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PubMed1995

The chloroplast gene encoding ribosomal protein S4 in Chlamydomonas reinhardtii spans an inverted repeat--unique sequence junction and can be mutated to suppress a streptomycin dependence mutation in ribosomal protein S12.

The ribosomal protein gene rps4 was cloned and sequenced from the chloroplast genome of Chlamydomonas reinhardtii. The N-terminal 213 amino acid residues of the S4 protein are encoded in the single-copy region (SCR) of the genome, while the C-terminal 44 amino acid residues are encoded in the inverted repeat (IR). The deduced 257 amino acid sequence of C. reinhardtii S4 is considerably longer (by 51-59 residues) than S4 proteins of other photosynthetic species and Escherichia coli, due to the presence of two internal insertions and a C-terminal extension. A short conserved C-terminal motif found in all other S4 proteins examined is missing from the C. reinhardtii protein. In E. coli, mutations in the S4 protein suppress the streptomycin-dependent (sd) phenotype of mutations in the S12 protein. Because we have been unable to identify similar S4 mutations among suppressors of an sd mutation in C. reinhardtii S12 obtained using UV mutagenesis, we made site-directed mutations [Arg68 (CGT) to Leu (CTG and CTT)] in the wild-type rps4 gene equivalent to an E. coli Gln53 to Leu ribosomal ambiguity mutation (ram), which suppresses the sd phenotype and decreases translational accuracy. These mutants were tested for their ability to transform the sd S12 mutation of C. reinhardtii to streptomycin independence. The streptomycin-independent isolates obtained by biolistic transformation all possessed the original sd mutation in rps12, but none had the expected donor Leu68 mutations in rps4. Instead, six of 15 contained a Gln73 (CAA) to Pro (CCA) mutation five amino acids downstream from the predicted mutant codon, irrespective of rps4 donor DNA. Two others contained six- and ten-amino acid, in-frame insertions at S4 positions 90 and 92 that appear to have been induced by the biolistic process itself. Eight streptomycin-independent isolates analyzed had wild-type rps4 genes and may possess mutations identical to previously isolated suppressors of sd that define at least two additional chloroplast loci. Cloned rps4 genes from streptomycin-independent isolates containing the Gln73 to Pro mutation and the 6-amino acid insertion in r-protein S4 transform the sd strain to streptomycin independence.

Amino Acid Sequence↗
PubMed1992

X-ray-induced specific-locus mutations in the ad-3 region of two-component heterokaryons of Neurospora crassa. X. Heterozygous effects of multilocus deletion mutations of genotype ad-3A or ad-3B.

Previous studies on X-ray-induced irreparable adenine-3 mutations (designated [ad-3]IR), induced in heterokaryon 12 of Neurospora crassa, demonstrated that they were not recessive and exhibited heterozygous effects in terms of markedly reduced linear growth rates (de Serres, 1965). Complementation tests with a series of tester strains carrying multilocus deletion mutations in the ad-3 and immediately adjacent genetic regions demonstrated that X-ray-induced irreparable mutations map, in the main part, as a series of overlapping multilocus deletion mutations that extend both proximally and distally into the immediately adjacent genetic regions, as well as into the 'X' region (a region of unknown, but essential function) between ad-3A and ad-3B (de Serres, 1968, 1989). Further studies (de Serres and Miller, 1988) have shown that the heterozygous effects of multilocus deletion mutations in the ad-3 region can be modified genetically and biochemically. In the present paper, the heterozygous effects of X-ray-induced multilocus deletion mutations of genotype ad-3A or ad-3B, induced in heterokaryon 12 (Webber and de Serres, 1965; de Serres, 1988, 1989), have been determined. These data show that 57.7% (15/26) of X-ray-induced multilocus deletion mutations covering the ad-3A locus have heterozygous effects, in terms of reduced linear growth rates, in forced dikaryons with a gene/point mutant at the ad-3B locus and 80.0% (20/25) in forced dikaryons with a multilocus deletion mutation covering the ad-3B locus. In addition, 35.1% (20/57) of X-ray-induced multilocus deletion mutations covering the ad-3B locus have heterozygous effects in forced dikaryons with a gene/point mutant at the ad-3A locus, and 100.0% (35/35) in forced dikaryons with a multilocus deletion mutation covering the ad-3A locus. These results demonstrate that the dominant or recessive characteristics of X-ray-induced specific-locus mutations resulting from multilocus deletion mutations are allele specific.

