Compound heterozygous p. Arg949Trp and p. Gly970Ala mutations deteriorated the function of PEX1p: A study on PEX1 in a patient with Zellweger syndrome.

The peroxisome is responsible for a variety of vital pathways in primary metabolism, including the very long-chain fatty-acid oxidation and plasmalogen lipid biosynthesis. Autosomal recessive disorder of the Zellweger spectrum (ZSD) is a major subset of peroxisome biogenesis disorders (PBDs) that can be caused by mutations in any of the 14 PEX genes. Zellweger syndrome (ZS) is the foremost common and severe phenotype within the heterogeneous ZSD. However, missense mutations encode proteins with residual functions, which are associated with phenotypes that are milder than ZS. Mutations in the PEX1 gene are among the most prevalent. PEX1 and PEX6 proteins, belonging to the AAA family of ATPases, form a hexameric complex, which is associated with peroxisome membranes and essential for peroxisome biology. In this study, a two-month-old Iranian boy with hypotonia, poor feeding, and difficulty in breathing was diagnosed with Zellweger syndrome. The parents of the patient were second cousins and healthy and no similar cases were observed in the parents' family. The PEX1 gene was sequenced in the patient and his parents. The compound heterozygous mutations, p. Arg949Trp and p. Gly970Ala, were identified in the patient, while the parents were heterozygous for these alleles. Sequence analysis of the mutant PEX1 D2 domain revealed that mutation p. Arg949Trp precisely occurred in a conserved arginine residue (P4 Arg), which hinders the substrate processing of the complex. Several database records have reported mutation p. Arg949Trp(R949W) but its clinical significance is given as uncertain. We report here a novel mutation, p. Gly970Ala, which is not recorded before and may prevent proper interaction of PEX1 and PEX6 proteins. In summary, the clinical findings and peroxisome profile of the patient suggested that compound heterozygosity for these two missense mutations resulted in a nonfunctional PEX1/PEX6 complex causing the severe ZS phenotype.

Mild reduction of plasmalogens causes rhizomelic chondrodysplasia punctata: functional characterization of a novel mutation.

Rhizomelic chondrodysplasia punctata (RCDP) is an autosomal recessive disorder due to the deficiency in ether lipid synthesis. RCDP type 1, the most prominent type, is caused by the dysfunction of the receptor of peroxisome targeting signal type 2, Pex7 (peroxisomal biogenesis factor 7), and the rest of the patients, RCDP types 2 and 3, have defects in peroxisomal enzymes catalyzing the initial two steps of alkyl-phospholipid synthesis, glyceronephosphate O-acyltransferase and alkylglycerone phosphate synthase (Agps). We herein investigated defects of two patients with RCDP type 3. Patient 1 had a novel missense mutation, T1533G, resulting in the I511M substitution in Agps. The plasmalogen level was mildly reduced, whereas the protein level and peroxisomal localization of Agps-I511M in fibroblasts were normal as in the control fibroblasts. Structure prediction analysis suggested that the mutated residue was located in the helix α15 on the surface of V-shaped active site tunnel in Agps, likely accounting for the mild defects of plasmalogen synthesis. These results strongly suggest that an individual with mildly affected level of plasmalogen synthesis develops RCDP. In fibroblasts from patient 2, the expression of AGPS mRNA and Agps protein was severely affected, thereby giving rise to the strong reduction of plasmalogen synthesis.

Fanconi-Bickel syndrome versus osteogenesis imperfeeta: An Iranian case with a novel mutation in glucose transporter 2 gene, and review of literature.

Fanconi-Bickel syndrome is an extremely rare hereditary metabolic disease, characterized by hepatomegaly due to glycogen storage, refractory hypophosphatemic rickets, marked growth retardation and proximal renal tubular acidosis. Recurrent bone fractures are one of the hallmark findings. It is a single gene disorder; the responsible gene belongs to the facilitative glucose transporters 2 (GLUT2) family gene or (SLC2A2) mapped to the q26.1-26.3 locus on chromosome 3, and encodes the GLUT protein 2. This protein is expressed in pancreatic ί-cells, hepatocytes, renal tubules, and intestinal mucosa. Several mutations in the GLUT2 gene have been reported in different ethnicities. Herein we report an Iranian girl with a missed diagnosis of osteogenesis imperfecta. She was referred with the history of frequent fractures, and severe motor delay and was suspected to osteogenesis imperfecta. Following the case we detected refractory rickets instead of OI, sever growth failure, proximal renal tubulopathy and RTA, and enlarged kidneys, progressive hepatomegaly, and GSD on liver biopsy. Glucose and galactose tolerance tests confirmed abnormal carbohydrate metabolism. Molecular analysis on GLUT2 gene revealed a homozygous novel mutation in exon 5; it was 15 nucleotide deletion and 7 nucleotide insertion and caused a frame shift mutation, produced a premature truncated protein (P.A229QFsX19). This mutation has not been reported before in the relevant literature.

