Unrelated Iranian Families Research Articles

Genetic screening in two Iranian families with early-onset Alzheimer's disease identified a novel PSEN1 mutation

A subset of early-onset Alzheimer's disease is inherited as an autosomal-dominant trait and is associated with mutations in the genes encoding β-amyloid precursor protein, presenilin 1, or presenilin 2. In this study, we identified 2 PSEN1 mutations (1 novel and 1 known) in 2 unrelated Iranian families with autosomal-dominant Alzheimer's disease. The disease progressed rapidly with a mean age at onset of 33 and 42 years and an age at death ranging from 43 to 48 years.

Mutations of the aminoacyl-tRNA-synthetases SARS and WARS2 are implicated in the etiology of autosomal recessive intellectual disability.

Intellectual disability (ID) is the hallmark of an extremely heterogeneous group of disorders that comprises a wide variety of syndromic and non-syndromic phenotypes. Here, we report on mutations in two aminoacyl-tRNA synthetases that are associated with ID in two unrelated Iranian families. In the first family, we identified a homozygous missense mutation (c.514G>A, p.Asp172Asn) in the cytoplasmic seryl-tRNA synthetase (SARS) gene. The mutation affects the enzymatic core domain of the protein and impairs its enzymatic activity, probably leading to reduced cytoplasmic tRNASer concentrations. The mutant protein was predicted to be unstable, which could be substantiated by investigating ectopic mutant SARS in transfected HEK293T cells. In the second family, we found a compound heterozygous genotype of the mitochondrial tryptophanyl-tRNA synthetase (WARS2) gene, comprising a nonsense mutation (c.325delA, p.Ser109Alafs*15), which very likely entails nonsense-mediated mRNA decay and a missense mutation (c.37T>G, p.Trp13Gly). The latter affects the mitochondrial localization signal of WARS2, causing protein mislocalization. Including AIMP1, which we have recently implicated in the etiology of ID, three genes with a role in tRNA-aminoacylation are now associated with this condition. We therefore suggest that the functional integrity of tRNAs in general is an important factor in the development and maintenance of human cognitive functions.

Molecular characterization of AIPL1 gene region in the Iranian population: application of novel informative haplotypes and detection of mutational founder effect

Leber’s congenital amaurosis (LCA) is considered as one of the main causes of congenital blindness. In view of the genetically heterogeneous nature of the disease, indirect diagnosis using linkage analysis has proven to be useful in molecular diagnosis procedure. Mutations in AIPL1 gene are one of the leading causes of LCA. In the present study, the application of three single nucleotide polymorphic (SNP) markers related to the gene, including rs7212734, rs11658369 and rs8066853 was evaluated for the first time in the Iranian population. The markers were genotyped using tetra-primer ARMS PCR in 154 unrelated healthy individuals. Haplotype frequency and other characteristics of the markers were examined by using the GENEPOP, PowerMarker and Cubic Exact Solution software. The data indicated the presence of six different haplotypes in the Iranian population. Among them, three haplotypes showed high informativeness with frequencies ≥0.05. Three unrelated Iranian LCA families were analyzed. The p.W278* mutation in exon 6 of AIPL1 was found in all the families using APEX microarray chips. SNP analysis for the families showed the conservation of T−A−A haplotype linked to p.W278* mutation. All families bearing the mutation were from the Isfahan province and a founder effect was suggested in this population. Prenatal diagnosis using the markers led to the successful prediction of the fetus genotypes in all the at risk pregnancies and showed that rs7212734, rs11658369 and rs8066853 can be considered as the three informative markers for linkage analysis in carrier detection and molecular diagnosis of LCA in the Iranian population.

Mutational Screening of PKD1 and PKD2 Genes in Iranian Population Diagnosed with Autosomal Dominant Polycystic Kidney Disease.

