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At least 613 records · Page 34Linked to original sources
PubMed2006

Plasma levels of bradykinin are suppressed in factor XII-deficient mice.

A genetically-transmissible factor (F) XII-inactivated allele has been produced in mice by targeted replacement of exons 3-8 of the FXII gene with the neomycin resistance gene. Interbreeding of these mice provided offspring homozygous for two inactivated FXII alleles (FXII(-/-)). Male and female FXII-deficient mice bred normally in all genotypic combinations of the heterozygous and homozygous states, and the offspring survived to adulthood, suggesting that a total FXII deficiency does not affect embryonic development and survival. Neither FXII transcripts nor FXII antigen was found in various tissues of adult FXII(-/-) mice. No obvious unchallenged coagulopathies were present in FXII(-/-) adult mice, despite greatly prolonged activated partial thromboplastin times in this mouse cohort. FXII(-/-) mice were then used to assess the in vivo importance of the plasma FXII/prekallikrein/kininogen pathway in provision of resting plasma bradykinin (BK) levels and in generation of plasma BK stimulated by contact with an artificial surface, using a new and greatly improved plasma BK assay developed during these studies. It was found that approximately 50% of resting BK, and all of the contact-stimulated plasma BK, was provided by this FXII-dependent pathway, without a requirement for FXI. These results provide clear evidence that surface-stimulated BK production, in mice, is dependent on the activation of FXII.

Animals↗
PubMed1989

Gene replacement and retrieval with recombinant M13mp bacteriophages.

We have developed an allele exchange system for shuttling sequences of DNA to and from their original chromosomal loci. Cloned segments of the histidine operon of Salmonella typhimurium and the lactose operon of Escherichia coli served as target sequences and were used to develop the system. Replacement and retrieval of target sequences used the phage M13mp vectors and proceeded through an M13 lysogen intermediate. The intermediates and products of allele exchange were characterized by genetic and hybridization analyses. Several unique properties of M13 lysogens were exploited to devise positive selections to detect integration and excision. These positive selections were used to manipulate phenotypically silent alleles.

Chromosome Deletion↗
PubMed1992

Molecular basis of a unique African variant (A-IV 5) of human apolipoprotein A-IV and its significance in lipid metabolism.

Human apolipoprotein A-IV (apoA-IV) exhibits a genetically determined structural polymorphism amenable to analysis by isoelectric focusing and immunoblotting techniques. We have determined the allele frequency and molecular basis of a unique ApoA-IV*5 allele which is widely distributed among blacks but is absent in other populations. The frequency of the ApoA-IV*5 allele in blacks (N = 308) was estimated to be 3.2%. In comparison to the common ApoA-IV*1 allele, analysis of coding and non-coding sequences of the ApoA-IV*5 allele revealed an in-frame insertion of 12 nucleotides near the carboxyl terminal region of the mature protein. The insertion involves an exact duplication of the second of the four repeats and codes for 4 amino acids glutamic acid (GAA), glutamine (CAG), glutamine (CAG), and glutamine (CAG) and is responsible for the charge shift of the the apoA-IV 5 isoform slightly toward the anode as compared to the wild type apoA-IV 1 isoform on the isoelectric focusing gel. This in-frame insertion occurs in a region which is highly conserved among rat, mouse, and humans. In addition to the 12 nucleotide insertion, the four individuals sequenced for the ApoA-IV*5 allele also revealed a same-sense mutation by replacing G to T at the third position of codon 316. Our preliminary data suggest that this unique black allele marker may be of potentially significance in studies of human lipid metabolism and in microevolution.

Adolescent↗
PubMed1992

T-cell receptor gamma/delta: comparison of gene configurations and function between humans and chimpanzees.

