Temtamy preaxial brachydactyly syndrome (TPBS; OMIM: 605282) is an autosomal-recessive multiple congenital anomaly syndrome. It is mainly diagnosed by the presence of bilateral, symmetric preaxial brachydactyly and hyperphalangism of digits, in addition to characteristic facial features. Other associated manifestations were reported in the syndrome in the form of delayed motor and mental development, dental and ocular anomalies, sensorineural hearing loss, skeletal deformities, and decreased bone mineral density (Temtamy et al., 1998, 2012; Temtamy and Aglan, 2008).
Loss of human CHSY1 function was found to cause autosomal-recessive TPBS in 11 patients worldwide (Li et al., 2010; Tian et al., 2010). Five additional Egyptian TPBS patients associated with CHYS1 mutations were subsequently reported by Temtamy et al. (2012) with a review of the literature and definition of the phenotypic spectrum of the syndrome.
This case report adds a new Egyptian patient with TPBS with a new mutation in the CHSY1 gene.
A 3-year-old female Egyptian patient was presented to us at the Limb Malformations and Skeletal Dysplasia Clinic (LMSDC), National Research Centre (NRC). She is the second child of apparently normal first-cousin parents. The pregnancy and delivery histories were irrelevant.
On examination the patient was found to be cooperative with normal motor development and delayed speech. Anthropometric measurements were below normal for height (−3.8 SD), weight (−3.8 SD), and head circumference (−3.1 SD). She had a broad forehead, a round flat face with malar hypoplasia, wide palpebral fissures, faint blue sclera, microstomia, and micrognathia (Fig. 1a). Orodental manifestations included microdontia, diastema, high arched palate, and prominent median palatine raphe. Limb examination revealed hyperextensibility of all joints, low inserted short thumbs, wide space between the thumb and index finger, medial deviation of all fingers, soft-tissue syndactyly between digits 2 and 4, simian creases, and camptodactyly of the fourth and fifth digits bilaterally. The feet showed medial deviation of all toes with short broad big toes and soft-tissue syndactyly between toes 2 and 4 (Fig. 1b and c). Abdominal distension was noted and mild hepatomegaly was confirmed by abdominal ultrasound; needle biopsy revealed fatty infiltration. Muscle tone, deep reflexes, and external genitalia were normal. Radiological examination of hands revealed bilateral markedly short hypoplastic first metacarpal bone, short metacarpal bones of the index and middle fingers, hyperphalangism of the index finger, trapezoid shape of the proximal phalanx of the middle fingers, and hypoplasia of the middle phalanges of the fourth and fifth fingers. Radiographs of the feet showed fusion of tarsal bones, short broad metatarsal of the first toes, hyperphalangism of the big toes, hypoplastic terminal phalanges, and camptodactyly of all toes (Fig. 1d and e). Echocardiography was normal. Eye evaluation and fundus examination revealed no abnormalities, and hearing was normal. Bone densitometry showed borderline osteopenia at both the left femur and spine (z-score −0.9 and −0.86, respectively).
Mutation analysis of CHSY1
Genomic DNA was extracted from peripheral blood lymphocytes of the patient and her parents after obtaining signed informed consent aligned with the guidelines of the Research Ethical Committee of the NRC. The CHSY1 gene was amplified using five pairs of primers. The primers were designed using PRIMER 3 INPUT SOFTWARE version 0.4.0 (Table 1). The coding regions and their exon/intron boundaries of ∼50 bp sequence were investigated to identify any splice site variation. Our standard PCR cycling conditions were: initial denaturation at 96°C for 5 min; 30 cycles of denaturation at 96°C for 30 s; annealing at 60°C for 30 s; extension at 72°C for 30 min; and an additional extension at 72°C for 5 min. The PCR products were purified using the QIAquick PCR purification kit (Qiagen, Germany) and directly sequenced in both directions using the Big Dye Termination kit (Applied Biosystems, Foster City, California, USA) and analyzed on the ABI Prism 310 Genetic Analyzer (Applied Biosystems) according to the manufacturer’s instructions. A novel homozygous nonsense mutation in exon 2 of the CHSY1 gene, c.613G>T (p.E205X), was found in our patient and both parents were heterozygous (Fig. 2a). Further, the mutation was not detected in 200 normal chromosomes of Egyptian origin by PCR and restriction fragment length polymorphism analysis (PCR-RFLP) using MboII restriction endonuclease (Fig. 2c).