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is the fourth most frequent cause of endstage renal disease (ESDR), occurring at a varying frequency of 1/400 to 1/800 persons. The disease affects all ethnic groups worldwide, and there is a need for population based studies to be carried out for better diagnostic, genetic counselling, and treatment purpose. Eighteen unrelated probands (10 males and 8 females) with a familial history of ADPKD were selected for the study. Their clinical evaluation was performed to diagnose and assess disease progression. PKD1 and PKD2 genes were genotyped in each proband by next generation sequencing (NGS). Mutational analysis of PKD1 and PKD2 genes using NGS in eighteen unrelated Iranian ADPKD families revealed a total of eighteen heterozygous variations. PKD1 genotype revealed eight frameshift deletion mutations, two frameshift insertion mutations, five nonsense mutations and one splice mutation and PKD2 showed one frameshift deletion mutation and one frameshift insertion mutation. Four of the variants reported were novel and were present in the PKD1 gene. Further, PKD1 truncating mutations reached ESRD earlier than patients with non-truncating PKD1 mutations (52 ± 3.2 years vs. 58 ± 10.8 years, p = 0.01). The PKD1 and PKD2 genotyping of ADPKD Iranian patients with familial history showed no mutational hotspot. The screening has given four novel variants that will contribute to diagnosis, genetic counselling, and treatment of ADPKD patients in general.

Linkage Study Revealed Complex Haplotypes in a Multifamily due to Different Mutations in CAPN3 Gene in an Iranian Ethnic Group.

Calpainopathy is an autosomal recessive form of limb girdle muscular dystrophies which is caused by mutation in CAPN3 gene. In the present study, co-segregation of this disorder was analyzed with four short tandem repeat markers linked to the CAPN3 gene. Three apparently unrelated Iranian families with same ethnicity were investigated. Haplotype analysis and sequencing of the CAPN3 gene were performed. DNA sample from one of the patients was simultaneously sent for next-generation sequencing. DNA sequencing identified two mutations. It was seen as a homozygous c.2105C>T in exon 19 in one family, a homozygous novel mutation c.380G>A in exon 3 in another family, and a compound heterozygote form of these two mutations in the third family. Next-generation sequencing also confirmed our results. It was expected that, due to the rare nature of limb girdle muscular dystrophies, affected individuals from the same ethnic group share similar mutations. Haplotype analysis showed two different homozygote patterns in two families, yet a compound heterozygote pattern in the third family as seen in the mutation analysis. This study shows that haplotype analysis would help in determining presence of different founders.

Report of limb girdle muscular dystrophy type 2a in 6 Iranian patients, one with a novel deletion in CAPN3 gene

Calpain3 is a calcium-dependent intracellular protease involved in an autosomal recessive form of muscular dystrophy known as limb-girdle muscular dystrophy type 2A. Many pathogenic mutations have been identified in calpain3, encoded by the CAPN3 gene, which leads to weakness of the pelvic and shoulder girdle muscles. In the present study, whole exome sequencing was performed on six unrelated Iranian families who presented with progressive muscle weakness, with a strong suspicion of Calpainopathies. Genetic analysis of CAPN3 gene revealed five causative variants which had not been reported in the Iranian population before including a novel 6 bp deletion (c.795_800delCATTGA) and four previously reported mutations (c.1939G > T, c.2243G > A, c.2257delGinsAA, and c.2380 + 2T > G). Our findings indicate that exome sequencing can be a very effective and affordable method to diagnose heterogeneous muscular dystrophies, especially in consanguineous populations such as Iran.

Molecular and clinical characterization of Waardenburg syndrome type I in an Iranian cohort with two novel PAX3 mutations

Waardenburg syndrome (WS) is a disease of abnormal neural-crest derived melanocyte development characterized by hearing loss and pigmentary disturbances in hair, eyes and skin. WS is subdivided into four major types, WS1–WS4, where WS1 is recognized by the presence of dystopia canthorum, with PAX3 being the only known gene involved. This study aimed at investigating PAX3 mutations and clinical characteristics of WS1 in a group of Iranian patients.A total of 12 WS1 patients from four unrelated Iranian families were enrolled. Waardenburg consortium guidelines were used for WS1 diagnosis. A detailed family history was traced and a thorough clinical examination was performed for all participants. Furthermore, WS1 patients underwent screening for PAX3 mutations using PCR-sequencing.Dystopia canthorum, broad high nasal root and synophrys were observed in all patients. Early graying, hair discoloration, hypoplastic blue eyes (characteristic brilliant blue iris) and hearing loss were the most common features observed, while heterochromia iridis was the least frequently observed sign among the studied Iranian WS1 patients. Genetic analysis of PAX3 revealed four mutations including c.667C>T, c.784C>T, c.951delT and c.451+3A>C. Two of the four mutations reported here (c.951delT and c.451+3A>C) are being reported for the first time in this study.Our data provide insight into genotypic and phenotypic spectrum of WS1 in an Iranian series of patients. Our results expand the spectrum of PAX3 mutations and may have implications for the genetic counseling of WS in Iran.