The human and chimpanzee T-cell receptor gamma-delta (TCR gamma delta) bearing cells represent a minor subset (3-8%) of T lymphocytes. In the periphery, the TCR gamma delta population has a restricted combinatorial repertoire. The TCRD-V1 and -V2 gene products are expressed in a mutually exclusive fashion, whereas, the TCRD-V2 and the TCRG-V9 encoded proteins show, in general, a coordinated expression. Restriction fragment length polymorphism analysis showed conservation of the restriction sites that identify the TCRG-V9 and TCRD-V2 rearrangements. The human TCRG-V9 locus has two alleles, TCRG-V9A1 and TCRG-V9A2 differing at codon position 31. The chimpanzee TCRG-V9 gene product differs from the products of the human TCRG-V9A1 and TCRG-V9A2 allele by two and three amino acid replacements, respectively. The human and the chimpanzee TCRG-V9-TCRD-V2 lymphocytes show a similar specific proliferative and cytolytic response to human Daudi Burkitt's lymphoma cells. Therefore, the amino acid replacements found in the chimpanzee TCRG-V9 gene product do not change the superantigen specificity across this species barrier.

Alleles↗
PubMed2004

Evidence for positive selection at the pantophysin (Pan I) locus in walleye pollock, Theragra chalcogramma.

Nucleotide polymorphism at the pantophysin (Pan I) locus in walleye pollock, Theragra chalcogramma, was examined using DNA sequence data. Two distinct allelic lineages were detected in pollock, resulting from three amino acid replacement mutations in the first intravesicular domain of the protein. The common Pan I allelic group, comprising 94% of the samples, was less polymorphic (pi = 0.005) than the uncommon group (pi = 0.008), and nucleotide diversity in both was higher than for two allelic lineages in the related Atlantic cod, Gadus morhua. Phylogenetic analyses of Pan I sequences from these two species did not clearly resolve orthology among allelic groups, in part because of recombination that has occurred between the two pollock lineages. Conventional tests of neutrality comparing polymorphisms within and between homologous regions of the Pan I locus in walleye pollock and Atlantic cod did not detect the effects of selection. This result is likely attributed to low levels of synonymous divergence among allelic lineages and a lack of mutation-drift equilibrium inferred from nucleotide mismatch frequency distributions. However, the ratio of nonsynonymous to synonymous substitutions per site (dN/dS) exceeded unity in two intravesicular domains of the protein and the influence of positive selection at multiple codon sites was strongly inferred through the use of maximum-likelihood analyses. In addition, the frequency spectrum of linked neutral variation showed indirect effects of adaptive hitchhiking in pollock resulting from a selective sweep of the common allelic lineage. Recombination between the two allelic classes may have prevented complete loss of the older, more polymorphic lineage. The results suggest that recurrent sweeps driven by positive selection is the principle mode of evolution at the Pan I locus in gadid fishes.

Adaptation, Biological↗
PubMed2002

A novel cis AB allele derived from a B allele through a single point mutation.

BACKGROUND: The very rare cis AB phenotype, first described in 1964, corresponds to a special ABO allele encoding a glycosyltransferase that is capable of synthesizing both A and B substances. Until now, gene sequences of only two cis AB alleles were partially characterized. One involved the A1*02 allele with a single nonsynonymous substitution at codon 268, whereas the second arose from a single nonsynonymous substitution at codon 266 in exon 7 of a B1*01 allele. STUDY DESIGN AND METHODS: A cis AB phenotype was identified in a French family. The serologic characteristics of this phenotype were determined. The cis AB allele was characterized from exon 6 to exon 7 by cloning and sequencing. RESULTS: The cis AB.tlse(*)01 allele is identical to B(1*)01 except for a single point mutation at nucleotide position 700, where a T replaces a C, implying a change of amino acid 234 (the B(1*)01 proline being replaced by a serine). CONCLUSION: The cis AB.tlse(*)01 allele clearly differs from all previously reported ABO, including the two previous cis AB described.

ABO Blood-Group System↗
PubMed2000

G protein beta 3 gene: structure, promoter, and additional polymorphisms.