Disease-associated mutations in the actin-binding domain of filamin B cause cytoplasmic focal accumulations correlating with disease severity.

Dominant missense mutations in FLNB, encoding the actin-cross linking protein filamin B (FLNB), cause a broad range of skeletal dysplasias with varying severity by an unknown mechanism. Here these FLNB mutations are shown to cluster in exons encoding the actin-binding domain (ABD) and filamin repeats surrounding the flexible hinge 1 region of the FLNB rod domain. Despite being positioned in domains that bind actin, it is unknown if these mutations perturb cytoskeletal structure. Expression of several full-length FLNB constructs containing ABD mutations resulted in the appearance of actin-containing cytoplasmic focal accumulations of the substituted protein to a degree that was correlated with the severity of the associated phenotypes. In contrast, study of mutations leading to substitutions in the FLNB rod domain that result in the same phenotypes as ABD mutations demonstrated that with only one exception disease-associated substitutions, surrounding hinge 1 demonstrated no tendency to form actin-filamin foci. The exception, a substitution in filamin repeat 6, lies within a region previously implicated in filamin-actin binding. These data are consistent with mutations in the ABD conferring enhanced actin-binding activity but suggest that substitutions affecting repeats near the flexible hinge region of FLNB precipitate the same phenotypes through a different mechanism.

Identification of CANT1 mutations in Desbuquois dysplasia.

Desbuquois dysplasia is a severe condition characterized by short stature, joint laxity, scoliosis, and advanced carpal ossification with a delta phalanx. Studying nine Desbuquois families, we identified seven distinct mutations in the Calcium-Activated Nucleotidase 1 gene (CANT1), which encodes a soluble UDP-preferring nucleotidase belonging to the apyrase family. Among the seven mutations, four were nonsense mutations (Del 5' UTR and exon 1, p.P245RfsX3, p.S303AfsX20, and p.W125X), and three were missense mutations (p.R300C, p.R300H, and p.P299L) responsible for the change of conserved amino acids located in the seventh nucleotidase conserved region (NRC). The arginine substitution at position 300 was identified in five out of nine families. The specific function of CANT1 is as yet unknown, but its substrates are involved in several major signaling functions, including Ca2+ release, through activation of pyrimidinergic signaling. Importantly, using RT-PCR analysis, we observed a specific expression in chondrocytes. We also found electron-dense material within distended rough endoplasmic reticulum in the fibroblasts of Desbuquois patients. Our findings demonstrate the specific involvement of a nucleotidase in the endochondral ossification process.

Prenatal diagnosis in a mentally retarded woman with mosaic ring chromosome 18.

We present a pregnant woman with mental retardation and mosaic for ring 18 referred for prenatal diagnosis. Major clinical features included short stature with clinodactyly in feet, foot deformity and club feet, hypotonia, kyphosis, and absence of breast development, low set ears, high arched palate, dental decay and speech disorder. Prenatal diagnosis was carried. Using amniocentesis. The fetus had a normal karyotype described as 46,XX. The fetus was evaluated for clinical features after delivery; she was healthy with no abnormal clinical characterizations.

Brachyphalangy, polydactyly and tibial aplasia/hypoplasia syndrome (OMIM 609945): case report and review of the literature.