New evidence for the role of calpain 10 in autosomal recessive intellectual disability: identification of two novel nonsense variants by exome sequencing in Iranian families.

Knowledge of the genes responsible for intellectual disability, particularly autosomal recessive forms, is rapidly expanding. Increasing numbers of the gene show great heterogeneity and supports the hypothesis that human genome may contain over 2000 causative genes with a critical role in brain development. Since 2004, we have applied genome-wide SNP genotyping and next-generation sequencing in large consanguineous Iranian families with intellectual disability, to identify the genes harboring disease-causing mutations. The current study paved the way for identification of responsible genes in two unrelated Iranian families. We found two novel nonsense mutations, p.C77* and p.Q115*, in the calpain catalytic domain of CAPN10, which is a cysteine protease known to be involved in pathogenesis of noninsulin-dependent diabetes mellitus. Another different mutation in this gene (p.S138_R139ins5) has previously been reported in an Iranian family. All of these patients have common clinical features in spite of specific brain structural abnormalities on MRI. Different mutations in CAPN10 have already been found in three independent Iranian families. These results have strongly supported the possible role of CAPN10 in human brain development. Altogether, we proposed CAPN10 as a promising candidate gene for intellectual disability, which should be considered in diagnostic gene panels.

Investigation of ATP6V1B1 and ATP6V0A4 genes causing hereditary hearing loss associated with distal renal tubular acidosis in Iranian families.

Hearing defects are the most common sensory disorders, affecting 1 out of every 500 newborns. ATP6V1B mutations are associated with early sensorineural hearing loss, whereas ATP6V0A4 mutations are classically associated with either late-onset sensorineural hearing loss or normal hearing. ATP6V1B1 and ATP6V0A4 genetic mutations cause recessive forms of distal renal tubular acidosis. Ten unrelated deaf Iranian families with distal renal tubular acidosis were referred to the Genetics Research Centre, University of Social Welfare and Rehabilitation Sciences, Tehran. All exons of the ATP6V1B1 and ATP6V0A4 genes were sequenced in affected family members. We identified a previously reported ATP6V1B1 frameshift mutation (P385fsX441) in two families and a nucleotide substitution in exon 10 (P346R) in three families. In addition, one patient was homozygous for a novel nucleotide substitution in exon 3. ATP6V1B1 genetic mutations were detected in more than half of the families studied. Mutations in this gene therefore seem to be the most common causative factors in hearing loss associated with distal renal tubular acidosis in these families.

A Novel Mutation in the α2-Globin Gene in Two Unrelated Iranian Families

Background: α-globin is encoded by two adjacent genes, αl and α2. Evidence suggests that these genes are not expressed equally and that the α2-globin gene encodes the majority of α-globin. This finding predicts that a thalassemic mutation of the α2-globin gene would result in a more severe loss of α-chain synthesis than a similar mutation in the αl-globin gene. Objectives: In the present study we described a novel non-deletion α-thalassemia defect in the 5'UTR region of the α2-globin gene. Materials and Methods: For molecular analysis, genomic DNA was isolated from peripheral blood cells by a salting out procedure. The common alpha deletion mutations were ruled out using the published primers and conditions. The amplification of the entire β and α1 globin genes was also carried out and their DNA was sequenced. No mutation was detected. Results: The mutation under study was located on an AP-1 transcription factor binding site and inherited in two unrelated Iranian families with hypochromic microcytic anemia. Conclusions: The patients in this study had moderate microcytosis and hypochromia without hemolysis, jaundice and splenomegaly. Molecular analysis in these patients revealed a non-deletion type of mutation in the promoter region, which is highly consistent with findings of other studies.

