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Intronic Sequences (intronic + sequence)
Kinds of Intronic Sequences Selected AbstractsPolymorphisms located in the region containing BHMT and BHMT2 genes as maternal protective factors for orofacial cleftsEUROPEAN JOURNAL OF ORAL SCIENCES, Issue 4 2010Adrianna Mostowska Mostowska A, Hozyasz KK, Biedziak B, Misiak J, Jagodzinski PP. Polymorphisms located in the region containingBHMTandBHMT2genes as maternal protective factors for orofacial clefts. Eur J Oral Sci 2010; 118: 325,332. © 2010 The Authors. Journal compilation © 2010 Eur J Oral Sci Nonsyndromic cleft lip with or without cleft palate (NCL/P) is one of the most common craniofacial malformations; however, its aetiology is still unclear. Because the effects of maternal nutrition on fetal development are well known, we decided to pursue the question of whether polymorphic variants of genes encoding enzymes involved in choline metabolism might be associated with the maternal risk of having a baby with NCL/P. Analysis of 18 single nucleotide polymorphisms (SNPs) of betaine-homocysteine methyltransferase (BHMT), betaine-homocysteine methyltransferase-2 (BHMT2), choline dehydrogenase (CHDH), choline kinase (CHKA), dimethylglycine dehydrogenase (DMGDH), choline-phosphate cytidylyltransferase A (PCYT1A), and phosphatidylethanolamine N -methyltransferase (PEMT) provided evidence that polymorphisms located in the region containing BHMT and BHMT2 were protective factors against NCL/P affected pregnancies in our population. The strongest signal was found for the SNP located in the intronic sequence of BHMT2. Women carrying two copies of the rs625879 T allele had a significantly decreased risk of having offspring with orofacial clefts. These results were significant, even after correction for multiple comparisons. Moreover, the gene,gene interaction analysis revealed a significant epistatic interaction of BHMT2 (rs673752), PEMT (rs12325817), and PCYT1A (rs712012) with maternal NCL/P susceptibility. Altogether, our study identified a novel gene, the nucleotide variants of which were be associated with a decreased risk of having a baby with NCL/P. [source] A new Groucho TLE4 protein may regulate the repressive activity of Pax5 in human B lymphocytesIMMUNOLOGY, Issue 4 2002Michèle Milili Summary During mouse B-cell development, Pax5 is an essential transcription factor that acts as an activator of B-cell-specific genes and as a repressor of alternative lineage fates. The repressive function is mediated by the recruitment of members of the Groucho co-repressor family. Using an RNA display approach, we have isolated a transcript, called QD, specifically expressed in human pro-B and pre-B cells, which is derived from the human Groucho TLE4 gene. The QD transcript contains the first four TLE4 exons and an intronic sequence 3, of exon 4, demonstrating that QD is a splice variant of TLE4. The putative resulting protein of 94 amino acids corresponds to approximately half of an N-terminal tetramerization domain. We also show specific expression of TLE4 transcripts in human B cells and of TLE4 proteins in B-cell nuclei. Moreover, we demonstrate that recombinant QD protein binds to the TLE4 Q domain and is able to abolish the TLE4/Pax5 interaction. Thus, QD could act as a negative regulator of TLE4 function, in early B-cell differentiation. [source] Disruption of FRNK expression by gene targeting of the intronic promoter within the focal adhesion kinase geneJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 4 2007Haruko Hayasaka Abstract FRNK, a non-catalytic variant of focal adhesion kinase (FAK), is expressed in major blood vessels throughout mouse development and is postulated to play a role in regulating cell adhesion and signaling in vascular smooth muscle cells (VSMCs). The FRNK transcriptional start site lies within an intron of the FAK gene, suggesting that the FRNK gene is a "gene within a gene". Here, we identified a 1 kb intronic sequence of the FAK gene that is necessary for endogenous FRNK expression. Deletion of this sequence in gene-targeted mice abolished FRNK expression, showing the direct involvement of the FAK intron in the regulation of FRNK expression. The level of FAK expression was normal in the FRNK-deficient mice, indicating that FAK and FRNK are transcriptionally regulated by distinct promoters. The FRNK-deficient mice were viable, fertile, and displayed no obvious histological abnormalities in any of the major blood vessels. Western blot analysis showed that FRNK,deficient and wild-type (WT) cells had comparable levels of steady-state and adhesion-dependent FAK autophosphorylation. Despite the fact that ectopic expression of FRNK suppresses focal adhesion formation in cultured cells, these results suggest that endogenous FRNK is not essential for development or the formation of the mouse vasculature. J. Cell. Biochem. 