Brachyphalangy, polydactyly and tibial aplasia/hypoplasia syndrome (OMIM 609945) is a rare congenital disorder. Only seven patients have been reported to date, and the etiology of this syndrome is unknown. Autosomal dominant inheritance with variable expression has been suggested based on the presence of minor features in some parents and the fact that neither parental consanguinity nor pairs of affected siblings were observed. We report on the first patient with this syndrome who was born to consanguineous parents. Neither the mother nor the father, who were first cousins, had clinical features suggestive of a manifestation of brachyphalangy, polydactyly and tibial aplasia/hypoplasia syndrome. The patient had no siblings, and the family history was unremarkable. Clinical problems included brachydactyly of hands and feet, splaying of fingers and toes, preaxial polydactyly of feet, bilateral tibial aplasia, shortened radius and ulna, and characteristic facial dysmorphic signs. The detailed description of this patient adds to our knowledge of the clinical manifestations of brachyphalangy, polydactyly and tibial aplasia/hypoplasia syndrome and will eventually also contribute to the elucidation of the underlying gene defects.

Whole-Exome Sequencing Identifies Three Candidate Homozygous Variants in a Consanguineous Iranian Family with Autism Spectrum Disorder and Skeletal Problems.

Autism spectrum disorder (ASD) is characterized by 3 core symptoms with impaired social communication, repetitive behavior, and/or restricted interests in early childhood. As a complex neurodevelopmental disorder (NDD), the phenotype and severity of autism are extremely heterogeneous. Genetic factors have a key role in the etiology of autism. In this study, we investigated an Azeri Turkish family with 2 ASD-affected individuals to identify probable ASD-causing variants. First, the affected individuals were karyotyped in order to exclude chromosomal abnormalities. Then, whole-exome sequencing was carried out in one affected sibling followed by cosegregation analysis for the candidate variants in the family. In addition, SNP genotyping was carried out in the patients to identify possible homozygosity regions. Both proband and sibling had a normal karyotype. We detected 3 possible causative variants in this family: c.5443G>A; p.Gly1815Ser, c.1027C>T; p.Arg343Trp, and c.382A>G; p.Lys128Glu, which are in the FBN1, TF, and PLOD2 genes, respectively. All of the variants cosegregated in the family, and SNP genotyping revealed that these 3 variants are located in the homozygosity regions. This family serves as an example of a multimodal polygenic risk for a complex developmental disorder. Of these 3 genes, confluence of the variants in FBN1 and PLOD2 may contribute to the autistic features of the patient in addition to skeletal problems. Our study highlights the genetic complexity and heterogeneity of NDDs such as autism. In other words, in some patients with ASD, multiple rare variants in different loci rather than a monogenic state may contribute to the development of phenotypes.

Phenotypic Overlap of Roberts and Baller-Gerold Syndromes in Two Patients With Craniosynostosis, Limb Reductions, and ESCO2 Mutations.

Baller-Gerold (BGS, MIM#218600) and Roberts (RBS, MIM#268300) syndromes are rare autosomal recessive disorders caused, respectively, by biallelic alterations in RECQL4 (MIM*603780) and ESCO2 (MIM*609353) genes. Common features are severe growth retardation, limbs shortening and craniofacial abnormalities which may include craniosynostosis. We aimed at unveiling the genetic lesions underpinning the phenotype of two unrelated children with a presumptive BGS diagnosis: patient 1 is a Turkish girl with short stature, microcephaly, craniosynostosis, seizures, intellectual disability, midface hemangioma, bilateral radial and thumb aplasia, tibial hypoplasia, and pes equinovarus. Patient 2 is an Iranian girl born to consanguineous parents with craniosynostosis, micrognathism, bilateral radial aplasia, thumbs, and foot deformity in the context of developmental delay. Upon negative RECQL4 test, whole exome sequencing (WES) analysis performed on the two trios led to the identification of two different ESCO2 homozygous inactivating variants: a previously described c.1131+1G>A transition in patient 1 and an unreported deletion, c.417del, in patient 2, thus turning the diagnosis into Roberts syndrome. The occurrence of a Baller-Gerold phenotype in two unrelated patients that were ultimately diagnosed with RBS demonstrates the strength of WES in redefining the nosological landscape of rare congenital malformation syndromes, a premise to yield optimized patients management and family counseling.

Investigation of Chromosomal Abnormalities and Microdeletion/ Microduplication(s) in Fifty Iranian Patients with Multiple Congenital Anomalies.