A transversion mutation in non-coding exon 3 of the TMC1 gene in two ethnically related Iranian deaf families from different geographical regions; evidence for founder effect

ObjectivesTransmembrane channel-like 1 (TMC1) gene is a member of the transmembrane channel-like (TMC) gene family that encodes an integral membrane protein of the inner ear. It is suggested that mutation in this gene is one of the main causes of autosomal recessive non-syndromic hearing loss (ARNSHL) in different populations. The aim of this study was to determine the contribution of the TMC1 gene mutations in causing hearing loss in Iran. MethodsIn total 54 unrelated Iranian families containing 159 affected individuals with ARNSHL detected by audiometric and otologic examinations were analyzed. Haplotype analysis of all members of 45 GJB2- & GJB6-negative families, using four microsatellite markers linked to DFNB7/11 was performed. ResultsCo-segregation of hearing loss with all investigated markers for the DFNB7/11 locus was found in one family. DNA sequencing of all coding and non-coding exons and intron boundaries of the TMC1 gene identified c.-258A>C mutation in non-coding exon 3 only in individuals with hearing loss. This mutation has been previously reported in another Iranian family (G9) that share similar ethnicity. This variant was not detected in 300 ethnically matched healthy controls. ConclusionsThese results increase the probability that this nucleotide variation may be a pathogenic mutation. This study showed that the ethnicity may be more useful than geographical location to design research strategy for determining which genes should be considered when a heterogeneous disorder is under investigation.

The first molecular genetics analysis of individuals suffering from nephropatic cystinosis in the Southwestern Iran.

Nephropatic Cystinosis (NC) is a rare metabolic disorder due to mutation in the CTNS gene in which more than 90 different mutations have already been reported so far. This study was performed to investigate mutations of the CTNS gene and its promoter in a number of Iranian patients with NC. Polymerase chain reaction and direct sequencing were performed for molecular characterization of the CTNS gene in 25 patients from 24 unrelated Iranian families with NC. None of the patients showed the 57 kb deletion in heterozygous or homozygous manner. One was homozygous for a novel mutation, which was termed as "c.153-155insCT", while one of the cases was homozygous and another was compound heterozygous for the second novel mutation c.923G>A. Moreover three known mutations c.18-21delGACT, c.1017G>A, and c.681G>A in 11 of the patients were detected. No apparent mutation was observed in the rest of patients (44%, n=11). The present data exhibit a fundament for molecular carrier detection and prenatal diagnosis of a relatively large percentage of Iranian patients suffering from NC, at least in the Southwestern Iran, where Arab ethnicity is one of the common ethnicities of the region.

Ullrich Congenital Muscular Dystrophy (UCMD): Clinical and Genetic Correlations.

Ullrich congenital muscular dystrophy (UCMD) corresponds to the severe end of the clinical spectrum of neuromuscular disorders caused by mutations in the genes encoding collagen VI (COL VI). We studied four unrelated families with six affected children that had typical UCMD with dominant and recessive inheritance. Four unrelated Iranian families with six affected children with typical UCMD were analyzed for COLVI secretion in skin fibroblast culture and the secretion of COLVI in skin fibroblast culture using quantitative RT-PCR (Q-RT-PCR), and mutation identification was performed by sequencing of complementary DNA. COL VI secretion was altered in all studied fibroblast cultures. Two affected sibs carried a homozygous nonsense mutation in exon 12 of COL6A2, while another patient had a large heterozygous deletion in exon 5-8 of COL6A2. The two other affected sibs had homozygote mutation in exon 24 of COL6A2, and the last one was homozygote in COL6A1. In this study, we found out variability in clinical findings and genetic inheritance among UCMD patients, so that the patient with complete absence of COLVI was severely affected and had a large heterozygous deletion in COL6A2. In contrast, the patients with homozygous deletion had mild to moderate decrease in the secretion of COL VI and were mildly to moderately affected.

A new ATP7B gene mutation with severe condition in two unrelated Iranian families with Wilson disease

Wilson disease is associated with a defect in copper metabolism and caused by different mutations in ATP7B gene. The aim of this study was to determine mutation frequency of ATP7B exons 8 and 14 in Wilson disease patients from the south of Iran. The exons 8 and 14 of ATP7B gene were analyzed in 65 unrelated Wilson disease patients by Denaturing High Performance Liquid Chromatography, and samples with abnormal peak profile were selected for direct DNA sequencing. Seven out of 65 (10.8%) patients had mutations at exon 14, including c.3061-1G>A in four and c.3207C>A in three patients. In addition, four different mutations were identified at exon 8 of six patients (9.2%). Three of these mutations have been previously reported, including c.2304delC in two patients, c.2293G>A and 2304dupC each in one patient. Furthermore, a novel mutation, c.2335T>G (p.Trp779Gly), was identified in two unrelated patients. The patients with this novel mutation demonstrated severe neuropsychiatric condition. All together, 13 out of 65 (20%) patients had mutations within exons 8 and 14. We also identified a lower frequency of the most common mutations of exons 8 and 14 in the southern Iranian population.

