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C Substitution (c + substitution)
Selected AbstractsThe evolutionarily conserved residue A653 plays a key role in HERG channel closingTHE JOURNAL OF PHYSIOLOGY, Issue 11 2009Svetlana Z. Stepanovic Human ether-a-go-go- related gene (HERG) encodes the rapid, outwardly rectifying K+ current IKr that is critical for repolarization of the cardiac action potential. Congenital HERG mutations or unintended pharmaceutical block of IKr can lead to life-threatening arrhythmias. Here, we assess the functional role of the alanine at position 653 (HERG-A653) that is highly conserved among evolutionarily divergent K+ channels. HERG-A653 is close to the ,glycine hinge' implicated in K+ channel opening, and is flanked by tyrosine 652 and phenylalanine 656, which contribute to the drug binding site. We substituted an array of seven (I, C, S, G, Y, V and T) amino acids at position 653 and expressed individual variants in heterologous systems to assess changes in gating and drug binding. Substitution of A653 resulted in negative shifts of the V1/2 of activation ranging from ,23.6 (A653S) to ,62.5 (A653V) compared to ,11.2 mV for wild-type (WT). Deactivation was also drastically altered: channels with A653I/C substitutions exhibited delayed deactivation in response to test potentials above the activation threshold, while A653S/G/Y/V/T failed to deactivate under those conditions and required hyperpolarization and prolonged holding potentials at ,130 mV. While A653S/G/T/Y variants showed decreased sensitivity to the IKr inhibitor dofetilide, these changes could not be correlated with defects in channel closure. Homology modelling suggests that in the closed state, A653 forms tight contacts with several residues from the neighbouring subunit in the tetramer, playing a key role in S6 helix packing at the narrowest part of the vestibule. Our study suggests that A653 plays an important functional role in the outwardly rectifying gating behaviour of HERG, supporting channel closure at membrane potentials negative to the channel activation threshold. [source] G-substrate gene promoter SNP (,1323T>C) modifies plasma total cholesterol and triglyceride phenotype in familial hypercholesterolemia: Intra-familial association study in an eight-generation hyperlipidemic kindredGERIATRICS & GERONTOLOGY INTERNATIONAL, Issue 2 2004Yukiko Nobe Background: Plasma lipid and lipoprotein generally reflect the complex influences of multiple genetic loci, for instance, even familial hypercholesterolemia (FH), a representative example of monogenic hyperlipidemia, often presents with phenotypic heterogeneity. Methods: In the course of investigating familial coronary artery disease in Utah, we studied 160 members of an eight-generation extended family of FH, to examine possible genetic modification of lipoprotein phenotype by ,modifier locus'. G-substrate (GSBS) is an endogenous substrate for cGMP-dependent protein kinase. We carried out an intrafamilial correlation analysis of modifier effect of ,1323T>C substitution in the GSBS gene among 85 LDLR-mutation carriers and 75 non-carriers. Results: In the LDLR - mutation carriers, the plasma cholesterol levels were highest among ,1323C homozygotes (mean ± SD = 454 ± 101 mg/dL), lowest among ,1323T homozygotes (mean ± SD, 307 ± 72 mg/dL) and intermediate among ,1323T/C heterozygotes (mean ± SD, 314 ± 62 mg/dL; P = 0.015). Similarly, in the LDLR-mutation carriers, the plasma triglyceride levels were highest among ,1323C homozygotes (mean ± SD, 371 ± 381 mg/dL), lowest among ,323T homozygotes (mean ± SD, 171 ± 94 mg/dL), and intermediate among ,1323T/C heterozygotes (mean ± SD, 218 ± 130 mg/dL; P = 0.003). No such gene-interactive effect was observed among non-carriers of the LDLR-mutation. Conclusion: These results indicate a significant modification of the phenotype of FH with defective LDLR allele, by GSBS-1323C allele in the kindred studied. [source] A Novel ATP1A2 Gene Mutation in an Irish Familial Hemiplegic Migraine KindredHEADACHE, Issue 1 2008Desiree M. Fernandez MRCP Objective., We studied a large Irish Caucasian pedigree with familial hemiplegic migraine (FHM) with the aim of finding the causative gene mutation. Background., FHM is a rare autosomal-dominant subtype of migraine with aura, which is linked to 4 loci on chromosomes 19p13, 1q23, 2q24, and 1q31. The mutations responsible for hemiplegic migraine have been described in the CACNA1A gene (chromosome 19p13), ATP1A2 gene (chromosome 1q23), and SCN1A gene (chromosome 2q24). Methods., We performed linkage analyses in this family for chromosome 1q23 and performed mutation analysis of the ATP1A2 gene. Results., Linkage to the FHM2 locus on chromosome 1 was demonstrated. Mutation screening of the ATP1A2 gene revealed a G to C substitution in exon 22 resulting in a novel protein variant, D999H, which co-segregates with FHM within this pedigree and is absent in 50 unaffected individuals. This