Identification of a new heterozygous point mutation in the COL1A2 gene leading to skipping of exon 9 in a patient with joint laxity,hyperextensibility of skin and blue sclerae

A heterozygous deletion of exon 9 in the COL1A2 - mRNA of a patient with symptoms of both the Ehlers - Danlos - Syndrome and the Osteogenesis Imperfecta is described. In the genomic DNA of the patient, exon 9 is homozygously present. We identified a novel heterozygous point mutation in the splice donor site of intron 9, leading to a G→A substitution in position +5. This mutation leads to heterozygous skipping of exon 9 in the COL1A2 - mRNA of this patient. The deletion results in a shortened (by 18 amino acids) but in frame l2(1) chain, which probably leads to the formation of abberantly processed triple helices. Hum Mutat 12:138, 1998. © 1998 Wiley-Liss, Inc.     

Noval mutation (Y184C) in exon 4 of the beta-sarcoglycan gene identified in a Portuguese patient. Mutations in brief no. 177. Online

We report a novel beta-sarcoglycan gene mutation identified in a 21-year-old Portuguese male with a progressive myopathy of intermediate severity, who had been misdiagnosed as Becker Muscular Dystrophy (BMD) based on clinical observations and muscle immunocytochemical anaylsis with dystrophin antibodies only. Since no detectable deletions or duplications were found in the dystrophin gene, we screened for mutations in the sarcoglycan genes by PCR-SSCP. The patient's sample showed a band of increased mobility in exon 4 of the beta-sarcoglycan gene which, upon sequencing, was found to represent a homozygous A-->G transversion at nucleotide 551, resulting in a tyrosine to cysteine substitution at position 184 (Y184C). Carrier status was ascertained in both parents and a sister. These aberrant conformers were not detected in 85 unrelated control individuals screened by PCR-SSCP analysis. All seven beta-sarcoglycan mutations reported to date are associated with a severe phenotype and occur in exons 3 and 4, which correspond to the immediate extracellular domain of the protein. This region contains five conserved cysteine residues. In our patient, the presence of an extra cysteine residue could interefere with intra- and/or inter-molecular disulphide bond formation. The intermediate phenotype could perhaps result from the assembly of both normal and abnormal complexes, depending on the formation of the disulphide bonds.     

A novel point mutation in a splice acceptor site of intron 1 of the human low density lipoprotein receptor gene which causes severe hypercholesterolemia: an unexpected absence of exon skipping. Mutations in brief no. 139. Online

Familial hypercholesterolemia (FH) is a genetic disorder caused by mutations in the low density lipoprotein (LDL)-receptor gene. We found a new mutation in the splice acceptor site of intron 1 of the LDL receptor gene, which is designated as 68-1 G->C according to the nomenclature suggested by Beaudet and Tsui (1993), in a Japanese FH homozygote. She was born from consanguineous marriage and has this mutation as a true homozygous form. Her cultured fibroblasts showed no LDL receptor protein synthesis. This mutation caused activation of a cryptic splice acceptor side in the downstream exon 2, leading to frameshift and appearance of premature in-frame stop codon. The mutation was detected by Dde I restriction enzyme. The identical mutation was not found among 24 patients with homozygous and 120 patients with heterozygous FH. The mutation was very rare among the Japanese population.     

Identification of the mutation in the alkaptonuria mouse model. Mutations in brief no. 216. Online

Alkaptonuria (aku), an inborn error of metabolism caused by the loss of homogentisate 1,2-dioxygenase (HGD), has been described in a mouse model created by ethylnitrosourea mutagenesis but the mutation in these mice has not previously been identified. We used RT-PCR to amplify the Hgd cDNA from Hgd(aku)/Hgd(aku) mice. Two products shorter than the wild-type product were amplified. Restriction mapping and DNA sequencing were then used to identify the Hgd(aku) mouse mutation, found to be a single base change in a splice donor consensus sequence, causing exon skipping and frame-shifted products. This base change allowed us to create a non-radioactive genotyping assay for this allele.     

