al. 2011). This is particularly interesting owing to the transient ischemic attacks in our patient with a homozygous truncating mutation in ARL6IP6. There are several examples of genes that span the phenotypic spectrum of common disease and rare severe diseases depending on the mutation class as reviewed recently (Alkuraya 2015). Therefore, if its candidacy is confirmed and replicated both in the context of CMTC and stroke susceptibility, ARL6IP6 will be an important addition to the list of genes, which will further enhance our understanding of the continuum between Mendelian and common complex diseases. Acknowledgments We thank the family for their enthusiastic participation. We also thank the Genotyping and Sequencing Core Facilities at KFSHRC for their technical help. This study was supported by KACST Grant 13-BIO1113-20 (FSA) and King Salman Center for Disability Research grant (FSA). Conflict of interest Authors declare no conflict of interest.
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