OBJECTIVE: Major birth defects are inborn structural or functional anomalies with long-term disability and adverse impacts on individuals, families, health-care systems, and societies. Approximately 20% of birth defects are due to chromosomal and genetic conditions. Inspired by the fact that neonatal deaths are caused by birth defects in about 20 and 10% of cases in Iran and worldwide respectively, we conducted the present study to unravel the role of chromosome abnormalities, including microdeletion/microduplication(s), in multiple congenital abnormalities in a number of Iranian patients. MATERIALS AND METHODS: In this descriptive cross-sectional study, 50 sporadic patients with Multiple Congenital Anomalies (MCA) were selected. The techniques employed included conventional karyotyping, fluorescence in situ hybridization (FISH), multiplex ligation-dependent probe amplification (MLPA), and array comparative genomic hybridisation (array-CGH), according to the clinical diagnosis for each patient. RESULTS: Chromosomal abnormalities and microdeletion/microduplication(s) were observed in eight out of fifty patients (16%). The abnormalities proved to result from the imbalances in chromosomes 1, 3, 12, and 18 in four of the patients. However, the other four patients were diagnosed to suffer from the known microdeletions of 22q11.21, 16p13.3, 5q35.3, and 7q11.23. CONCLUSION: In the present study, we report a patient with 46,XY, der(18)[12]/46,XY, der(18), +mar[8] dn presented with MCA associated with hypogammaglobulinemia. Given the patient's seemingly rare and highly complex chromosomal abnormality and the lack of any concise mechanism presented in the literature to justify the case, we hereby propose a novel mechanism for the formation of both derivative and ring chromosome 18. In addition, we introduce a new 12q abnormality and a novel association of an Xp22.33 duplication with 1q43q44 deletion syndrome. The phenotype analysis of the patients with chromosome abnormality would be beneficial for further phenotype-genotype correlation studies.

A molecular and clinical study of Larsen syndrome caused by mutations in FLNB.

BACKGROUND: Larsen syndrome is an autosomal dominant osteochondrodysplasia characterised by large-joint dislocations and craniofacial anomalies. Recently, Larsen syndrome was shown to be caused by missense mutations or small inframe deletions in FLNB, encoding the cytoskeletal protein filamin B. To further delineate the molecular causes of Larsen syndrome, 20 probands with Larsen syndrome together with their affected relatives were evaluated for mutations in FLNB and their phenotypes studied. METHODS: Probands were screened for mutations in FLNB using a combination of denaturing high-performance liquid chromatography, direct sequencing and restriction endonuclease digestion. Clinical and radiographical features of the patients were evaluated. RESULTS AND DISCUSSION: The clinical signs most frequently associated with a FLNB mutation are the presence of supernumerary carpal and tarsal bones and short, broad, spatulate distal phalanges, particularly of the thumb. All individuals with Larsen syndrome-associated FLNB mutations are heterozygous for either missense or small inframe deletions. Three mutations are recurrent, with one mutation, 5071G-->A, observed in 6 of 20 subjects. The distribution of mutations within the FLNB gene is non-random, with clusters of mutations leading to substitutions in the actin-binding domain and filamin repeats 13-17 being the most common cause of Larsen syndrome. These findings collectively define autosomal dominant Larsen syndrome and demonstrate clustering of causative mutations in FLNB.

Vitamin D-dependent rickets type II: report of a novel mutation in the vitamin D receptor gene.

Hereditary vitamin D-resistant rickets type or vitamin D-dependent rickets type II is a genetically determined and rare autosomal recessive disorder, most often caused by mutations in the vitamin D receptor gene. It usually presents with rachitic changes not responsive to vitamin D treatment and the circulating levels of 1,25 (OH)2 vitamin D-3 are elevated, differentiating it from vitamin D-dependent rickets type I. Alopecia capitis or alopecia totalis is seen in some families with vitamin D-dependent rickets type II. This is usually associated with a more severe phenotype. In this report, we present the clinical findings on a family which exhibited the typical clinical features of hereditary vitamin D-resistant rickets in two siblings. In addition, molecular analysis of the vitamin D receptor gene was performed by sequencing all coding exons. The cardinal findings in the index patient were alopecia totalis, renal tubular acidosis, mild generalized aminoaciduria, refractory rickets, high alkaline phosphatase, and hyperparathyroidism. Other routine biochemical tests were within normal limits, but 1+ glycine was detected in his urine. Skin biopsy results were compatible with alopecia areata. A previous child with similar phenotype was reported to be deceased at the age of 32 months. Mutation analysis of the vitamin D receptor gene by direct sequencing analysis of all coding exons showed a homozygous c.122GA(p.Cys41Tyr) variant in exon 2 with several arguments pointing to a pathogenic effect. We should be aware of this very rare disease whenever we see a patient with refractory rickets and alopecia.