102: 947,954, 2007. © 2007 Wiley-Liss, Inc. [source] Malonyl CoA decarboxylase deficiency: C to T transition in intron 2 of the MCD geneJOURNAL OF NEUROSCIENCE RESEARCH, Issue 6 2001Sankar Surendran Abstract Malonyl CoA decarboxylase (MCD) is an enzyme involved in the metabolism of fatty acids synthesis. Based on reports of MCD deficiency, this enzyme is particular important in muscle and brain metabolism. Mutations in the MCD gene result in a deficiency of MCD activity, that lead to psychomotor retardation, cardiomyopathy and neonatal death. To date however, only a few patients have been reported with defects in MCD. We report here studies of a patient with MCD deficiency, who presented with hypotonia, cardiomyopathy and psychomotor retardation. DNA sequencing of MCD revealed a homozygous intronic mutation, specifically a ,5 C to T transition near the acceptor site for exon 3. RT-PCR amplification of exons 2 and 3 revealed that although mRNA from a normal control sample yielded one major DNA band, the mutant mRNA sample resulted in two distinct DNA fragments. Sequencing of the patient's two RT-PCR products revealed that the larger molecular weight fragments contained exons 2 and 3 as well as the intervening intronic sequence. The smaller size band from the patient contained the properly spliced exons, similar to the normal control. Western blotting analysis of the expressed protein showed only a faint band in the patient sample in contrast to a robust band in the control. In addition, the enzyme activity of the mutant protein was lower than that of the control protein. The data indicate that homozygous mutation in intron 2 disrupt normal splicing of the gene, leading to lower expression of the MCD protein and MCD deficiency. J. Neurosci. Res. 65:591,594, 2001. © 2001 Wiley-Liss, Inc. [source] Intronic variants in BRCA1 and BRCA2 that affect RNA splicing can be reliably selected by splice-site prediction programs,HUMAN MUTATION, Issue 1 2009Maaike P.G. Vreeswijk Abstract A large number of sequence variants identified in BRCA1 and BRCA2 cannot be distinguished as either disease-causing mutations or neutral variants. These so-called unclassified variants (UVs) include variants that are located in the intronic sequences of BRCA1 and BRCA2. The purpose of this study was to assess the use of splice-site prediction programs (SSPPs) to select intronic variants in BRCA1 and BRCA2 that are likely to affect RNA splicing. We performed in vitro molecular characterization of RNA of six intronic variants in BRCA1 and BRCA2. In four cases (BRCA1, c.81,6T>A and c.4986+5G>T; BRCA2, c.7617+2T>G and c.8754+5G>A) a deleterious effect on RNA splicing was seen, whereas the c.135-15_-12del variant in BRCA1 showed no effect on RNA splicing. In the case of the BRCA2 c.68,7T>A variant, RNA analysis was not sufficient to establish the clinical significance. Six SSPPs were used to predict whether an effect on RNA splicing was expected for these six variants as well as for 23 intronic variants in BRCA1 for which the effect on RNA splicing has been published. Out of a total of 174 predictions, 161 (93%) were informative (i.e., the wild-type splice-site was recognized). No false-negative predictions were observed; an effect on RNA splicing was always predicted by these programs. In four cases (2.5%) a false-positive prediction was observed. For DNA diagnostic laboratories, these programs are therefore very useful to select intronic variants that are likely to affect RNA splicing for further analysis. Hum Mutat 0,1,8, 2008. © 2008 Wiley-Liss, Inc. [source] Molecular characterization of familial hypercholesterolemia in German and Greek patients,,HUMAN MUTATION, Issue 3 2004George V. Z. Dedoussis Abstract We used the denaturing gradient gel electrophoresis (DGGE) method to define mutations in the promoter region, the 18 exons, and their flanking intronic sequences of the low-density lipoprotein (LDL) receptor gene LDLR, causing familial hypercholesterolemia (FH) phenotype in 100 German and in 100 Greek hypercholesterolemic individuals. In addition, we tested all patients for the presence of mutations in codons 3456-3553 