Frameshift Mutation In Exon Research Articles

Genomic organization and characterization of the white locus of the Mediterranean fruitfly, Ceratitis capitata.

An approximately 14-kb region of genomic DNA encoding the wild-type white eye (w+) color gene from the medfly, Ceratitis capitata has been cloned and characterized at the molecular level. Comparison of the intron-exon organization of this locus among several dipteran insects reveals distinct organizational patterns that are consistent with the phylogenetic relationships of these flies and the dendrogram of the predicted primary amino acid sequence of the white loci. An examination of w+ expression during medfly development has been carried out, displaying overall similarity to corresponding studies for white gene homologues in Drosophila melanogaster and other insects. Interestingly, we have detected two phenotypically neutral allelic forms of the locus that have arisen as the result of an apparently novel insertion or deletion event located in the large first intron of the medfly white locus. Cloning and sequencing of two mutant white alleles, w1 and w2, from the we,wp and M245 strains, respectively, indicate that the mutant conditions in these strains are the result of independent events--a frameshift mutation in exon 6 for w1 and a deletion including a large part of exon 2 in the case of w2.

Fh-Souassi: a founder frameshift mutation in exon 10 of the LDL-receptor gene, associated with a mild phenotype in Tunisian families

Familial hypercholesterolemia (FH) has a higher prevalence in central Tunisia together with a milder clinical expression than in western countries. The molecular basis of FH in Tunisia remains unknown. Our aim was to identify FH-causing mutations in three unrelated families (21 subjects) from the area of Souassi (central Tunisia). In probands with a presentation of homozygous FH, the promoter and 18 exons of the low density lipoprotein (LDL)-receptor gene were sequenced in both orientations. A novel complex frameshift mutation was identified in exon 10, nucleotides 1477–1479 (TCT) at Serine 472 were replaced by an insertion of seven nucleotides (AGAGACA), producing a premature termination codon 43 amino acids downstream. Binding of 125I-labelled LDL at 4°C to cultured fibroblasts from two probands showed <2% normal LDL-receptor activity. AvaII digestion of PCR amplified genomic DNA identified this unique mutation in all families; homozygotes n=11, heterozygotes n=10. All mutation carriers shared the same haplotype (7 RFLPs), suggesting that they had a common ancestor. Despite high plasma LDL levels ( m=16.0±3.0 mmol/l) and extravascular cholesterol deposits, most homozygotes were diagnosed after puberty and had a delayed onset of cardiovascular complications. Moreover, most heterozygotes were free of clinical signs and had plasma LDL cholesterol in the normal range (4.7±1.3 mmol/l) without taking any lipid-lowering medication. This mild clinical phenotype which contrasted with the severity of the mutation, could not be explained by specific apolipoprotein E or lipoprotein lipase alleles.

Novel mutations in the emerin gene in Israeli families.

Emery-Dreifuss Muscular Dystrophy (EMD or EDMD) is a rare X-linked recessive disorder, characterized by progressive muscle wasting and weakness, contractures, and cardiomyopathy, manifesting as heart block. Mutation analysis at the EMD gene locus was performed in 4 unrelated Israeli families with X-linked EMD and in one sporadic case. In the 4 families 4 different mutations were found, 3 of which were novel. These included two frame shift mutations in exon 2 (333delT and 412insA) and one base pair substitution at the consensus +1 donor splice in intron 5 (1429G-->A). The fourth mutation in exon 6 (1675-1678delTCCG) has been previously described. No mutations were identified in the one sporadic case. Two of the three novel mutations were found in exon 2. A summary of the previously published mutations described in the EMD Mutation Database (http://www.path.cam.ac.uk/emd/) as well as the mutations described in our study suggest that the distribution of mutations in EMD gene is not entirely random and that exon 2 is prone to mutations. Hum Mutat 17:522, 2001.