Molecular analysis of the neuronal apoptosis inhibitory protein gene in families with spinal muscular atrophy.

BACKGROUND: Spinal muscular atrophy is an autosomal recessive disorder characterized by degeneration of anterior horn cells in the spinal cord leading to progressive muscular weakness and atrophy. The spinal muscular atrophy candidate interval genes including survival motor neuron, the responsible gene in spinal muscular atrophy phenotype expression, neuronal apoptosis inhibitory protein, and P44, potential modifying genes, are located on chromosome 5q13 in two highly homologous copies (telomeric and centromeric) within the spinal muscular atrophy region. METHODS: In this study, the neuronal apoptosis inhibitory protein gene deletion was analyzed in 34 spinal muscular atrophy families, with the consanguinity rate of 65% (22/34), in whom exon 7 of the survival motor neuron-1 gene was already confirmed and was deleted in 79% of the affected individuals. Deletion analysis of exons 5, 6, and 13 of the neuronal apoptosis inhibitory protein-t gene was carried out in our samples. RESULTS: We found 80% neuronal apoptosis inhibitory protein gene deletion in 5q-spinal muscular atrophy patients (91% spinal muscular atrophy-I, 50% spinal muscular atrophy-II and -III), and in 5% (two of forty) of spinal muscular atrophy parents. All the neuronal apoptosis inhibitory protein-deleted samples also lacked the survival motor neuron-1 gene. CONCLUSION: The neuronal apoptosis inhibitory protein gene deletion in spinal muscular atrophy-I was higher than the other spinal muscular atrophy types. The high frequency of neuronal apoptosis inhibitory protein deletion most likely reflects a higher frequency of survival motor neuron-1 deletions compared with survival motor neuron-1 to survival motor neuron-2 gene conversion in this population.

L-2-hydroxyglutaric aciduria: a report of six cases and review of the literature.

L-2-hydroxyglutaric aciduria is a rare and novel autosomal recessive inherited neurometabolic disorder. Since its first description by Duran in 1980, less than 100 cases have so far been reported. Occurring mostly in childhood, it is characterized by slowly progressive neurological dysfunction with cerebellar ataxia, pyramidal signs, intellectual decline, seizure, and extrapyramidal symptoms. MRI scanning is highly characteristic and screening for organic acid (L-2-hydroxyglutaric acid) in urine, serum, and cerebrospinal fluid is diagnostic. We investigated six Iranian children, aged 4, 14, 16, and 16 years, (the last one had two affected brothers and both of them died of similar illness at the ages of 20 and 22), by urinary organic acids assay and MRI scanning with suspicion of this rare disorder. Symptoms were suspicious for one of the leukoencephalopathies accompanied by macrocephaly. Affected cases were evaluated because of mild to moderate psychomotor retardation and regression. Head circumferences were above 2 standard deviations. Urine levels of L-2-hydroxyglutaric acid were strongly increased. MRI scanning of the brain showed hyperintense signal on T2W images of the subcortical white matter and basal ganglia in all of them. Because of its inheritance pattern (autosomal recessive) and the high rate of consanguineous marriages in Iran, the prevalence of this disorder might be high among the mentally-handicapped patients, especially those with macrocephaly. Therefore, this entity should be considered in the differential diagnosis of mentally-retarded patients with macrocephaly.

LMNA mutations in atypical Werner's syndrome.