18 bp insertion/duplication with internal missense mutation in human hepatic lipase gene exon 3. Mutations in brief no. 181. Online

Human hepatic lipase (hHL) plays an important role in hydrolysis of triglycerides from plasma lipoproteins. The enzyme also hydrolyzes HDL2 lipids resulting in smaller HDL particles with a lower cholesterol content and properties similar to HDL3. hHL is localized in liver sinusoids, ovary and adrenal gland. These findings propose an influence on processing of cholesterol. Here we report an insertion mutation in exon 3 of hHL. The 18 bp duplication contains an additional internal point mutation (GenBank-Accession #AF037404). The female mutation carrier suffered from severe adiposity with total cholesterol of 291,6mg/dl, HDL-cholesterol of 55,3mg/dl, LDL-cholesterol of 206,8mg/dl and triglycerides of 80,8mg/dl. Following cloning of a PCR-amplified fragment the mutation was confirmed by cycle sequencing. Sequence analysis revealed an inserted repeat of 18 nucleotides. Furthermore the patient carries an additional missense mutation A-->G at nucleotide 9 of the repeat which results in an amino acid exchange from Ile-->Val at codon 4 of the repeat. These data enable us to report the insertion of HisTyrThrValArgVal which might be responsible for the moderate shift in lipid metabolism of the heterozygous patient.     

An exon skipping mutation of a type V collagen gene (COL5A1) in Ehlers-Danlos syndrome.

The Ehlers-Danlos syndrome (EDS) is a heterogeneous group of inherited connective tissue disorders characterised by skin hyperextensibility, joint hypermobility, easy bruising, and cutaneous fragility. Nine discrete clinical subtypes have been classified. We have investigated the molecular defect in a patient with clinical features of Ehlers-Danlos syndromes types I/II and VII. Electron microscopy of skin tissue indicated abnormal collagen fibrillogenesis with longitudinal sections showing a marked disruption of fibril packing giving very irregular outlines to transverse sections. Analysis of the collagens produced by cultured fibroblasts showed that the type V collagen had a population of alpha 1 (V) chains shorter than normal. Peptide mapping suggested a deletion within the triple helical domain. RTPCR amplification of mRNA covering the whole of this domain of COL5A1 showed a deletion of 54 bp. Although six Gly-X-Y triplets were lost, the essential triplet amino acid sequence and C-propeptide structure were maintained allowing mutant protein chains to be incorporated into triple helices. Genomic DNA analysis identified a de novo G+3-->T transversion in a 5' splice site of one COL5A1 allele. This mutation is analogous to mutations causing exon skipping in the major collagen genes, COL1A1, COL1A2, and COL3A1, identified in several cases of osteogenesis imperfecta and EDS type IV. These observations support the hypothesis that type V, although quantitatively a minor collagen, has a critical role in the formation of the fibrillar collagen matrix.     

First mutation (S340X) in choroideremia gene in a Spanish family. Mutations in brief no. 173. Online

A study of choroideremia gene was performed in Spanish families affected with this disorder. One abnormal pattern was detected in exon eight corresponding to a new mutation not described before. The mutation was identified as a nonsense mutation S340X.     

A novel missense mutation D513G in exon 10 of the cystic fibrosis transmembrane conductance regulator (CFTR) gene identified in a French CBAVD patient. Mutations in brief no. 175. Online

Congenital bilateal absence of the vas deferens (CBAVD) with obstructive azoospermia is a congenital reproductive disorder that affects one in 10000 male individuals. The observation that many men presenting with CBAVD have mutations in their CFTR genes had led to the proposal that CBAVD may be a primary genital form of cystic fibrosis. We report here one novel mutation located in exon 10 of the CFTR gene. This mutation, named D513G (A-->G at position 1670), has been found in one of 83 patients with CBAVD from France, the analysis of exon 10 using a chemical clamp DGGE assay allowed us to identify three CF mutations AEF508 (37/166; 22%), AE1507 (1/166; 0/6%) and D513G (1/166; 0.6%), and two variants M470V and E528E (1716 G>A). The novel D513G mutation has not been found in more than 200 non-CF chromosomes and in a sample of 300 CF chromosomes from French classical CF patients.     

A novel mutation (V191G) in a German-British type 1 Gaucher disease patient. Mutations in brief no. 131. Online

Gaucher disease results from mutations in the glucocerebrosidase gene located on human chromosome 1q21. Three clinical forms of Gaucher disease have been described: type 1, nonneuropathic; type 2, acute neuropathic; and type 3, subacute neuropathic. We have identified a novel mutation in a German-British patient with type 1 Gaucher disease which results in V191G of the glucocerebrosidase polypeptide. Because the mutation abolishes a HphI cleavage site, its presence was confirmed by HphI RFLP analysis of PCR-amplified genomic DNA. In the second allele of the patient, the mutation identified was g.5841A G(N370S). Sequence analysis of the remainder of the coding region of the gene as well as the exon-intron boundaries showed identity to normal controls. Because mutation N370S has so far been found only in type 1 Gaucher disease and postulated to result in mild clinical presentation, and since the clinical course of this patient has been relatively mild with minimal skeletal involvement, we speculate that the V191G/N370S genotype may also result in good prognosis.     