of the gene encoding apolipoprotein B-100 (APOB). Twenty-six aberrant DGGE patterns were identified and subsequently directly sequenced. In LDLR, two novel missense mutations (c.1957G>T/p.V653F, c.647 G>A/p.C216Y) and one novel homozygous base substitution c.1-156 C>T in the repeat 2 of the promoter region were identified among German FH patients; one novel splice site c.1060+10C>G was identified among Greek FH patients. One of the German FH patients was a carrier for the mutations c.1171G>A/p.A391T and p.V653F, and two of the Greek FH patients were compound heterozygotes for the mutations c.1150C>T/p.Q384X and c.1158C>G/p.D386E. Two German FH patients carried the mutation p.R3500Q within APOB. Comparing the mutations within the LDLR gene of the two European FH populations, the German population seems to be more heterogeneous than the Greek cohort. Further studies in progress are trying to elucidate the responsiveness to drug therapy in association with LDLR genotype and the nutritional habits of the two FH populations. © 2004 Wiley-Liss, Inc. [source] Constitutional NF1 mutations in neurofibromatosis 1 patients with malignant peripheral nerve sheath tumors,,HUMAN MUTATION, Issue 5 2003Lan Kluwe Abstract Neurofibromatosis type 1 (NF1) patients have 10% of lifetime risk for developing malignant peripheral nerve sheath tumors (MPNST), one of the most aggressive cancers. We examined the spectrum of constitutional NF1 mutations among 24 NF1 patients with MPNST. We found mutations in 18 patients: four megabase deletions involving the NF1 gene, 13 truncating mutations, and only one missense mutation. One deletion included both exonic and intronic sequences. No typical splicing mutation was found. Five of these mutations were novel: c.3686delA, c.197_204+9del17, c.3044T>C (p.Leu1015Pro), c.2497delT, and c.6020_6027dup. The proportion of megabase deletions of the NF1 gene found in patients with MPNST (17%=4/24) was higher than that in a group of unselected NF1 patients (5.4%=27/500). © 2003 Wiley-Liss, Inc. [source] A novel intracellular isoform of VEGFR-1 activates Src and promotes cell invasion in MDA-MB-231 breast cancer cellsJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 3 2010Belén Mezquita Abstract Two types of VEGFR-1 receptors have been characterized: a full-length transmembrane receptor and a truncated extracellular soluble isoform (sVEGFR-1). We report here the characterization, in normal and cancer cells, of a new family of intracellular isoforms of VEGFR-1 resulting from alternative initiation of transcription in intronic sequences of the gene. While the classical isoforms of VEGFR-1 were barely detectable in MDA-MB-231 breast cancer cells, one of the intracellular isoforms transcribed from intron 21 (i21VEGFR-1) was the main isoform expressed in these cells. The new transcript encodes for a protein that contains only the phosphotransferase domain and the carboxyterminal tail of VEGFR-1. Treatment of MDA-MB-231 cells with siRNA specific for the tyrosine domain of VEGFR-1 suppressed the expression of i21VEGFR-1, downregulated phosphorylation of Src at tyrosine 418, and reduced markedly the invasion capacity of these cells in vitro. Accordingly, overexpression of transfected i21VEGFR-1 in MDA-MB-231 cells upregulated the active form of Src and increased invasiveness of MDA-MB-231 cells. The expression of i21VEGFR-1 in MDA-MB-231 cells was inhibited by retinoic acid. Both, activation of Src and downregulation by retinoic acid, have been reported in other intracellular members of the Fms/Kit/PDGFR family of tyrosine kinases, particularly in the intracellular isoform of c-kit, analogous structurally to i21VEGFR-1 and frequently expressed in cancer cells. J. Cell. Biochem. 110: 732,742, 2010. © 2010 Wiley-Liss, Inc. [source] Gene expression microarray analysis and genome databases facilitate the characterization of a chromosome 22 derived homogenously staining regionMOLECULAR CARCINOGENESIS, Issue 1 2004Suzanna L. Arcand Abstract Karyotype and fluorescence in situ hybridization (FISH) analyses previously identified a homogenously staining region (HSR) derived from chromosome 22 in OV90, an epithelial ovarian cancer (EOC) cell line. Affymetrix® expression microarrays in combination with the UniGene and Human Genome Browser databases were used to identify the candidate genes comprising the amplicon of the HSR, based on comparison of expression profiles of OV90, EOC cell lines lacking HSRs and primary cultures of normal ovarian surface epithelial (NOSE) cells. A group of probe sets displaying a minimum 3-fold overexpression with a high reliability score (P-call) in OV90 were identified which represented genes that mapped within a 1,2 Mb interval on chromosome 22. A large number of probe sets, some of which represent the same genes, displayed no evidence of overexpression and/or low reliability scores (A-call). An investigation of the probe set sequences with the Affymetrix® and Sanger Institute Chromosome 22 Group databases revealed that some of the probe sets displaying discordant results for the same gene were complementary to intronic sequences and/or the antisense strand. Microarray results were validated by RT-PCR. Genomic analysis suggests that the HSR was derived from the amplification of a 1.1 Mb interval defined by the chromosomal map positions of ZNF74 and Hs.372662, at 22q11.21. The deduced amplicon is derived from a complex region of chromosome 22 that harbors low-copy repeats (LCRs). The amplicon contains 18 genes as likely targets for gene amplification. This study illustrates that large-scale expression microarray analysis in combination with genome databases is sufficient for deducing target genes associated with amplicons and stresses the importance of investigating probe set design before engaging in validation studies. © 2004 Wiley-Liss, Inc. [source] Evaluation of a fetus at risk for dihydropteridine reductase deficiency by direct mutation analysis using denaturing gradient gel electrophoresisPRENATAL DIAGNOSIS, Issue 10 2001H. Serap Kalkano Abstract Dihydropteridine reductase (DHPR) is an enzyme involved in the recycling of tetrahydrobiopterin (BH4), which is an obligate co-factor of the aromatic amino acid hydroxylases. DHPR deficiency is a rare, autosomal recessive disorder caused by mutations in the QDPR gene. DHPR-deficient patients are diagnosed by a lack of response to a low phenylalanine diet and by severe neurological symptoms. Final diagnosis is made by measurements of neurotransmitters and pterin metabolites in cerebrospinal fluid (CSF) and urine, in addition to DHPR enzyme activity, which can be assessed in whole red blood cells. Treatment of DHPR deficiency can be difficult and the outcome is not always satisfying, even if all treatment strategies are followed. Therefore prenatal diagnosis is of great importance in affected families. Prenatal diagnosis is possible by measuring DHPR activity in different cell types but this is time consuming. More than 25 different mutations have to date been identified in the QDPR gene and direct identification of a mutation in a fetus would be easy and rapid. We have developed a method based on denaturing gradient gel electrophoresis (DGGE) for the analysis of the QDPR gene. The method is useful for rapid and simultaneous scanning of all exons and flanking intronic sequences of the QDPR gene. We describe the first prenatal diagnosis conducted using this method. Copyright © 2001 John Wiley & Sons, Ltd. [source] Systematic search for mutations in the human tissue inhibitor of metalloproteinases-3 (TIMP-3) gene on chromosome 22 and association study with schizophreniaAMERICAN JOURNAL OF MEDICAL GENETICS, Issue 3 2001Chao-Chun Hung Abstract Several linkage studies have suggested that chromosome 22q12,q13 is a putative region for schizophrenic genes. In this study, the human tissue inhibitor of metalloproteinase-3 (TIMP-3) gene was investigated as positional candidate gene for schizophrenia because of its regulatory function on extracellular matrix proteins, cell adhesion molecules, and neural cell adhesion molecules in the brain. We systematically searched for the nucleotide variants by sequencing all the exons and their flanking intronic sequences in a sample of Chinese schizophrenic patients from Taiwan. Two silent mutations in the exon 3 were identified: c.249T,C at codon 83 (His) and c.261C,T at codon 87 (Ser). However, no mutations causing amino acid alteration or aberrant splicing of transcripts were observed. Hence, it is unlikely that the TIMP-3 gene itself may play an important role in the genetic susceptibility to schizophrenia. Further case control association study revealed a significant difference of genotype distribution of the c.249T,C between schizophrenic patients and control. This finding supports that 22q12 is a schizophrenia susceptible region, and it is likely that there might be other genetic mutations in the neighborhood of the TIMP-3 gene locus that may contribute to the susceptibility of schizophrenia. © 2001 