BACKGROUND: Werner's syndrome is a progeroid syndrome caused by mutations at the WRN helicase locus. Some features of this disorder are also present in laminopathies caused by mutant LMNA encoding nuclear lamin A/C. Because of this similarity, we sequenced LMNA in individuals with atypical Werner's syndrome (wild-type WRN). METHODS: Of 129 index patients referred to our international registry for molecular diagnosis of Werner's syndrome, 26 (20%) had wildtype WRN coding regions and were categorised as having atypical Werner's syndrome on the basis of molecular criteria. We sequenced all exons of LMNA in these individuals. Mutations were confirmed at the mRNA level by RT-PCR sequencing. In one patient in whom an LMNA mutation was detected and fibroblasts were available, we established nuclear morphology and subnuclear localisation. FINDINGS: In four (15%) of 26 patients with atypical Werner's syndrome, we noted heterozygosity for novel missense mutations in LMNA, specifically A57P, R133L (in two people), and L140R. The mutations altered relatively conserved residues within lamin A/C. Fibroblasts from the patient with the L140R mutation had a substantially enhanced proportion of nuclei with altered morphology and mislocalised lamins. Individuals with atypical Werner's syndrome with mutations in LMNA had a more severe phenotype than did those with the disorder due to mutant WRN. INTERPRETATION: Our findings indicate that Werner's syndrome is molecularly heterogeneous, and a subset of the disorder can be judged a laminopathy.

Hereditary bilateral conductive hearing loss caused by total loss of ossicles: a report of familial expansile osteolysis.

OBJECTIVE: The objective of this study was to report on three members of a family with familial expansile osteolysis; the important point about these patients was that none of them had middle-ear ossicles. STUDY DESIGN AND SUBJECTS: A retrospective case review including three cases with familial expansile osteolysis. SETTING: Department of Otolaryngology in a tertiary referral center. INTERVENTIONS: Each patient underwent computerized tomography of the temporal bone in the coronal view, audiometric and tympanometric evaluations, biochemical investigation, whole body isotope scans by Tc-99 mMDP and X-ray. Also the patients' pedigree was studied. Two of the patients had exploratory middle-ear surgery as well. RESULTS: The temporal-bone computed-tomography scan in the coronal view of all three patients and also exploratory middle-ear surgery, which was done on two of the patients, showed no ossicles in the middle ear of either ear in all three cases. This feature hadn't been reported in previous studies. Hearing loss was revealed in the medical histories since childhood. Audiometry indicated mild to moderate conductive and mixed hearing loss and also an AD-type tympanogram pattern along with an absence of acoustic reflexes in both ears of the cases. Both serum alkaline phosphatase and hydroxyproline levels were elevated. There was an increase in uptake and activity at multiple foci of the whole skeleton. No improvement in hearing thresholds was obtained after reconstruction of the middle ear. CONCLUSION: The total absence of middle-ear ossicles can probably be regarded as a new symptom in some patients with familial expansile osteolysis. Common ossiculoplasty for improving the hearing thresholds in this condition may be unsuccessful; therefore, both surgeons and patients must be completely aware of the contingent undesirable results.

Yunis-Varón syndrome: the first report of two Iranian cases.

The Yunis-Varón syndrome represents a rare autosomal recessive syndrome of easy recognition characterized by defective growth of the cranial bone along with complete or partial absence of the clavicles (cleidocranial dysplasia), absence of thumbs and halluces, distal aphalangia, ectodermal anomalies, growth retardation and poor outcome. The molecular genetic basis is unknown. Here, we report an 8 months old girl with Yunis-Varón syndrome, born to a consanguineously married, with normal parents. She had micrognathia, wide fontanels, prominent eyes, poor sucking, congenital heart diseases, asymmetric face, ambiguous genitalia, reduction anomaly in right hand including thumb, and hypoplastic distal phalanges of 3th fingers, and hypo plastic clavicles. She has glaucoma and lenses opacity. There is another similar case in her family. Karyotype is normal. She is the first Iranian known case of Yunis-Varón syndrome.

Fraccaro syndrome: report of two Iranian cases: an infant and an adult in a family.

49,XXXXY is rare chromosomal pattern and these patients have mental retardation, small penis, cryptorchidism and skeletal anomalies. We reported a 10 month-old boy who has hypotonia, microcephaly, hypertelorism, depressed nasal bridge, epicanthic folds and bilateral multiple ear tags, high arched palate, down set ears, micrognathia and congenital heart disease such as patent ductus arteriosus (PDA), Atrial septal defect (ASD), mild pulmonary stenosis. Among the skeletal anomalies, he has kyphoscoliosis, clinodactyly of the fourth and fifth fingers of both hands, and bilateral club foot and unilateral dysplasia of the hip. Karyotype was found as 49,XXXXY[44]/48,XXXY[6] and this cytogenetic analysis was help to establish clinical diagnosis Fraccaro syndrome.