Recent studies have shown that a polymorphism (C825T) in the gene encoding the G protein beta 3 subunit (GNB3) is associated with hypertension and obesity. We characterized the entire GNB3 gene, which spans 7.5 kb and is composed of 11 exons and 10 introns. Its promoter lacks a TATA box but harbors GC-rich regions. The functional activity of the GNB3 promoter was verified with reporter gene assays that also demonstrated its inducibility by phorbol esters. A novel polymorphism in the promoter region A(-350)G occurred with frequencies (G allele) of 76%, 97%, and 61% in Africans, Chinese, and Germans, respectively. Reporter gene constructs with either the A or the G allele did not differ with regard to inducement of the reporter protein. A silent nucleotide exchange in the coding region (A657T) occurred with T allele frequencies ranging from 0.5% to 2.4%. Another polymorphism (G814A) results in the replacement of glycine by serine at position 272. In Germans, the A allele occurred at a frequency of 10%. Finally, a C1429T polymorphism in the 3' untranslated region of GNB3 was identified that occurred at T allele frequencies of 38%, 17%, and 30% in Africans, Chinese, and Germans, respectively. Haplotype prediction indicated in Germans an almost complete association of GNB3 825T with 1429T, and vice versa. An analysis of these polymorphic loci in nonhuman primates revealed that the ancestral GNB3 gene harbored the (-350)G, 825C, and 1429C alleles. This is the first complete characterization of the human GNB3 gene and its promoter region, which will enable refined epidemiological and biochemical investigations of GNB3 in hypertension and obesity.

Adolescent↗
PubMed1999

Molecular analysis in Spanish patients with muscle carnitine palmitoyltransferase deficiency.

The most common mutation in muscle carnitine palmitoyltransferase II (CPT II) deficiency is a missense mutation that replaces a leucine for a serine residue at amino acid position 113 of the CPT II protein (S113L). We performed molecular analysis in a group of 14 Spanish patients with CPT II deficiency from ten unrelated families. The S113L mutation was observed in 8 of the 14 patients studied. Seven patients were homozygous for the mutation, 1 patient was heterozygous, and 6 patients did not carry the mutation on either allele. Seven healthy relatives belonging to three different families carried the mutation on one allele. One patient carried the missense mutation that replaces a tyrosine for a serine at amino acid position 628 on one allele. Our data indicate that the S113L is also the most common mutation in Spanish patients with CPT II deficiency in muscle, and that further pathogenic mutations remain to be identified.

Carnitine O-Palmitoyltransferase↗
PubMed2003

Genome manipulation by homologous recombination in Drosophila.

By utilising a cell's recombinational machinery, researchers in many different model organisms have been able to perform gene targeting experiments in which specific sequence alterations are introduced into virtually any endogenous gene. Not only can functional knock-outs be generated by gene targeting, interesting alleles with mutations encoding specific amino acid replacements can also be made. A practical gene targeting method has only recently become available for Drosophila. This article reviews the Drosophila gene targeting method, with emphases placed on different approaches that are being used to generate different mutations.

Alleles↗
PubMed2007

Extreme context specificity in differential allelic expression.

Variability in cis-regulation of gene expression has been implicated in the phenotypic manifestation of complex traits including common, multifactorial diseases. The differential expression of alleles due to polymorphism in cis-regulatory elements is common in the human genome, but there is a paucity of information about the context specificity of these control elements. In this study, we examined the differential allelic expression (DAE) of BMP5 in human mesenchymal tissues obtained from 16 donors undergoing joint replacement for treatment of osteoarthritis. We observed significant differences in BMP5 allelic output, with allelic ratios greater than 4:1 (P < 10(-20)) in the tissues of some donors. We also discovered a significant variability in allelic expression within the different tissues of donors. For 12 of our donors, we examined the allelic expression of BMP5 in two different regions of cartilage: cartilage adjacent to the site of the osteoarthritic lesion and cartilage distal from the lesion. Five of these 12 donors demonstrated highly significant differences (P < or = 10(-8)) in allelic expression between the different regions of their cartilage. Using DAE as a phenotype, we attempted to map tissue-specific cis-regulatory polymorphisms, and we identified a single nucleotide polymorphism located downstream of BMP5, which was significantly associated with DAE in some but not all of the examined tissues. These findings suggest that allelic expression can be highly context specific and that when interrogating the cis-regulatory control of a particular gene, one cannot necessarily assume that allelic expression is conserved across different tissues or even across different regions of the same tissue.