Mutations in the Hepatocyte Nuclear Factor-1β Gene Are Associated with Familial Hypoplastic Glomerulocystic Kidney Disease

Familial glomerulocystic kidney disease (GCKD) is a dominantly inherited condition characterized by glomerular cysts and variable renal size and function; the molecular genetic etiology is unknown. Mutations in the gene encoding hepatocyte nuclear factor (HNF)-1beta have been associated with early-onset diabetes and nondiabetic renal disease-particularly renal cystic disease. We investigated a possible role for the HNF-1beta gene in four unrelated GCKD families and identified mutations in two families: a nonsense mutation in exon 1 (E101X) and a frameshift mutation in exon 2 (P159fsdelT). The family members with HNF-1beta gene mutations had hypoplastic GCKD and early-onset diabetes or impaired glucose tolerance. We conclude that there is genetic heterogeneity in familial GCKD and that the hypoplastic subtype is a part of the clinical spectrum of the renal cysts and diabetes syndrome that is associated with HNF-1beta mutations.

The chromosome 9q genes TGFBR1, TSC1, and ZNF189 are rarely mutated in bladder cancer.

This study assessed a series of bladder tumours and bladder tumour cell lines for sequence variation in the Krüppel-like zinc finger gene ZNF189, the tuberous sclerosis complex gene 1 (TSC1), and the TGF beta receptor type I (TGFBR1). All three genes have been mapped to 9q regions commonly deleted in transitional cell carcinoma of the bladder. Mutation analysis of the coding sequence of these genes revealed several variant bands that were shown to represent polymorphisms. Mutation analysis of the ZNF189 gene in bladder cancer cell lines identified one amino acid substitution (lysine-->isoleucine) at position 323 in exon 4. For the TSC1 gene, two mutations were identified in two out of 27 independent cell lines. Both mutations result in a truncated protein. Furthermore, one out of 36 bladder tumours had a frameshift mutation in exon 7 of the TSC1 gene. No tumour-specific mutations were found in the TGFBR1 gene. The length of the polyalanine tract present in exon 1 of the TGFBR1 gene was also investigated. It has been suggested that the allele with six alanines (6A) is more frequent in patients with bladder and other cancers, so bladder cancer patients were compared with normal controls. In both groups, the percentage of heterozygotes was 17%. These data do not support a role for the 6A allele in bladder cancer susceptibility.

Non-erythroid form of acute intermittent porphyria caused by promoter and frameshift mutations distant from the coding sequence of exon 1 of the HMBS gene

Acute intermittent porphyria (AIP) is a low-penetrant, autosomal dominant disorder caused by mutations in the HMBS gene. The gene is transcribed from two promoters to produce ubiquitous and erythroid isoforms of porphobilinogen deaminase, which differ only at their NH2 ends. In the classical form of AIP, both isoforms are deficient, but about 5% of families have the non-erythroid variant in which only the ubiquitous isoform is affected. Previously identified mutations in this variant have been within or close to the coding region of exon 1 of the HMBS gene, the only exon that is expressed solely in the ubiquitous isoform. Here, we describe mutations in the ubiquitous promoter (-154delG) and in exon 3 (41delA) that cause the non-erythroid variant. Reporter gene and electrophoretic mobility shift assays show that the G nucleotide at position -154, the most 5' of several transcription-initiation sites in the ubiquitous HMBS promoter, which lies immediately 3' to a transcription-factor IIB binding motif, is essential for normal transcription. The frameshift mutation in exon 3 introduces a stop codon into mRNA for the ubiquitous isoform only. Our investigations identify two new mechanisms for production of the non-erythroid variant of AIP and demonstrate that mutational analysis for diagnosis of this variant needs to include wider regions of the HMBS gene than indicated by previous reports. Furthermore, they show that deletion of one of several transcription initiation sites in the promoter of a housekeeping gene that lacks both TATA and initiator elements can produce disease.

Novel germline BRCA1 mutations detected in women in singapore who developed breast carcinoma before the age of 36 years.