"Familial" versus "Sporadic" intellectual disability: contribution of common microdeletion and microduplication syndromes

BackgroundInterstitial Microdeletion and Microduplication syndromes have been proposed as a significant cause of sporadic intellectual disability (ID) but the role of such aberrations in familial ID has not yet been investigated. As the balanced chromosomal abnormalities commonly lead to the recurrent ID or multiple congenital anomalies, this study was designed to evaluate whether it was justified to investigate such aberrations in familial ID patients. Three hundred and twenty eight patients from 101 unrelated Iranian families with more than two ID patients in the first-degree relatives, have been investigated. Assessment of a panel of 21 common Microdeletion and Microduplication syndromes (CMMS) was carried out using Multiplex Ligation-Dependent Probe Amplification (MLPA) technique.ResultsAmong the families studied, 27.7% had 4-12, 35.6% had 3 and 36.6% had 2 affected individuals in the first-degree relatives. An autosomal dominant inheritance of Williams-Beuren syndrome (WBS) was detected in a family with no clinical suspicion of WBS. The prevalence of CMMS was therefore,0.99%.ConclusionThis is the first investigation of a panel of CMMS in a large sample set of "familial ID patients". The findings of this study showed the low prevalence of CMMSs in "familial ID" patients in spite of the significant contribution of such aberrations in "sporadic ID" which has a very useful practical impact by avoiding unnecessary diagnostic tests in "familial ID" patients.

Two Novel Mutations in SCN1A Gene in Iranian Patients with Epilepsy

Epilepsy as a common chronic neurological disorder is characterized by recurrent unprovoked seizures. Febrile seizures are the most common type of epilepsy in infants and children. Our aim was the molecular analysis of SCN1A gene in affected Iranian patients with GEFS+ and Dravet syndrome diagnosed clinically to explain genotype-phenotype correlation and exact classification. The 34 unrelated Iranian families with epilepsy were selected and screened for SCN1A mutations by MLPA, ARMS, and PCR-RFLP confirmed by direct sequencing. MLPA analysis showed normal patterns, but direct sequencing revealed that generally 20/34 (0.588) probands have common reported single nucleotide polymorphisms (SNPs) (p.A1067G; rs2298771) with allelic frequency as 0.706/0.294 in patients and 0.515/0.485 in control group, respectively, for A/G. No significant differences between groups were observed. Moreover, four novel allelic variants as missense substitutions included two new sequence variation (p.F412 I, p.Y1274N) and two previously reported mutations (p.R101G, p.S103G) that were detected in 4/34 probands but not in control groups and other healthy normal family members. Clinical diagnosis could nearly establish the classification, but mutation screening helps clinicians to confirm their data. We found mutation in four probands and confirmed the net diagnosis. Our data suggest that the clinical symptom variations could be also explained, considering the role of modifier genes such as mitochondrial mutations or other genes responsible for drug metabolism pathways including multiple drug resistance family genes (ABCB1) or MTHFR.

First Study of CF Mutations in the CFTR Gene of Iranian Patients: Detection of F508, G542X, W1282X, A120T, R117H, and R347H Mutations

Thirty-seven unrelated Iranian CF families were screened for the presence of seven common mutations (DeltaF508, G542X, W1282X, G551D, N1303K, 1717-1G-->A, and 621-1G-->T) using ARMS PCR and exons 4 and 7 of the CFTR gene by SSCP method. This study resulted in the identification of 26.8 per cent of all CF alleles: DeltaF508 (16.2 per cent), W1282X (4 per cent), G542X (2.7 per cent), R117H (1.3 per cent), R347H (1.3 per cent), and A120T (1.3 per cent) mutations were detected. To the best of our knowledge, it is the first report of an Asian subject carrying the A120T mutation. Our findings suggest heterogeneity in the Iranian population, stressing the need to draw attention to sequence analysis in order to find population-specific mutations.