Adenine↗
PubMed1998

Mutations of the human tyrosinase gene associated with tyrosinase related oculocutaneous albinism (OCA1). Mutations in brief no. 204. Online.

Mutations in the human tyrosinase gene produce tyrosinase-related oculocutaneous albinism (OCA1, MIM #203100). Tyrosinase is a copper containing enzyme and is responsible for catalyzing the rate limiting step in melanin biosynthesis, the hydroxylation of tyrosine to dopaquinone. We report 13 new mutations in the tyrosinase gene associated with OCA1A (without pigment) and OCA1B (with pigment) including 9 missense mutations (H19Q, R521, R77C, G97R, C289R, L312V, P313R, F340L and H404P), two nonsense mutations (W80X and R116X) and two frameshift mutations (53delG and 223 delG). Our previous work has defined clusters of missense mutations that appear to represent functional domains of the enzyme, and three of the missense mutations fall into these clusters including two (F340L and H404P) that flank the copper B bindng site and the missense mutation R52I that is located in the amino terminal end cluster of the protein. The G97R missense mutation is the first identified within the epidermal growth factor (EGF)-like sequence and the H19Q missense mutation alters the cleavage site of the signal peptide sequence. Mutational analysis can provide a definitive diagnosis of the type of OCA as well as help structure/function analysis.

Albinism, Oculocutaneous↗
PubMed2005

Mutation rates in the dystrophin gene: a hotspot of mutation at a CpG dinucleotide.

An analysis of mutations was performed in 141 Duchenne muscular dystrophy (DMD) patients previously found to be negative for large deletions by standard multiplex PCR assays. Comprehensive mutation scanning of all coding exons, adjacent intronic splice regions, and promoter sequences was performed by DOVAM-S, a robotically enhanced, high throughput method that detects essentially all point mutations. Samples negative for point mutations were further analyzed for duplications by multiplex amplifiable probe hybridization (MAPH). Presumptive causative mutations were detected in 90% of the patients (70% protein truncating point mutations, 13% duplications, and 7% deletions not detected by the standard multiplex screening method). A total of 40 of the mutations are putatively novel. Most duplications involve multiple exons with an average and median size of about 160 and 153 kb, respectively. This is the first analysis of the absolute and relative rates of point mutations in the dystrophin gene. Relative to microdeletions (0.68 x 10(-9) per bp per generation), transitions at CpG dinucleotides are enhanced 150-fold while complex indels, the least common mutation type, are less frequent than microdeletions by a factor of five. The frequency of microdeletions and microinsertions at mononucleotide repeats increases exponentially with length. When compared to the well-studied human factor IX gene (F9), the results are similar, with two exceptions: a hotspot of mutation in the dystrophin gene (c.8713C>T/p.R2905X) at a CpG dinucleotide and an altered size distribution of microdeletions. The hotspot reflects a difference in the underlying pattern of mutation, while the altered size distribution of microdeletions reflects certain abundant sequence motifs within the dystrophin coding sequence (relative to factor IX).

Adolescent↗
PubMed2002

Ten novel ORF15 mutations confirm mutational hot spot in the RPGR gene in European patients with X-linked retinitis pigmentosa.

RGPR was the first gene found to be mutated in XLRP, the subtype of RP displaying the most severe form of retinal degeneration with partial or complete blindness in the third or fourth decade of life. Despite the RP3 locus on Xp21.1 accounting for 60-90% of XLRP, only 10-20% of identified RPGR mutations were reported in earlier analyses. This discrepancy appeared to be resolved when Vervoort et al. identified a mutational hot spot in a new purine-rich 3' exon (ORF15) that accounted for 60% of their XLRP patients [Vervoort et al., 2000]. In our mutation screening of 37 unrelated European XLRP patients we identified two recently described deletions and 10 novel mutations in exon ORF15 of RPGR, 4 of which were nonsense and 6 frameshift mutations. The latter included one duplication and 5 deletion mutations, all of which lead to a downstream premature termination. No mutations were detected in the additionally screened new exon ORF14. The data reported here, together with previous findings, document a significant clustering of mutations as well as polymorphisms in ORF15 of RPGR. In our unselected XLRP patient population, ORF15 mutations constitute 32% of cases, a finding that contradicts the results of Vervoort and coworkers [Vervoort et al., 2000] but is in agreement with a more recent study on North American XLRP patients [Breuer et al., 2002]. The observed prevalence is sufficient to justify an initial mutation screening of ORF15 in the genetically heterogeneous group of XLRP.