In recent years, although BRCA1 has been extensively investigated, the contribution of inherited BRCA1 mutations to breast carcinoma in Asian populations is largely unknown. The authors undertook this study to determine the prevalence and spectrum of germline BRCA1 mutations among women in Singapore with early onset breast carcinoma. Forty-three of 72 eligible patients whose breast carcinoma was diagnosed before the age of 36 years were studied, independent of family history. DNA samples from 50 unrelated individuals randomly selected from the National Thalassemia Registry served as controls. Mutational screening was performed by single-strand conformation polymorphism analysis and protein truncation test, and alterations were confirmed by sequencing. First-degree relatives of patients with definite BRCA1 mutations were offered screening. A total of 6 novel alterations in BRCA1 were identified, including 2 frameshift mutations in exon 11 (2846insA and 2885delA), 3 rare sequence variants, and 1 polymorphism. Three women (7%) carried deleterious mutations, and the mutation was present in at least 1 unaffected first-degree relative of the proband. The same mutation (2846insA) was identified in 2 of the 7 unrelated subjects of Malay ethnicity. One mutation and three rare variants were identified in four women with no family history of breast or ovarian carcinoma whereas all women with affected first-degree relatives did not harbor BRCA1 mutations. No mutation was identified in the controls. The spectrum of germline BRCA1 mutations among the patients in this study was distinct from that in Caucasian populations although a similar prevalence was observed. Larger studies are necessary to clarify the significance of the mutation 2846insA in the Malay community and the penetrance of specific mutations in the Singapore population.

Elastic-Fiber Pathologies: Primary Defects in Assembly—and Secondary Disorders in Transport and Delivery

Accession numbers and the URL for data in this article are as follows:Online Mendelian Inheritance in Man (OMIM), www.ncbi.nlm.nih.gov/Omim (for congenital contractural arachnodactyly [MIM 121050], adCL [MIM 123700], MFS [MIM 154700]), SVAS [MIM 185500], Williams-Beuren syndrome [MIM 194050], Costello syndrome [MIM 218040], GM1-gangliosidosis [MIM 230500, MIM 230600, and MIM 230650], MPSI [MIM 252800], and MPS IVB [MIM 253010])

Abnormal nephron development associated with a frameshift mutation in the transcription factor hepatocyte nuclear factor-1β1: 1See Editorial by Woolf, p. 1202

The transcription factor hepatocyte nuclear factor (HNF)-1 beta functions as a homodimer or as a heterodimer with the structurally related protein HNF-1 alpha. Both are expressed sequentially in rat kidney development, with HNF-1 beta being detected from the earliest inductory phases. HNF-1 beta gene mutations are associated with a unique disorder characterized by maturity-onset diabetes of the young (MODY) and early-onset and progressive nondiabetic renal dysfunction, which may lead to chronic renal failure. The HNF-1 beta gene was screened for mutations in six subjects with early-onset diabetes and a history of renal dysfunction in the subjects or their families. A novel frameshift mutation in exon 4 of the HNF-1 beta gene and a deletion of CCTCT at codons 328 to 329 were detected in one subject. She was diagnosed as diabetic at the age of 21 in her second pregnancy. Glucose tolerance rapidly deteriorated over 18 months as a result of beta-cell dysfunction. The HNF-1 beta mutation arose de novo on a paternal chromosome and cosegregated with renal abnormalities in her family. The proband had bilateral small cysts in normal-sized kidneys and a reduced creatinine clearance of 66 mL/min (NR 80-120). Her first pregnancy was terminated at 17 weeks following an ultrasound diagnosis of bilateral, nonfunctioning cystic kidneys. Her first-born child had a small multicystic, dysplastic right kidney and a dysplastic left kidney with a reduced creatinine clearance (40 mL/min per 1.73 m2). Histologic examination of the large (5.8 vs. 1.4 g), polycystic fetal kidneys showed no normal nephrogenesis. These studies indicate that HNF-1 beta plays a central role in normal kidney development and pancreatic beta-cell function, and suggest that one mechanism by which HNF-1 beta gene mutations may cause renal dysfunction are by their effects on nephron development.