residue is also highly conserved across species. Conclusions., We propose that D999H is a novel FHM ATP1A2 mutation. [source] A human phospholamban promoter polymorphism in dilated cardiomyopathy alters transcriptional regulation by glucocorticoids,HUMAN MUTATION, Issue 5 2008Kobra Haghighi Abstract Depressed calcium handling by the sarcoplasmic reticulum (SR) Ca-ATPase and its regulator phospholamban (PLN) is a key characteristic of human and experimental heart failure. Accumulating evidence indicates that increases in the relative levels of PLN to Ca-ATPase in failing hearts and resulting inhibition of Ca sequestration during diastole, impairs contractility. Here, we identified a genetic variant in the PLN promoter region, which increases its expression and may serve as a genetic modifier in dilated cardiomyopathy (DCM). The variant AF177763.1:g.203A>C (at position ,36,bp relative to the PLN transcriptional start site) was found only in the heterozygous form in 1 out of 296 normal subjects and in 22 out of 381 cardiomyopathy patients (heart failure at age of 18,44 years, ejection fraction=22±9%). In vitro analysis, using luciferase as a reporter gene in rat neonatal cardiomyocytes, indicated that the PLN-variant increased activity by 24% compared to the wild type. Furthermore, the g.203A>C substitution altered the specific sequence of the steroid receptor for the glucocorticoid nuclear receptor (GR)/transcription factor in the PLN promoter, resulting in enhanced binding to the mutated DNA site. These findings suggest that the g.203A>C genetic variant in the human PLN promoter may contribute to depressed contractility and accelerate functional deterioration in heart failure. Hum Mutat 29(5), 640,647, 2008. © 2008 Wiley-Liss, Inc. [source] Molecular analysis of genomic DNA allows rapid, and accurate, prenatal diagnosis of peroxisomal D-bifunctional protein deficiencyPRENATAL DIAGNOSIS, Issue 1 2002B. C. Paton Abstract Prenatal diagnosis was requested for a couple with a previous child affected by the peroxisomal disorder D-bifunctional protein deficiency. Prior analysis of the D-bifunctional protein cDNA sequence from the propositus had shown that it was missing 22,bp. This was subsequently attributed to a point mutation in the intron 5 donor site (IVS5+1G>C) of the D-bifunctional protein gene. Consistent with parental consanguinity, the patient was shown to be homozygous for this mutation, which is associated with loss of a Hph 1 restriction site in the genomic sequence. Prenatal testing of the fetus using genomic DNA isolated from uncultured amniocytes indicated that both alleles of the D-bifunctional protein had the IVS5+1G>C substitution. The peroxisomal defect was later confirmed biochemically using cultured amniocytes, which were found to have elevated levels of very long chain fatty acids (VLCFA). This is the first report of prenatal diagnosis of D-bifunctional protein deficiency using molecular analysis of genomic DNA. Copyright © 2002 John Wiley & Sons, Ltd. [source] Frequencies of the -330 (T , G) IL-2 and -590 (T , C) IL-4 gene polymorphisms in a population from south-eastern BrazilINTERNATIONAL JOURNAL OF IMMUNOGENETICS, Issue 4 2002R. M. Scarel-Caminaga Summary Polymorphisms in the promoter regions of cytokine genes may affect their transcription. A T/G substitution at position ,330 of the interleukin-2 (IL-2) gene and a T/C substitution at position ,590 of the interleukin-4 (IL-4) gene have been described previously. The ,590 (T , C) IL-4 gene polymorphism was associated with asthma and atopy in US and Japanese populations. Population genetics is a useful tool for determination of the biological significance of genetic polymorphisms. The aim of this study was to investigate the frequencies of polymorphisms in the promoter regions of the IL-2 and IL-4 genes in a population from south-eastern Brazil and to compare them with those published for other populations. Allele frequencies were estimated in 114 unrelated individuals from São Paulo State. These subjects had an average age of 41.2 years (± 12.4 years) and the ethnic composition of the sample was: 78.07% Caucasian, 11.4% Black and 10.53% Mulatto. DNA from subjects was extracted from epithelial buccal cells, and the PCR-RFLP technique was employed to investigate the ,330 (T , G) IL-2 and ,590 (T , C) IL-4 gene polymorphisms. The allele frequency of the IL-2 gene polymorphism obtained in our study was similar to that found in UK Caucasoid groups. The T allele frequency of the IL-4 gene polymorphism observed in the Caucasian Brazilian group was similar to that found in UK and Australian populations, while the frequency observed for the Black Brazilian group was similar to that found in Japanese and Kuwaiti Arab populations. The results for the ,330 (T , G) IL-2 and ,590 (T , C) IL-4 polymorphisms are consistent with the high contribution of European lineages to the population in south-eastern Brazil. [source] | |||||||||||||