Carrier Proteins↗
PubMed2002

Mutation screening of the fibrillin-1 (FBN1) gene in 76 unrelated patients with Marfan syndrome or Marfanoid features leads to the identification of 11 novel and three previously reported mutations.

Mutations in the gene encoding fibrillin-1 (FBN1) cause Marfan syndrome (MFS) and other related connective tissue disorders. In this study we performed SSCP to analyze all 65 exons of the FBN1 gene in 76 patients presenting with classical MFS or related phenotypes. We report 7 missense mutations, 3 splice site alterations, one indel mutation, one nonsense mutation and two mutations causing frameshifts: a 16bp deletion and a single nucleotide insertion. 5 of the missense mutations (Y1101C, C1806Y, T1908I, G1919D, C2251R) occur in calcium-binding Epidermal Growth Factor-like (EGFcb) domains of exons 26, 43, 46 and 55, respectively. One missense mutation (V449I) substitutes a valine residue in the non-calcium-binding epidermal growth factor like domain (EGFncb) of exon 11. One missense mutation (G880S) affects the "hybrid" motif in exon 21 by replacing glycine to serine. The 3 splice site mutations detected are: IVS1-1G>A in intron 1, IVS38-1G>A in intron 38 and IVS46+5G>A in intron 46. C628delinsK was identified in exon 15 leading to the substitution of a conserved cysteine residue. Furthermore two frameshift mutations were found in exon 15 (1904-1919del ) and exon 63 (8025insC) leading to premature termination codons (PTCs) in exon 17 and 64 respectively. Finally we identified a nonsense mutation (R429X) located in the proline rich domain in exon 10 of the FBN1 gene. Y1101C, IVS46+5G>A and R429X have been reported before.

Adolescent↗
PubMed1995

Characteristics of UV-induced mutation spectra in human XP-D/ERCC2 gene-mutated xeroderma pigmentosum and trichothiodystrophy cells.

To study the relationships between mutagenesis and carcinogenesis, we compared the mutations and their frequency induced by ultraviolet irradiation at 254 nm (UV-C) in XP-D (GM-08207B/XP6BE), TTD/XP-D (TTD1VI-LAS-KMT11) and wild-type (MRC-5V1) human cells. XP-D and TTD/XP-D cells, mutated in the same XP-D/ERCC2 gene, are deficient in nucleotide excision repair. Whereas XP-D patients develop early skin tumors, TTD patients do not exhibit abnormal levels of cancers. After verification of UV hypersensitivity and DNA repair defect of the immortalized cell lines XP-D and TTD compared with a wild-type cell line, UV-induced mutagenesis was studied with a new shuttle vector pR2, carrying the target lacZ' gene. The UV-mutation frequencies in XP-D and TTD cells were similar and significantly increased compared with normal cells. Sequence analysis of 312 independent mutant plasmids revealed that more rearrangements were induced in TTD cells (16%) than in XP-D (5%) and normal cells (1%), while XP-D cells exhibited a twofold higher rate of tandem mutations compared with TTD and normal cells. In the three cell lines, a predominance of G:C to A:T transitions was found, especially in chiefly on the cytosine at 5'-TC-3' sites. The types of UV-induced point mutations in TTD cells were, however, more similar to those in normal cells than those found in XP-D cells. XP-D mutations were preferentially located in 5'-TCPur-3' sites, while mutations in normal and TTD cells were mostly at 5'-TCC-3' sites. Analysis of mutation spectra revealed differences in the location of the mutational hotspots between the three lines. Although the mutation frequency of the UV-irradiated pR2 vector is much higher in TTD and XP-D cells than in normal cells, the mutation spectrum is closer between TTD and normal cells as compared with XP-D cells. These dissimilarities could contribute to an explanation of some of the differences between the two syndromes.

Amino Acid Sequence↗
PubMed2004

Mutational biases associated with potential iron-binding DNA motifs in rodent lacI and human p53 mutational databases.