Alleles↗
PubMed2001

A polymorphism in CYP17 and endometrial cancer risk.

Among women, the A2 allele of CYP17 has been associated with elevated levels of endogenous steroid hormones; however, it does not seem to be a strong independent risk factor for breast cancer. We assessed the association between the A2 allele of CYP17 and invasive endometrial cancer risk in a case-control study nested within the Nurses' Health Study cohort (cases: n = 184; controls: n = 554). We also evaluated whether endometrial cancer risk associated with CYP17 genotype was modified by established endometrial cancer risk factors. In addition, we further examined the relationship between CYP17 genotype and endogenous plasma steroid hormone levels among postmenopausal controls not using hormone replacement therapy (HRT). Women with the A2 allele of CYP17 were at decreased risk of endometrial cancer (A1/A1 genotype (reference); A1/A2 genotype: odds ratio, 0.89; 95% confidence interval, 0.62-1.27; A2/A2 genotype: odds ratio, 0.43; 95% confidence interval, 0.23-0.80; P trend, 0.02). We also observed the inverse association between the A2 allele and endometrial cancer risk to be stronger among women with a first-degree family history of endometrial and/or colorectal cancer (P for interaction, 0.05). Among 165 controls, we did not observe women with the A2 allele to have significantly elevated levels of any steroid hormone fraction. When these women were combined and analyzed with those women on whom we had previously examined the relationship between CYP17 genotype and circulating hormone levels (total n = 469), only modest associations were observed for the A2/A2 genotype and steroid hormone fractions estrone (versus A1/A1 genotype: +10.9%; P = 0.05) and estradiol (+8.5%; P = 0.17). These data suggest that the A2 allele of CYP17 decreases endometrial cancer risk, but has only weak effects on endogenous estrogen levels among postmenopausal women.

Adult↗
PubMed1993

Mutations in the segment polarity genes wingless and porcupine impair secretion of the wingless protein.

We have characterized the molecular nature of mutations in wingless (wg), a segment polarity gene acting during various stages of Drosophila development. Embryo-lethal alleles have undergone mutations in the protein-encoding domain of the gene, including deletions and point mutations of conserved residues. In a temperature sensitive mutation, a conserved cysteine residue is replaced by a serine. In embryo-viable alleles, the wg transcriptional unit is not affected. Immunostaining of mutant embryos shows that the embryo-lethal alleles produce either no wg antigen or a form of the protein that is retained within cells. Interestingly, embryos mutant for the segment polarity gene porcupine show a similar retention of the wg antigen. We have also transfected wild type wg alleles into Drosophila tissue culture cells, which then display wg protein on the cell surface and in the extracellular matrix. In similar experiments with mutant alleles, the proteins are retained in intracellular compartments and appear not to be secreted. These data provide further evidence that wg acts as a secreted factor and suggest that porcupine provides an accessory function for wg protein secretion or transport.

Alleles↗
PubMed2004

Oestrogen replacement therapy may improve memory functioning in the absence of APOE epsilon4.

