RESUMO
BACKGROUND: Mutations in the periaxin (PRX) gene cause autosomal recessive demyelinating neuropathy Charcot-Marie-Tooth (CMT) type 4F. To date, 10 non-sense or frameshift PRX mutations have been reported in patients with early-onset neuropathy and further disease course consistent with either Dejerine-Sottas neuropathy or slow-progressive demyelinating CMT. METHODS: We sequenced 59 patients from 55 Czech families including four unrelated patients of Romani (Gypsy) origin with early-onset CMT displaying decreased nerve conduction velocities. RESULTS: We identified a novel homozygous mutation c.3286_3356del71 (K1095fsX18) in one Romani patient showing very slow disease progression. Amongst non-Romani Czech CMT patients, PRX mutations have been proven to be very rare.
Assuntos
Doença de Charcot-Marie-Tooth/genética , Doença de Charcot-Marie-Tooth/fisiopatologia , Doenças Desmielinizantes/genética , Proteínas de Membrana/genética , Roma (Grupo Étnico)/genética , Deleção de Sequência/genética , Idade de Início , Sequência de Bases , Pré-Escolar , Feminino , Testes Genéticos , Humanos , Lactente , Masculino , LinhagemRESUMO
Mutations in the early growth response 2 gene (EGR2) cause demyelinating neuropathies differing in severity and age of onset. We tested 46 unrelated Czech patients with dominant or sporadic demyelinating CMT neuropathy for mutations in the EGR2 gene. One novel de-novo mutation (Arg359Gln, R359Q) was identified in heterozygous state in a patient with a typical CMT1 phenotype, progressive moderate thoracolumbar scoliosis and without clinical signs of cranial nerve dysfunction. This patient is presently less affected compared to previously described Dejerine-Sottas neuropathy (DSN) patients carrying another substitution at codon 359 (Arg359Trp, R359W). This report shows that EGR2 mutations are rare in Czech patients with demyelinating type of CMT and suggests that different substitutions at codon 359 of EGR2 can cause significantly different phenotypes.
Assuntos
Arginina/genética , Doença de Charcot-Marie-Tooth/genética , Proteínas de Ligação a DNA/genética , Glutamina/genética , Mutação , Escoliose/genética , Transativadores/genética , Adolescente , Doença de Charcot-Marie-Tooth/complicações , Doença de Charcot-Marie-Tooth/fisiopatologia , Doença de Charcot-Marie-Tooth/radioterapia , Análise Mutacional de DNA/métodos , Feminino , Humanos , Masculino , Condução Nervosa/fisiologia , Exame Neurológico , Nervos Periféricos/fisiopatologia , Radiografia , Escoliose/complicações , Escoliose/diagnóstico por imagem , Escoliose/fisiopatologia , Regulador Transcricional ERGRESUMO
Exaggerated neurogenic inflammation has been recognized to be one reason for many CRPS symptoms. Since angiotensin-converting enzyme (ACE) is a key enzyme for the termination of neurogenic inflammation, it has been selected as a candidate gene for CRPS predisposition. A previous report of an insertion/deletion (I/D) polymorphism in intron 16 within the ACE gene implicated an increased risk to develop CRPS I associated with the D allele. However, in the present study the D allele frequency was not increased in CRPS I cases (0.51 for D allele, 0.49 for I allele). Furthermore, there was no co-segregation of any genotype (DD, ID, II) with the CRPS phenotype in 12 selected familial CRPS I cases from six CRPS I families. In conclusion, the results presented herein render this particular ACE gene polymorphism unlikely to be a predisposing factor for CRPS I.
Assuntos
Predisposição Genética para Doença/genética , Peptidil Dipeptidase A/genética , Polimorfismo Genético/genética , Distrofia Simpática Reflexa/enzimologia , Distrofia Simpática Reflexa/genética , Análise Mutacional de DNA , Feminino , Frequência do Gene , Testes Genéticos , Genótipo , Humanos , Íntrons/genética , Masculino , Mutação/genética , Neuropeptídeos , Linhagem , Fenótipo , Distrofia Simpática Reflexa/diagnósticoRESUMO
Previously we reported the amplification of the peripheral myelin protein 22 (PMP22) gene in cell lines of human osteogenic and glioma tumors. PMP22 normally is expressed at high levels in Schwann cells of the peripheral nervous system and is suggested to function as a structural protein of the myelin sheath. One of the most common inherited peripheral neuropathies, Charcot-Marie-Tooth Type 1A (CMT1A), is associated with a duplication of a 1.5-Mb DNA region on chromosome 17p11.2 - p12 containing PMP22. On the other hand, PMP22 is identical to gas3, whose expression is induced in growth-arresting NIH3T3-fibroblasts and is thought to play a role in cell proliferation. The precise role of gas3/PMP22 remains to be determined. Here we show that in the tumor cell lines RH30 and SF763 the amplified region including PMP22 comprises the whole 1.5-Mb CMT1A region. We could prove expression of PMP22 by reverse transcriptase-polymerase chain reaction (RT-PCR) and discovered an unusual PMP22 transcript in these tumor cell lines. Western blot analyses resulted in detection of a 22-kDa protein by the PMP22-specific antibody 558/2 and in exclusion of myelin protein zero (MPZ) expression in these cell lines.