Wiley-Liss, Inc. [source] Effect of Polymorphisms in Four Candidate Genes for Fertility on Litter Size in a German Pig LineREPRODUCTION IN DOMESTIC ANIMALS, Issue 4 2010A Spötter Contents We carried out an SNP discovery project in pigs for candidate genes playing potentially important roles in embryonic development. Using eight pigs one each from eight breeds (Meishan, Mangalitza, Duroc, Pietrain, German Landrace, Hampshire, Husum Red Pied, German Large White), 36 SNPs were identified in intronic sequences of 21 porcine candidate genes based on sequencing of PCR products. The primer pairs were designed using porcine EST sequences allowing amplification of introns. These SNPs were tested for their association with the number of piglets born alive in German Large White sows using a discordant approach. Significant effects (p < 0.001 and p < 0.05, respectively) of intronic SNPs on litter size were found for four genes: mitogen-activated protein kinase kinase kinase 3 (MAP3K3), vascular endothelial growth factor receptor (KDR), erbb2 interacting protein (ERBB2IP) and peroxisome proliferator-activated receptor delta (PPARD). These SNPs can be further tested in upcoming association studies for their influence on litter size in different breeds using larger sample sizes. [source] Refined candidate region specified by haplotype sharing for Escherichia coli F4ab/F4ac susceptibility alleles in pigsANIMAL GENETICS, Issue 1 2010M. Jacobsen Summary Infection of the small intestine by enterotoxigenic Escherichia coli F4ab/ac is a major welfare problem and financial burden for the pig industry. Natural resistance to this infection is inherited as a Mendelian recessive trait, and a polymorphism in the MUC4 gene segregating for susceptibility/resistance is presently used in a selection programme by the Danish pig breeding industry. To elucidate the genetic background involved in E. coli F4ab/ac susceptibility in pigs, a detailed haplotype map of the porcine candidate region was established. This region covers approximately 3.7 Mb. The material used for the study is a three generation family, where the founders are two Wild boars and eight Large White sows. All pigs have been phenotyped for susceptibility to F4ab/ac using an adhesion assay. Their haplotypes are known from segregation analysis using flanking markers. By a targeted approach, the candidate region was subjected to screening for polymorphisms, mainly focusing on intronic sequences. A total of 18 genes were partially sequenced, and polymorphisms were identified in GP5, CENTB2, APOD, PCYT1A, OSTalpha, ZDHHC19, TFRC, ACK1, MUC4, MUC20, KIAA0226, LRCH3 and MUC13. Overall, 227 polymorphisms were discovered in the founder generation. The analysis revealed a large haplotype block, spanning at least 1.5 Mb around MUC4, to be associated with F4ab/ac susceptibility. [source] R58fs Mutation in the HGD Gene in a Family with Alkaptonuria in the UAEANNALS OF HUMAN GENETICS, Issue 1 2009Yousef M Abdulrazzaq Summary This study was conducted to determine the prevalence of alkaptonuria in the UAE population and to identify the genotype of affected individuals. In a 3 stage sampling technique 2981 pupils from Government schools in Al Ain and private schools in Dubai were selected to take part in the study, of whom 2857 provided urine samples. Urine collected was analysed for homogentisic acid by gas chromatography-mass spectrometry. Genomic DNA was isolated from the white blood cells of all family members of the affected case following standard established protocols. Specific PRC primers were designed to amplify all 14 exons of the HGD gene with the flanking intronic sequences including the splice site sequences. 2857 children returned a viable urine sample, of which one was highly positive for homogentisic acid. All 12 members of this girl's family were studied and one, a 22 year old brother, was found to excrete HGA. Another, a sister who had not provided a urine sample, was discovered by genetic testing. There were no complaints of joint pain or other symptoms in any member of this family. Parents were first cousins. We found a single nucleotide deletion c.342delA, located in exon 3, which resulted in a frameshift at amino acid position 58 (p.Arg58fs or p.R58fs). Alkaptonuria may be more common than it is thought to be with an allele prevalence estimated at 0.0107 (95% CI 0.000392 , 0.03473). The R58fs mutation is old, perhaps having occurred several thousand years ago, and has spread over a large geographical area. [source] | |||||||||||||