Frameshift mutations with severe and moderate clinical phenotypes in Thai hemophilia A patients.

Six frameshift mutations in exon 14 of the factor VIII gene were identified in Thai hemophilia A patients. Although all these mutations created premature stop codons and expected to cause severe disease, the molecular defects and clinical severity were in discrepancy in some patients. Four mutations (delT3490, delACAC3618-21, delGA4429-30, and delA4658) were found in the patients with the severe clinical phenotype while two (delA3629-37 and insA4372-9) were observed in the patients who had moderate severity, with FVIII:C of 4.2 and 2.8%. The frameshift mutations in these two patients were due to deletion and insertion of an 'A' nucleotide in the stretches of 9As and 8As in codons 1191-4 and 1439-41, respectively. This indicates that deletion or insertion in the stretches of poly A nucleotides in exon 14 of the factor VIII gene is a likely cause of the moderate clinical severity in some cases of Thai hemophilia A patients.

A complete deficiency of coagulation factor XIII A-subunit due to a novel compound heterozygote of Ser 413 Leu missense and an nt 389 (ins G) frameshift mutation.

Coagulation factor XIII consists of two A- and two B-subunits, and either gene mutation can cause a complete deficiency. In a newborn patient with persistent bleeding from the umbilical cord stump, the plasma A-subunit protein was not detectable. Direct PCR sequencing revealed an nt 389 (ins G) frameshift mutation in exon 4 resulting in a new stop codon and a Ser 413 Leu missense mutation in exon 10 in either allele. His mother and father were heterozygous for the nt 389 (ins G) and the Ser 413 Leu, respectively, with about 50% reduction of the plasma A-subunit proteins. In all family members examined only those with either mutation showed the reduced subunit A protein levels. Thus, this complete deficiency of factor XIII was due to a novel compound heterozygous mutation in the A-subunit gene.

Moderation of Phenotypic Severity in Dystrophic and Junctional Forms of Epidermolysis Bullosa Through In-Frame Skipping of Exons Containing Non-Sense or Frameshift Mutations

Non-sense mutations on both alleles of either the type VII collagen gene (COL7A1) or the genes encoding laminin 5 (LAMA3, LAMB3, or LAMC2) usually result in clinically severe forms of recessive dystrophic or junctional epidermolysis bullosa, respectively. In this study we assessed two unrelated families whose mutations in genomic DNA predicted severe recessive dystrophic epidermolysis bullosa or junctional epidermolysis bullosa phenotypes but in whom the manifestations were milder than expected. The recessive dystrophic epidermolysis bullosa patients had a homozygous single base-pair frameshift mutation in exon 19 of COL7A1 (2470insG). Clinically, there was generalized blistering but only mild scarring. Skin biopsy revealed positive type VII collagen immunoreactivity and recognizable anchoring fibrils. The junctional epidermolysis bullosa patients were compound heterozygotes for a frameshift/non-sense combination of mutations in exons 3 and 17 of LAMB3 (29insC/Q834X). These patients did not have the lethal form of junctional epidermolysis bullosa but, as adults, displayed the milder generalized atrophic benign epidermolysis bullosa variant. There was undetectable laminin 5 staining at the dermal-epidermal junction using an antibody to the beta3 chain, but faintly positive alpha3 and gamma2 chain labeling, and there was variable hypoplasia of hemidesmosomes. To explain the milder recessive dystrophic epidermolysis bullosa and junctional epidermolysis bullosa phenotypes in these families, reverse transcription-polymerase chain reaction, using RNA extracted from frozen skin, was able to provide evidence for some rescue of mutant mRNA transcripts with restoration of the open- reading frame. In the recessive dystrophic epidermolysis bullosa patients, transcripts containing in-frame skipping of exon 19 of COL7A1 in the cDNA were detected, and in the junctional epidermolysis bullosa patients transcripts with in-frame skipping of exon 17 of LAMB3 were identified. The truncated proteins encoded by these transcripts are expected to lack certain critical domains involved in cell-matrix attachment, but may still be able to contribute to adhesion thereby moderating the severity of the skin blistering. This study shows the limitations in predicting phenotype in epidermolysis bullosa solely based on mutation analysis of genomic DNA and emphasizes the importance of immunohistochemistry, electron microscopy, and mRNA assessment as parallel investigations.