Assuntos
Neoplasias Ósseas/genética , Neoplasias Encefálicas/genética , Regulação Neoplásica da Expressão Gênica/genética , Glioma/genética , Proteínas da Mielina/genética , Osteossarcoma/genética , Sequência de Aminoácidos , Testes de Carcinogenicidade , Humanos , Hibridização In Situ , Dados de Sequência Molecular , RNA Mensageiro/análise , Células Tumorais CultivadasRESUMO
Elongation of long-chain fatty acids was investigated in yeast mutants lacking endogenous de novo fatty acid synthesis. In this background, in vitro fatty acid elongation was dependent strictly on the substrates malonyl-CoA, NADPH and a medium-chain or long-chain acyl-CoA primer of 10 or more carbon atoms. Maximal activity was observed with primers containing 12-14 carbon atoms, while shorter-chain-length acyl-CoA were almost (octanoyl-CoA) or completely (hexanoyl-CoA, acetyl-CoA) inactive. In particular, acetyl-CoA was inactive as a primer and as extender unit. The Michaelis constants for octanoyl-CoA (0.33 mM), decanoyl-CoA (0.83 mM) lauroyl-CoA (0.05 mM), myristoyl-CoA (0.4 mM) and palmitoyl-CoA (0.13 mM) were determined and were comparable for fatty acid synthesis and elongation. In contrast, the affinity of malonyl-CoA was 17-fold lower for elongation (Km = 0.13 mM) than for the fatty acid synthase (FAS) system. With increasing chain length of the primer (> or = 12:0), fatty acid elongation becomes increasingly sensitive to substrate inhibition. Due to the activation of endogenous fatty acids, ATP exhibits a stimulatory effect at suboptimal but not at saturating substrate concentrations. In the yeast cell homogenate, the specific activity of fatty acid elongation is about 10-20-fold lower than that of de novo fatty acid synthesis. The same elongation activity is observed in respiratory competent and in mitochondrially defective cells. The products of in vitro fatty acid elongation are fatty acids of 15-17 or 22-26 carbon atoms, depending on whether tridecanoyl-CoA or stearoyl-CoA is used as a primer. In vitro, the elongation products are converted in part, by alpha-oxidation, to their odd-chain-length lower homologues or are hydrolyzed to fatty acids. In contrast, no odd-chain-length elongation products or very-long-chain fatty acids (VLCFA) shorter than 26:0 are observed in vivo. Hence, VLCFA synthesis exhibits a higher processivity in vivo than in the cell homogenate. In addition, the in vivo process appears to be protected against side reactions such as hydrolysis or alpha-oxidation. Yeast mutants defective in 12:0 or 13:0 elongation were derived from fas-mutant strains according to their failure to grow on 13:0-supplemented media. In vivo, 12:0 elongation was reduced to 0-10% of the normal level, while 16:0 elongation and VLCFA synthesis were unimpaired. It is concluded that yeast contains either two different elongation systems, or that the respective mutation interferes differentially with medium-chain and long-chain fatty acid elongation. The yeast gene affected in the elongation-defective mutants was isolated and, upon sequencing, identified as the known ELO1 sequence. It encodes a putative membrane protein of 32-kDa molecular mass with no obvious similarity to any of the known FAS component enzymes.
Assuntos
Aciltransferases/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Acil Coenzima A/metabolismo , Proteína de Transporte de Acila S-Maloniltransferase , Aciltransferases/genética , Clonagem Molecular , Ácidos Graxos não Esterificados/biossíntese , Genes Fúngicos , Teste de Complementação Genética , Genótipo , Cinética , Saccharomyces cerevisiae/crescimento & desenvolvimento , Especificidade por SubstratoRESUMO
Sakacin 674, a bacteriocin produced by Lactobacillus sake Lb764 and which inhibits the growth of Listeria monocytogenes, was purified to homogeneity by ammonium sulphate precipitation and sequential ion exchange, hydrophobic interaction and reversed phase chromatography. The complete amino acid sequence of sakacin 674 was determined by Edman degradation. The bacteriocin consisted of 43 amino acid residues and had a calculated molecular mass of 4436.6 Da, which is in good agreement with the molecular mass of 4437.2 as determined by mass spectrometry. The structural gene encoding sakacin 674 (sakR) was located on the chromosome. This gene was cloned and sequenced. It encoded a primary translation product of 61 amino acid residues which was cleaved between amino acids 18 and 19 to yield the active sakacin 674. Sakacin 674 resembled other known bacteriocins and was very similar to sakacin P.