Identification of 9 novel IDS gene mutations in 19 unrelated Hunter syndrome (mucopolysaccharidosis Type II) patients. Mutations in brief no. 202. Online

Hunter syndrome is an X-linked lysosomal storage disorder caused by a deficiency of the lysosomal enzyme iduronate-2-sulfatase (IDS). The IDS deficiency can be caused by several different types of mutations in the IDS gene. We have performed a molecular and mutation analysis of a total 19 unrelated MPS II patients of different ethnic origin and identified 19 different IDS mutations, 9 of which were novel and unique. SSCP analysis followed by DNA sequencing revealed four novel missense mutations: S143F, associated with the 562C-->T polymorphism, C184W, D269V and Y348H. Two novel nonsense mutations were found: Y103X (433C-->A) and Y234X (826C-->G). In two patients two novel minor insertions (42linsA and 499insA) were identified. In one patient a complete IDS deletion was found, extending from locus DXS1185 to locus DXS466).     

A novel mutation of the down-regulated in adenoma gene in a Japanese case with congential chloride diarrhea. Mutations in brief no. 198. Online

Congenital chloride diarrhea (CLD) is an autosomal recessive disease characterized by excretion of watery stool with a high chloride content. Pathogenesis of CLD is a deficient absorption of chloride in exchange for bicarbonate in the ileum and the colon. In 1996, it was reported that 36 patients with CLD had mutations in the down-regulated in adenoma (DRA) gene; 32 Finnish patients had a three base deletion (951delGGT), 2 Polish patients had a one base mutation (371AtoT) and 2 Polish patients had a one base deletion (344delT). In this study we analyzed the DRA gene in a Japanese boy patient with CLD and in members of his family. The patient was found to have a two base deletion (TT) at nucleotide 1526-1527 within codon 509 which results in a frameshift leading to a permature stopping at codon 517. The patient was homozygous for the deletion, his parents and brother were heterozygous, and his sister was normal. This is the first case of CLD identified to carry a mutation of the DRA gene in Asia.     

A novel splice site mutation of the EXT2 gene in a Finnish hereditary multiple exostoses family. Mutations in brief no. 197. Online

Hereditary multiple exostoses is a dominantly inherited disease characterized by multiple benign osteochondromas. The affected individuals have an increased risk of developing sarcoma. A large Finnish family with hereditary multiple exostosis was analyzed to find the disease-causing mutation. Blood samples were obtained from 35 family members, including 21 affected and 14 unaffected individuals. Using 2-point linkage analysis the EXT phenotype was shown to be linked to the recently cloned EXT2 gene on chromosome 11p11. The coding region of the gene was sequenced and a previously unreported splice site mutation found. This G to T transversion within a 5-prime splice donor site following exon 6 was shown to cause aberrant splicing of RNA. The described change is considered to be a novel disease-causing mutation in the EXT2 gene.     

T426I a new mutation in the thyroid hormone receptor beta gene in a sporadic patient with resistance to thyroid hormone and dysmorphism. Mutations in brief no. 192. Online

Resistance to thyroid hormone (RTH) is a rare inherited autosomal syndrome caused by mutations in the thyroid hormone receptor beta (TRb) gene. Although RTH is generally a familiar disease, 15% of sporadic cases have been also reported. So far, about 80 different mutations of TRb gene have been identified in patients affected by RTH. All these mutations localize to the binding domain and most of them cluster within two "hot spots" (codons 310-349 and codons 429-460). Here we describe in a patient with RTH, a new mutation in codon 426 (T426I) of the TRb gene leading to a threonine to isoleucine substitution. This is a "de nova" mutation which localizes in the so-called "cold" region, outside the two known "hot spots". The patient had the hallmark of RTH: elevated FT3 and FT4, normal TSH, and clinical features of both hypo and hyperthyroidism. Moreover, several dysmorphisms were present including triangular face appearance, synophris, low set ears, micrognathia with malocclusion, large upper incisors and apparent lack of lower cuspids which have not previously described in RTH patients.     