Diffuse Type Gastric and Lobular Breast Carcinoma in a Familial Gastric Cancer Patient with an E-Cadherin Germline Mutation

E-Cadherin alterations have been reported frequently in sporadic diffuse type gastric and lobular breast carcinomas. Germline mutations of this gene have been identified recently in several gastric cancer families. We analyzed seven patients with a family history of the disease who had diffuse type gastric cancer diagnosed before the age of 45 for germline mutations in CDH1, the gene encoding the E-cadherin protein. We identified a frameshift mutation in exon 3 in one patient with a strong family history of gastric cancer. The same germline mutation was found in the patient's mother, who had metachronous development of lobular breast and diffuse type gastric carcinomas. Immunohistochemistry for E-cadherin protein expression revealed an abnormal staining pattern in both of these tumors, suggesting complete inactivation of the cell adhesion molecule. Thus, our finding suggests that besides diffuse type gastric cancer, lobular breast carcinomas may be associated with germline CDH1 mutations.

Identification of novel RPGR (retinitis pigmentosa GTPase regulator) mutations in a subset of X-linked retinitis pigmentosa families segregating with the RP3 locus.

The X-linked form of retinitis pigmentosa (XLRP) is a severe disease of the retina, characterised by night blindness and visual field constriction in a degenerative process, culminating with complete loss of sight within the third decade of life. Genetic mapping studies have identified two major loci for XLRP: RP3 (70%-75% of XLRP) and RP2 (20%-25% of XLRP). The RPGR (retinitis pigmentosa GTPase regulator) gene has been cloned within the RP3 genomic interval and it has been shown that 10%-20% of XLRP families have mutations in this gene. Here, we describe a single-strand conformational polymorphism-based mutation screening of RPGR in a pool of 29 XLRP families for which the disease segregates with the RP3 locus, in order to investigate the proportion of RP3 families with RPGR mutations and to relate the results to previous reports. Five different new mutations have been identified: two splice site mutations for exon 1 and three frameshift mutations in exons 7, 10 and 11. The percentage of RPGR mutations identified is 17% (5/29) in our genetically well-defined population. This figure is comparable to the percentage of RP2 gene mutations that we have detected in our entire XLRP patient pool (10%-15%). A correlation of RPGR mutations with phenotype in the families described in this study and the biochemical characterisation of reported mutations may provide insights into the function of the protein.

A novel frameshift mutation in exon 6 (the site of Asn 291) of the lipoprotein lipase gene in type I hyperlipidemia

A new heterozygous lipoprotein lipase gene defect has been identified in a type I hyperlipidemic patient at the position of notable amino acid Asn 291. The patient is a 33-year-old male. His body mass index (BMI) was 18.5 kg/m 2. The total cholesterol (TC), triglycerides (TG) and high density lipoprotein-cholesterol (HDL-C) concentration from his fasting plasma were 4.8, 11.9 and 0.4 mmol/l, respectively. The lipoprotein lipase (LPL) activity and mass in the postheparin plasma (PHP) from the patient were 0.58 mmol/ml/h (normal range: 7.7±2.6) and 244 ng/ml (normal range: 192±30), respectively. The hepatic lipase activity of the PHP from the patient was 10.6 mmol/ml/h (normal range: 9.9±3.6). DNA analysis of the LPL gene revealed that this patient had a heterozygous one nucleotide deletion of A coding Asn 291, resulting in a premature termination of the LPL protein at amino acid residue 303. The other abnormality in the LPL gene of the proband was an amino acid residue 194 defect (Ile194→Thr), which is known to cause a defective enzyme. A medium-chain triglyceride (MCT) loading test was conducted to find how this triglyceride affects plasma lipoprotein metabolism in this patient in a short term ( Fig. 3). The plasma total cholesterol (TC) or high density lipoprotein (HDL)-C levels did not change significantly after oral administration of a fatty meal containing long chain triglycerides (LCT) or MCT. The plasma TG level, on the other hand, increased from 11.9 to 19.2 mmol/l (+61%) at 6 h after loading a fatty meal containing LCT, whereas the plasma TG levels tended to even decrease at 6 h after oral administration of an MCT, tricaprin (from 11.6 to 10.5 mmol/l (−9.4%)). These results suggest that MCT, as opposed to LCT, is useful for treatment of type I hyperlipidemia with a novel mutation at the notable amino acid Asn 291 of the LPL gene.