The role of Fenton oxidants in DNA damage, aging, and cancer is appreciated, but not well understood. Six potential iron-binding (PIB) DNA motifs were previously identified as sites of preferential strand cleavage. Since DNA-metal binding domains are a known determinant of oxidative DNA damage, and the location of strand breaks explains where oxidant attack occurs, we sought to determine whether the likelihood of base change mutations is a function of neighboring PIB motifs. We developed a sliding window function that computes the density of PIB motifs on both strands, within 4-12bp, for each location along a target gene. This range of window sizes reflects known diffusion distances of Fenton reaction products. Using mutational databases, odds of mutation at each base were calculated relative to PIB motif density, for all PIB motif types in aggregate, or for individual PIB motifs. Using mutational data from lacI transgenic animals, we observed a non-random distribution of PIB motifs, associated with increased odds of mutation, showing a strand bias. Sensitivity analysis confirmed that the optimum association between PIB motif density and mutations occurs when a 7bp radius is used for the window size. Randomly simulated mutations showed no association with PIB motif density. When the method was applied to human TP53 mutation data, we saw similar results, but no strand bias. As PIB motif density rises, linear trends are observed for increasing odds of mutation. Sensitivity analysis revealed associations between PIB motifs and GC --> AT transitions and GC --> TA transversions-the most commonly observed types of mutations arising from oxidative DNA damage. DNA-metal binding motifs are found in a wide variety of biological contexts, including many where conformational sensitivity to redox state is important. These techniques can help elucidate how DNA-iron-binding may affect lesions and subsequent mutations from multiple agents.

Amino Acid Motifs↗
PubMed2002

Mutation cluster region, association between germline and somatic mutations and genotype-phenotype correlation in upper gastrointestinal familial adenomatous polyposis.

Studies of adenomatous polyposis coli (APC) mutations in familial adenomatous polyposis (FAP) have focused on large bowel disease. It has been found that: 1) germline APC mutations around codon 1300 are associated with severe colorectal polyposis; 2) somatic APC mutations in colorectal tumors tend to cluster approximately between codons 1250 and 1450; and 3) patients with germline mutations close to codon 1300 tend to acquire somatic mutations (second hits) in their colorectal polyps by allelic loss, whereas the tumors of other FAP patients have truncating second hits. Using new and published data, we have investigated how germline and somatic APC mutations influence the pathogenesis of upper gastrointestinal polyps in FAP. We have compared the results with those from colorectal disease. We found that somatic mutations in upper gastrointestinal polyps cluster approximately between codons 1400 and 1580. Patients with germline APC mutations after codon 1400 tend to show allelic loss in their upper gastrointestinal polyps; the tumors of other patients have truncating somatic mutations after codon 1400. Finally, patients with germline mutations after codon 1400 tend to have more severe duodenal polyposis (odds ratio, 5.72; 95% confidence interval, 1.13 to 28.89; P = 0.035). Thus, in both upper gastrointestinal and colorectal tumors, a specific region of the APC gene is associated with severe disease, clustering of somatic mutations, and loss of the wild-type allele. However, the region concerned is different in upper gastrointestinal and colorectal disease. The data suggest that loss of all APC SAMP repeats is probably necessary for duodenal and gastric tumorigenesis in FAP, as it is in colonic tumors. Compared with colonic tumors, however, retention of a greater number of beta-catenin binding/degradation repeats is optimal for tumorigenesis in upper gastrointestinal FAP.

Adenomatous Polyposis Coli↗
PubMed1997

Spontaneous mutation frequencies and spectra in p53 (+/+) and p53 (-/-) mice: a test of the 'guardian of the genome' hypothesis in the Big Blue transgenic mouse mutation detection system.

TSG-p53/Big Blue double transgenic mice offer a powerful tool for examining the effect of a p53 germline mutation on spontaneous somatic mutation in vivo. After sequencing the DNA-binding domain of the lacI gene, we previously reported no differences in mutant frequency between p53 nullizygous (-/-) and p53 wild-type (+/+) mice in liver, spleen and brain. However, jackpot mutations elsewhere in the gene may have obscured a real difference in mutation frequency and the small sample size of mutations not at CpG dinucleotides (n = 23) may have been insufficient to reveal differences in mutation spectra. Herein we have sequenced the entire lacI gene, including the promoter and lacZ operator regions. 123 additional independent mutations have been found including 70 mutations not at CpG sites. The mutation frequency was determined by correcting for jackpot mutations. There were no statistically significant differences in mutation frequency or spectrum between the p53 (+/+) and p53 (-/-) genotypes in any of the three tissues.