Identification of a D579G homozygote cystic fibrosis patient with pancreatic sufficiency and minor lung involvement. Mutations in brief no. 221. Online

Here we describe the identification of an italian patient homozygote for the D579G mutation affected by a mild form of Cystic Fibrosis with pancreatic sufficiency, minor lung involvement and marked viscosity of the cervical mucous. The D579G mutation causes an A1868G transition, a substitution of an aspartic acid to a glycine residue, generating an important amino acid change (charged to hydrophobic) in the nucleotide-binding domain (NBD). The mutation was first described by Brancolini et al. (1995) on two pancreatic sufficient CF patients, compound heterozygotes for delta508F. Patients were from Southern Italy (Puglia) as the D579G homozygote one, who is a 30 years old woman from Taranto (Puglia), daughter of second cousins born in Bari (Puglia). The identification of a homozygote D579G patient might confirm that this mutation does correlate with pancreatic sufficiency and a mild pulmonary phenotype.     

Identification of cystic fibrosis mutations in the United Arab Emirates. Mutations in brief no. 133. Online

We have designed a study aimed at identifying the genetic mutations responsible for cystic fibrosis (CF) in the population of the United Arab Emirates. The prevalence of CF in the UAE is at least 1/15,000 live births and the disease is associated with very severe clinical presentations. We have investigated 17 unrelated families. Ten UAE national families were of Bedouin descent: all 15 CF patients, who presented with very severe forms of the disease, were homozygous for a S549R mutation due to a T->G transversion at nucleotide postion 1779. Amongst a distinct population of Baluch origin, CF patients from 6 out of 7 affected families were DF508 homozyotes. Hence, the unique distribution of CF mutations in the United Arab Emirates--two mutations, S549R and DF508, characterize so far 94% of CF families--should allow efficient organizing and delivering of CF carrier screening programmes on the country's relatively limited population size.     

Identification of 8 new mutations in Brazilian families with Marfan syndrome. Mutations in brief no. 211. Online

Marfan Syndrome (MFS) is a connective tissue disease caused by mutations in the fibrillin-1 FBN1) gene. Screening for mutations in all the 65 exons of the FBN1 gene in 34 unrelated patients were performed to compare the efficiency of SSCP versus Heteroduplex analysis and to verify if the spectrum of mutations in Brazilian patients is similar to the one previously reported. Fourteen different band shifts were detected by SSCP analysis; among these only 6 were also were also detected through Heteroduplex analysis, suggesting that SSCP analysis was a more efficient method. Except for one, the molecular alteration was confirmed in the remaining 13 cases by sequencing; five of them were neutral polymorphisms and the eight others are new pathogenic mutations, as follows: 5 missense, one nonsense and two deletions leading to a premature termination codon (PTC). All of them are located in EGF-like-calcium binding motifs (EGF-like-cb). Our findings reinforce that cysteine substitutions and PTC mutations in the region between exons 24-32 are more likely not to be associated with the neonatal phenotypes.     

Identification of a new SCN5A mutation, D1840G, associated with the long QT syndrome. Mutations in brief no. 153. Online

The long QT syndrome (LQT) is an inherited cardiac disorder that can cause sudden cardiac death among apparently healthy young individuals due to malignant ventricular arrhythmias. LQT was found to be caused by mutations in four genes LTQ1, LQT2, LQT3 and LQT5, and linkage was reported for an additional locus, LQT4, on chromosome 4q25-27. We have studied a large (n=131) LQT-affected Jewish kindred and identified tight linkage between the LQT-affected status and LQT3 (lod score 6.13, with an estimated recombination fraction of zero). We identified a new point-mutation, A to G substitution at nucleotide 5519 of the SCN5A gene, changing the aspartate 1840 to glycine, D1840G. This is a non-conservative change of an amino acid completely conserved in sodium channels from Molusca to human. The mutation was identified in all affected individuals (n=23), and not identified in all the unaffected family members (n=40), and not in 200 chromosomes of healthy control individuals. The mutation was identified in 3/12 individuals with equivocal phenotype, thus, providing an accurate dignostic tool for all family members. This mutation is currently being used in a cellular electrophysiological model, to characterize the function of the mutated sodium channel in this syndrome.