Isolated congenital anosmia with morphologically normal olfactory bulb in two Iranian families: a new clinical entity?

Congenital total loss of the sense of smell occurs as a part of a syndrome or isolated anosmia. Kallmann syndrome is the most well known congenital anosmia associated with hypogonadotropic hypogonadism. Isolated congenital anosmia (ICA) is a very rare condition and appears to be due to changes in the olfactory epithelium or to aplasia of the olfactory nerve, bulb, and tract. Here we report two unrelated Iranian families with ICA. One family consisted of nine affected members, and the other family contained three affected members. Clinical history, physical examination, and smell testing by intravenous injection of combined vitamins (Alinamin trade mark, Takeda Pharmaceutical Co. Ltd., Japan) confirmed the disease in each affected member. No signs of hypogonadism or other neurological disorders were observed in any affected members. Family analysis with the complete ascertainment method under assumption of the same condition in the two families suggested that the disease is not inconsistent with an autosomal dominant mode with incomplete penetrance. The inheritance in one family appears unusual, i.e., there were no affected individuals in the third generation. When only two upper generations in the family are concerned, the segregation ratio was 0.39 +/- 0.11. Male-to-male transmissions were observed and both sexes were affected in both families. Magnetic resonance imaging (MRI) of the olfactory bulb and sulcus revealed no evidence of morphological changes in all affected members, suggesting that these patients have either a defect in the olfactory epithelium or a functional defect in the olfactory cortex.

Incidence of phenylketonuria in Iran estimated from consanguineous marriages.

The incidence of phenylketonuria (PKU; McKusick 261600) varies widely in different parts of the world. While the average incidence of PKU in caucasians is approximately 1 in 10 000, it differs severalfold among different populations. Neonatal mass screening is a valuable source of incidence data. Alternatively, incidence can be estimated from rates of consanguineous marriages. If the frequency of consanguineous ¢rst-cousin marriages in the general population (C), and the frequency of ¢rst-cousin marriages among parents of children homozygous for a given allele (C 0) are known, the frequency ðqÞ of that allele can be estimated from Dahlberg’s formula q 1⁄4 1⁄2Cð1 C0Þ =1⁄2Cð1 C0Þ þ 16ðC 0 CÞ (Dahlberg 1947). For rare autosomal recessive diseases, if both C and C0 are known, the frequency of the disease gene ðqÞ, and therefore of the disease, can be estimated. This approach has been used for estimating the incidence of several autosomal recessive diseases, including the incidence of PKU in Norway and in Italy. These estimates are in agreement with values obtained from neonatal screening data. This method is especially useful in those countries where neonatal mass screening for hyperphenylalaninaemia has not yet been instituted. We used this approach to estimate the incidence of PKU in Iran. Several lines of evidence point to a high incidence of PKU in this country: one of the highest values for the incidence of PKU has been reported from Iran’s neighbouring country, Turkey (1 in 4370 live births; Ozalp et al 1990). Our studies suggest major similarities in the spectrum of mutations responsible for PKU in the two countries (Hosseini-Mazinani and Koochmeshgi, unpublished data). Furthermore, incidence of PKU among the institutionalized mentally retarded in Iran is unusually high. In the present study, 82 unrelated Iranian PKU families were enrolled. These families had been referred to a national PKU centre from all over the country. Frequency of ¢rst-cousin marriages among parents of PKU subjects was determined using a questionnaire. In 46 families (56%), parents were ¢rst cousins. The frequency of ¢rst-cousin marriages among Iranian Muslims, who comprise 99% of the Iranian general population, is 17% (Farhud et al 1991). Assuming that consanguineous marriages have been a long-standing custom in Iran, q (the PKU gene frequency) can be calculated to be 0.0118. Woolf has calculated this value for Turkey to be 0.0108 (Woolf 1994). Both these values are lower that the PKU gene frequency in Ireland and West Scotland (0.0135^0.0142; Woolf et al 1975). The inbreeding coe⁄cient ðF Þ in the Iranian Muslim population is 0.0117 (Farhud et al 1991), J. Inherit. Metab. Dis. 25 (2002) 80^81 # SSIEM and Kluwer Academic Publishers. Printed in the Netherlands.