Animals↗
PubMed1999

Analysis of the hereditary pancreatitis-associated cationic trypsinogen gene mutations in exons 2 and 3 by enzymatic mutation detection from a single 2.2-kb polymerase chain reaction product.

BACKGROUND: Hereditary pancreatitis is associated with at least 2 mutations in the cationic trypsinogen gene. The purpose of the present study is to test the utility of T4 endonuclease VII for the detection of cationic trypsinogen R117H mutations. In addition, the possibility of screening for R117H, N21I, and A8V mutations in a single 2.2-kb polymerase chain reaction (PCR) product using T4 endonuclease VII was investigated. METHODS: Twenty-nine DNAs from control patients and patients with known cationic trypsinogen R117H, A8V, or N21I mutations were selected from the ongoing hereditary pancreatitis study of the Midwest Multicenter Pancreatic Study Group. The samples were coded and randomized, and a 911-bp sequence containing exon 3 or a 2,212- bp sequence containing exons 2 and 3 were amplified by PCR using fluorescent- labeled primers. The PCR products were digested with T4 endonuclease VII and screened for mutations on an automated DNA sequencer. RESULTS: In all cases with a mutation, a cleavage fragment on the direct and/or complementary DNA strand could easily be visualized, and its approximate size correlated with the predicted location of the known mutations within the PCR product. When the code for affected status was broken, there was 100% correlation between previous DNA sequence or restriction fragment length polymorphism findings and the T4 endonuclease VII digestion results for all 29 DNAs. CONCLUSION: T4 endonuclease VII accurately identified the known cationic trypsinogen gene mutations in exons 2 and 3. Enzymatic mutation detection appears to be an accurate and useful method for screening individuals for known trypsinogen gene mutations and may be useful in identifying previously unidentified mutations within large regions of interest.

Acute Disease↗
PubMed2005

Mutations in the known genes are not the major cause of MED; distinctive phenotypic entities among patients with no identified mutations.

Multiple epiphyseal dysplasia (MED) is a clinically and genetically heterogeneous chondrodysplasia. Mutations in six genes (COMP, COL9A1, COL9A2, COL9A3, MATN3 and DTDST) have been reported, but the genotype-phenotype correlations and the proportions of cases due to mutations in these genes are still not well characterized. We performed a clinical, radiological and molecular analysis of known MED genes on 29 consecutive MED patients. The mutation analysis resulted in identification of the DTDST mutation in four patients (14%), the COMP mutation in three (10%) and the MATN3 mutation in three (10%). Thus, a disease-causing mutation was identified in 10 patients altogether (34%). The phenotypic features observed in the patients with mutations were in accordance with previously described phenotypes, but two new distinct phenotypic entities were identified in patients in whom no mutation was found. One of them was characterized by severe, early-onset dysplasia of the proximal femurs with almost complete absence of the secondary ossification centres and abnormal development of the femoral necks. The other phenotype was characterized by 'mini-epiphyses', resulting in severe dysplasia of the proximal femoral heads. The findings suggest that mutations in the known genes are not the major cause of MED and are responsible for less than half of the cases. The existence of additional MED loci is supported by the exclusion of known loci by mutation analysis and finding of specific subgroups among these patients.

Adolescent↗
PubMed2006

Pathogenicity of MSH2 missense mutations is typically associated with impaired repair capability of the mutated protein.