The proband had the characteristic limb anomalies described in all reported cases with TPBS in the form of preaxial brachydactyly, hyperphalangism, camptodactyly, clinodactyly, and tarsal fusion. These were described by Temtamy et al. (2012) as universal findings in the syndrome. Although no motor delay was noted in our patient, delayed speech was recorded. Delayed motor development was reported in 38% of previously described cases, whereas learning disabilities and mental subnormality were present in 63% (Temtamy et al., 2012).
Short stature, rounded face, microcephaly, wide-eye look, microstomia, micrognathia, and osteopenia were additional findings in our patient. These are common findings, present in more than 70% of cases as reported by Temtamy et al. (2012). Our patient is the offspring of consanguineous parents, confirming the autosomal-recessive inheritance of the syndrome (Temtamy et al., 1998; Race et al., 2010).
Molecular studies of the CHSY1 gene in our patient revealed a novel homozygous nonsense mutation in exon 2 leading to a premature stop codon at amino acid E205. This mutation affects a highly conserved glutamic acid residue (Fig. 2b) across a wide range of species and was not found in 200 normal chromosomes, thus excluding the possibility of being a polymorphism. Further, the E205X mutation is predicted to result in a truncated protein consisting of 205 amino acids instead of the whole CHSY1 802 amino-acid residues (Kitagawa et al., 2001) unless the mutated transcript is degraded through the nonsense-mediated decay pathway, which would result in a protein-null allele. To the best of our knowledge this is the second nonsense mutation to be reported in the CHSY1 gene, as Li et al. (2010) reported a Turkish family in which three affected members were homozygous for a nonsense mutation in exon 1, c.205C>T, Q69X. We have previously described seven Egyptian patients from five unrelated families with TPBS carrying five distinct mutations in the CHYS1 gene; two frameshift mutations in exon 1, and three missense mutations in exons 2 and 3 (Li et al., 2010; Temtamy et al., 2012). It is worth noting that none of the seven Egyptian patients in addition to the new case reported here shared a similar mutation, suggesting a high genetic homogeneity with strong mutation rate for this gene in Egypt.
All reported patients with TPBS were offspring of consanguineous marriages and carried homozygous mutations except for two sibs who were compound heterozygous and presented with a milder phenotype. Our finding extends the number of CHSY1 mutations to 11 different mutations. The 11 mutations are three frameshift, five missense, one splice site, and two nonsense (Table 2). There was no reported difference in the severity of the phenotype with regard to the type of mutation, as patients with either missense or other type of mutations presented with severe manifestations. However, frameshift, splice site, and nonsense mutations induce early termination and loss of CHSY1 function and are predicted to result in a more severe phenotype. Given the diversity of the identified mutations, establishing a phenotype–genotype correlation is difficult as none of the mutations identified so far was recurrent.
This case report adds a new Egyptian patient with TPBS with a novel mutation in the CHSY1 gene. Follow-up of the patient is scheduled because of the progressive nature of the disease as reported by Temtamy et al. (2010). The study confirms our previous conclusion that the syndrome is an easily recognizable dysmorphic syndrome and that the rarity of worldwide reports could be due to underdiagnosis.
We recently studied a 9-month-old female patient with the typical clinical features of TPBS. She is the first-born child to first-cousin Egyptian parents. Additional skeletal findings were developmental dysplasia of the right hip and anterolateral dislocation of the right knee joint.
On molecular study she was found to have a pathogenic mutation in exon 2 of the CHSY1 gene, c.664G>T(G222w). It is similar to the mutation that we found in family 8 of our publication (Temtamy et al., 2012).
Conflicts of interest
There are no conflicts of interest.
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Keywords:© 2014 Middle East Journal of Medical Genetics
CHSY1; Egypt; loss of function mutations; parental consanguinity; skeletal disorder; Temtamy preaxial brachydactyly syndrome