Mutations in a new gene encoding a thiamine transporter cause thiamine-responsive megaloblastic anaemia syndrome.

Thiamine-responsive megaloblastic anaemia syndrome (TRMA; MIM 249270) is an autosomal recessive disorder with features that include megaloblastic anaemia, mild thrombocytopenia and leucopenia, sensorineural deafness and diabetes mellitus. Treatment with pharmacologic doses of thiamine ameliorates the megaloblastic anaemia and diabetes mellitus. A defect in the plasma membrane transport of thiamine has been demonstrated in erythrocytes and cultured skin fibroblasts from TRMA patients. The gene causing TRMA was assigned to 1q23.2-q23.3 by linkage analysis. Here we report the cloning of a new gene, SLC19A2, identified from high-through-put genomic sequences due to homology with SLC19A1, encoding reduced folate carrier 1 (refs 8-10). We cloned the entire coding region by screening a human fetal brain cDNA library. SLC19A2 encodes a protein (of 497 aa) predicted to have 12 transmembrane domains. We identified 2 frameshift mutations in exon 2. a 1-bp insertion and a 2-bp deletion, among four Iranian families with TRMA. The sequence homology and predicted structure of SLC19A2, as well as its role in TRMA, suggest that its gene product is a thiamine carrier, the first to be identified in complex eukaryotes.

Recurrent molecular abnormalities in type VII collagen in Southern Italian patients with recessive dystrophic epidermolysis bullosa.

In this study we searched for mutations in the type VII collagen gene (COL7A1) in 10 families from Southern Italy with severe generalised recessive dystrophic epidermolysis bullosa using PCR amplification of genomic DNA, heteroduplex analysis and direct nucleotide sequencing. Our principal aim was to identify any recurrent mutations in COL7A1 that might facilitate future mutation detection strategies in this population. Three recurrent COL7A1 mutations were delineated in six of the 10 families: a frameshift mutation in exon 4, 497insA, was detected in three affected individuals from three families, a deletion mutation at the acceptor splice site of intron 114/exon 115, 8441-14del21, was found in five patients in three of the families, and an intron 49 acceptor splice site mutation, 4783-1 G-to-A, was identified in three subjects in two families (GenBank accession no, L02870). Haplotype analyses showed evidence for propagation of common ancestral mutant COL7A1 alleles for each of these recurrent mutations. These results contribute significantly to understanding the nature of COL7A1 pathology in patients from Southern Italy and in designing future approaches to mutation detection.

Quantitative Analysis of Survival Motor Neuron Copies: Identification of Subtle SMN1 Mutations in Patients with Spinal Muscular Atrophy, Genotype-Phenotype Correlation, and Implications for Genetic Counseling