BACKGROUND & AIMS: Inherited deleterious mutations in mismatch repair genes MLH1, MSH2, and MSH6 predispose to hereditary nonpolyposis colorectal cancer. A major diagnostic challenge is the difficulty in evaluating the pathogenicity of missense mutations. Previously we showed that most missense variants in MSH6 do not impair MMR capability and are associated with no or low cancer susceptibility, whereas in MLH1, functional studies distinguished nontruncating mutations with severe defects from those not or slightly impaired in protein expression or function. The present study was undertaken to evaluate the pathogenicity of inherited missense mutations in MSH2. METHODS: Fifteen mutated MSH2 proteins including 14 amino acid substitutions and one in-frame deletion were tested for expression/stability, MSH2/MSH6 interaction, and repair efficiency. The genetic and biochemical data were correlated with the clinical data. Comparative sequence analysis was performed to assess the value of sequence homology as a tool for predicting functional results. RESULTS: None of the studied MSH2 mutations destroyed the protein or abolished MSH2/MSH6 interaction, whereas 12 mutations impaired the repair capability of the protein. Comparative sequence analysis correctly predicted functional studies for 13 of 14 amino acid substitutions. CONCLUSIONS: Interpretation was pathogenic for 12, nonpathogenic for 2, and contradictory for 1 mutation. The pathogenicity could not be distinguished unambiguously by phenotypic characteristics, although correlation between the absence of staining for MSH2 and pathogenicity of the missense mutation was notable. Unlike in MSH6 and MLH1, the pathogenicity of missense mutations in MSH2 was always associated with impaired repair capability of the mutated protein.

Adaptor Proteins, Signal Transducing↗
PubMed2001

Mutations as missing data: inferences on the ages and distributions of nonsynonymous and synonymous mutations.

This article describes a new Markov chain Monte Carlo (MCMC) method applicable to DNA sequence data, which treats mutations in the genealogy as missing data. The method facilitates inferences regarding the age and identity of specific mutations while taking the full complexities of the mutational process in DNA sequences into account. We demonstrate the utility of the method in three applications. First, we demonstrate how the method can be used to make inferences regarding population genetical parameters such as theta (the effective population size times the mutation rate). Second, we show how the method can be used to estimate the ages of mutations in finite sites models and for making inferences regarding the distribution and ages of nonsynonymous and synonymous mutations. The method is applied to two previously published data sets and we demonstrate that in one of the data sets the average age of nonsynonymous mutations is significantly lower than the average age of synonymous mutations, suggesting the presence of slightly deleterious mutations. Third, we demonstrate how the method in general can be used to evaluate the posterior distribution of a function of a mapping of mutations on a gene genealogy. This application is useful for evaluating the uncertainty associated with methods that rely on mapping mutations on a phylogeny or a gene genealogy.

Algorithms↗
PubMed1996

Loss or somatic mutations of hMSH2 occur in hereditary nonpolyposis colorectal cancers with hMSH2 germline mutations.

Hereditary nonpolyposis colorectal cancer (HNPCC) is a major cancer susceptibility syndrome known to be caused by the inheritance of mutations in DNA mismatch repair genes, such as hMSH2, hMLH1, hPMS1 and hPMS2. To investigate the role of genetic alterations of hMSH2 in HNPCC tumorigenesis, we analyzed 36 Japanese HNPCC kindreds as to hMSH2 germline mutations. Moreover, we also examined somatic mutations of hMSH2 or loss of heterozygosity at or near the hMSH2 locus in the tumors from the hMSH2-related kindreds. Germline mutations were detected in five HNPCC kindreds (5/36, 14%). Among them, three were nonsense mutations, one was a frameshift mutation and the other was a mutation in an intron where the mutation affected splicing. Loss of heterozygosity in four and somatic mutations in one were detected among the eight tumors with hMSH2 germline mutations. All these alterations were only detected in genomic instability(+) tumors, i.e., not in genomic instability(-) ones, indicating that mutations of hMSH2 were responsible for at least some of the tumors with genomic instability. These data establish a basis for the presymptomatic diagnosis of HNPCC patients, and constitute further evidence that both DNA mismatch repair genes and tumor suppressor genes may share the same requirement, i.e., two hits are necessary to inactivate the gene function.

Alleles↗
PubMed2003

Expanding the phenotype of LMNA mutations in dilated cardiomyopathy and functional consequences of these mutations.