Problems with diagnosis and genetic counseling occur for patients with autosomal recessive proximal spinal muscular atrophy (SMA) who do not show the most common mutation: homozygous absence of at least exon 7 of the telomeric survival motor neuron gene (SMN1). Here we present molecular genetic data for 42 independent nondeleted SMA patients. A nonradioactive quantitative PCR test showed one SMN1 copy in 19 patients (45%). By sequencing cloned reverse-transcription (RT) PCR products or genomic fragments of SMN1, we identified nine different mutations in 18 of the 19 patients, six described for the first time: three missense mutations (Y272C, T274I, S262I), three frameshift mutations in exons 2a, 2b, and 4 (124insT, 241-242ins4, 591delA), one nonsense mutation in exon 1 (Q15X), one Alu-mediated deletion from intron 4 to intron 6, and one donor splice site mutation in intron 7 (c.922+6T-->G). The most frequent mutation, Y272C, was found in 6 (33%) of 18 patients. Each intragenic mutation found in at least two patients occurred on the same haplotype background, indicating founder mutations. Genotype-phenotype correlation allowed inference of the effect of each mutation on the function of the SMN1 protein and the role of the SMN2 copy number in modulating the SMA phenotype. In 14 of 23 SMA patients with two SMN1 copies, at least one intact SMN1 copy was sequenced, which excludes a 5q-SMA and suggests the existence of further gene(s) responsible for approximately 4%-5% of phenotypes indistinguishable from SMA. We determined the validity of the test, and we discuss its practical implications and limitations.

Global sequence diversity of BRCA2: analysis of 71 breast cancer families and 95 control individuals of worldwide populations [published erratum appears in Hum Mol Genet 1999 Apr;8(4):717-9

The aim of this study was to evaluate the prevalence of simple sequence variation in the BRCA2 gene. To this end, 71 breast and breast-ovarian cancer (HBC/HBOC) families along with 95 control individuals from a wide range of ethnicities were analyzed by means of denaturing high-performance liquid chromatography (DHPLC) and direct sequence analysis. In the coding (10 257 bp) and non-coding (2799 bp) sequences of BRCA2, 82 sequence variants were identified. Three different, apparently disease-associated BRCA2 mutations were found in six HBC/HBOC families (8%): two splice site mutations in introns 5 and 21, and one frameshift mutation in exon 11. In the coding region, 53 simple sequence variants were found: 35 missense mutations, one 2 bp deletion (CT) resulting in a stop at codon 3364, one nonsense mutation with a stop at codon 3326, one deletion of a complete codon (AAA) resulting in the loss of leucine, and 15 silent mutations. In the non-coding region, 26 polymorphisms were detected. Of the 79 sequence variants that were not obviously disease-associated, eight were detected only in HBC/HBOC families. The remaining 71 variants were identified in both HBC/HBOC families and control individuals. Sixty three sequence variants (80%) were specific for a continent. Forty two percent (33 out of 79) of the sequence variants were detected exclusively in Africa, though only 13% of the 332 chromosomes screened were of African origin. Our data indicate that, in BRCA2, simple sequence variation is frequent [in the coding region 1 in 194 bp (straight theta = 2.2 x 10(-4)), and in the non-coding region 1 in 108 bp (straight theta = 4.4 x 10(-4)), respectively].

Cutis Laxa Arising from Frameshift Mutations in Exon 30 of the Elastin Gene (ELN)

Congenital cutis laxa, a rare syndrome with marked skin laxity and pulmonary and cardiovascular compromise, is due to defective elastic fiber formation. In several cases, skin fibroblast tropoelastin production is markedly reduced yet reversed in vitro by transforming growth factor-beta treatment. We previously showed that this reversal was due to elastin mRNA stabilization in one cell strain, and here this behavior was confirmed in skin fibroblasts from two generations of a second family. cDNA sequencing and heteroduplex analysis of elastin gene transcripts from three fibroblast strains in two kindreds now identify two frameshift mutations (2012DeltaG and 2039DeltaC) in elastin gene exon 30, thus leading to missense C termini. No other mutations were present in the ELN cDNA sequences of all three affected individuals. Transcripts from both alleles in each kindred were unstable and responsive to transforming growth factor-beta. Exons 22, 23, 26A, and 32 were always absent. Since exon 30 underwent alternative splicing in fibroblasts, we speculate that a differential splicing pattern could conceivably lead to phenotypic rescue. These two dominant-acting, apparently de novo mutations in the elastin gene appear to be responsible for qualitative and quantitative defects in elastin, resulting in the cutis laxa phenotype.