AIMS: Mutations in the lamin A/C gene (LMNA) have been reported to be involved in dilated cardiomyopathy (DCM) associated with conduction system disease and/or skeletal myopathy. The aim of this study was to perform a mutational analysis of LMNA in a large white population of patients affected by dilated cardiomyopathy with or without associated symptoms. METHODS: We performed screening of the coding sequence of LMNA on DNA samples from 66 index cases, and carried out cell transfection experiments to examine the functional consequences of the mutations identified. RESULTS: A new missense (E161K) mutation was identified in a family with early atrial fibrillation and a previously described (R377H) mutation in another family with a quadriceps myopathy associated with DCM. A new mutation (28insA) leading to a premature stop codon was identified in a family affected by DCM with conduction defects. No mutation in LMNA was found in cases with isolated dilated cardiomyopathy. Functional analyses have identified potential physiopathological mechanisms involving identified mutations, such as haploinsufficiency (28insA) or intermediate filament disorganisation (E161K, R377H). CONCLUSION: For the first time, a specific phenotype characterised by early atrial fibrillation is associated with LMNA mutation. Conversely, mutations in LMNA appear as a rare cause of isolated dilated cardiomyopathy. The variable phenotypes observed in LMNA-DCM might be explained by the variability of functional consequences of LMNA mutations.

Adolescent↗
PubMed2006

Optimization of patient selection for gefitinib in non-small cell lung cancer by combined analysis of epidermal growth factor receptor mutation, K-ras mutation, and Akt phosphorylation.

PURPOSE: Mutations in epidermal growth factor receptor (EGFR) are strongly predictive of gefitinib efficacy in non-small-cell lung cancer. However, the presence of EGFR mutant nonresponses and nonmutant responses points out the need for more comprehensive analysis. PATIENTS AND METHODS: For 69 non-small-cell lung cancer patients treated with gefitinib, we have extended our analysis to EGFR gene copy number by fluorescence in situ hybridization, mutations in K-ras, HER2, and exon 20 of EGFR by direct sequencing, and phosphatase and tensin homologue expression by immunohistochemistry, in addition to EGFR exons 18, 19, and 21, and phosphorylations of Akt and extracellular signal-regulated kinase reported previously. RESULTS: EGFR mutation and high gene copy number were associated with better objective response in univariate analysis. However, only gefitinib-sensitive EGFR mutation was independently predictive of both response (P = 0.011) and survival (P = 0.002) in multivariate analysis. No patients with K-ras mutation, including two EGFR mutants, showed response. In EGFR nonmutants, patients with either K-ras mutation or p-Akt overexpression exhibited poor response and time-to-progression whereas patients with high gene copy number tended to have better outcomes in univariate analysis. In multivariate analysis of time-to-progression in EGFR nonmutants, K-ras mutation or p-Akt overexpression was associated with shorter time-to-progression (P = 0.017). No patient with HER2 mutation showed response to gefitinib. Reduced phosphatase and tensin homologue expression was not associated with gefitinib sensitivity. CONCLUSION: Gefitinib-sensitive EGFR mutation is the single most important predictor of gefitinib sensitivity. In addition to EGFR mutation, K-ras mutation and Akt phosphorylation aid in better prediction of gefitinib responsiveness in non-small-cell lung cancer.

Adult↗
PubMed1999

H-ras and K-ras gene mutations in primary human soft tissue sarcoma: concomitant mutations of the ras genes.

ras gene mutations have been described with varying frequency in several types of human malignancies. To determine the incidence and type of ras mutations in human soft tissue tumors, we studied 45 sarcomas, including 27 malignant fibrous histiocytomas (MFHs), 10 liposarcomas, 2 rhabdomyosarcomas, and 6 leiomyosarcomas. Al of the tumors were investigated by direct sequence analysis with the automated DNA sequencing of polymerase chain reaction-amplified ras sequences. Twenty (44%) of the sarcomas examined harbored K-ras mutations, 18 (90%) of which were MFHs. All of the K-ras mutations were G-to-A transition mutations in the second position of codon 13 (glycine --> aspartic acid). Of the samples with K-ras activation, 7 (16% of the total of 45 tumors), including 6 MFHs and 1 leiomyosarcoma, also contained H-ras mutation. All of the tumors that showed H-ras alteration had G-to-T transversion mutations in the second base of codon 12 (glycine --> valine). These data possibly implicate that ras gene activation may be a relatively uncommon event in soft tissue tumors, with the exception of MFH. It is suggested that the oncogenic process underlying the development of tumors between these groups may be different and that ras gene mutations may play a role in the etiology and/or progression of MFH. It is noteworthy that when ras gene activation occurs in sarcoma, it predominantly affects the K-ras gene, particularly codon 13. Moreover, H-ras mutations in our samples were detected only in association with tumors that also displayed K-ras-mutated genes. This study demonstrates for the first time concomitant mutations in two different members of the ras gene family in sarcoma

Adolescent↗