There is currently intense controversy regarding the use of hormone replacement therapy (HRT) in postmenopausal women, in relation to its therapeutic efficacy in Alzheimer's disease (AD). It has been suggested that the benefits of HRT may be modified by apolipoprotein E (APOE) genotype (the major genetic risk factor for AD). Here we report the findings of the first study designed to systematically explore the interaction of (a) oestrogen replacement therapy (ERT) and (b) possession of an epsilon4 allele of APOE on specific elements of episodic learning and memory that are commonly used indices of age-related cognitive decline. This data represents a cross-sectional analysis of the interaction of ERT and APOE genotype on learning and memory in a cohort of 181 healthy postmenopausal women [ERT users (n = 101, mean age 65.40 +/- 6.34); ERT non-users (n = 80, mean age 67.03 +/- 6.80)] residing in Perth, Western Australia. The highest level of learning (trials 2-5; P < 0.05) and memory (e.g. total number of items recalled; P < 0.05) performance was observed in women taking ERT who were not carriers of the APOE epsilon4 allele. APOEepsilon4 carriers receiving ERT performed no better on episodic memory testing than APOE epsilon4 carriers who were not receiving ERT. These cognitive differences related to genetic profile, were noted on both recall and recognition (P = 0.005) tests of memory. The findings have significance for evaluating whether and when ERT may be clinically indicated. Specifically, ERT may benefit the cognitive functioning of women not carrying the APOE epsilon4 allele.

Aged↗
PubMed1995

Identification and characterization of the orf virus type I topoisomerase.

Vaccinia virus (VV) and Shope fibroma virus (SFV), representatives of the orthopox and leporipox genera, respectively, encode type I DNA topoisomerases. Here we report that the 957-nt F4R open reading frame of orf virus (OV), a representative of the parapox genus, is predicted to encode a 318-aa protein with extensive homology to these enzymes. The deduced amino acid sequence of F4R has 54.7 and 50.6% identity with the VV and SFV enzymes, respectively. One hundred forty amino acids are predicted to be conserved in all three proteins. The F4R protein was expressed in Escherichia coli under the control of an inducible T7 promoter, partially purified, and shown to be a bona fide type I topoisomerase. Like the VV enzyme, the OV enzyme relaxed negatively supercoiled DNA in the absence of divalent cations or ATP and formed a transient covalent intermediate with cleaved DNA that could be visualized by SDS-PAGE. Both the noncovalent and covalent protein/DNA complexes could be detected in an electrophoretic mobility shift assay. The initial PCR used to prepare expression constructs yielded a mutant allele of the OV topoisomerase with a G-A transition at nt 677 that was predicted to replace a highly conserved Tyr residue with a Cys. This allele directed the expression of an enzyme which retained noncovalent DNA binding activity but was severely impaired in DNA cleavage and relaxation. Incubation of pUC19 DNA with the wild-type OV or VV enzyme yielded an indistinguishable set of DNA cleavage fragments, although the relative abundance of the fragments differed for the two enzymes. Using a duplex oligonucleotide substrate containing the consensus site for the VV enzyme, we demonstrated that the OV enzyme also cleaved efficiently immediately downstream of the sequence CCCTT.

Amino Acid Sequence↗
PubMed2002

In Saccharomyces cerevisiae, ATP2 mRNA sorting to the vicinity of mitochondria is essential for respiratory function.

We recently demonstrated that polysome-associated mRNAs that co-isolate with mitochondria encode a subset of mitochondrial proteins, and that the 3' UTRs of these transcripts are essential for their localization to the vicinity of the organelle. To address the question of the involvement of the mRNA targeting process in mitochondrial biogenesis, we studied the role of ATP2 3' UTR. An altered ATP2 allele in which the 3' UTR was replaced by the ADH1 3' UTR exhibits properties supporting the importance of mRNA localization to the vicinity of mitochondria: (i) the mutated strain presents a respiratory dysfunction; (ii) mitochondrial import of the protein translated from the altered gene is strongly reduced, even though the precursor is addressed to the organelle surface; (iii) systematic deletions of ATP2 3' UTR revealed a 100 nucleotide element presenting RNA targeting properties. Additionally, when the ATM1 3' UTR was replaced by the ADH1 3' UTR, we obtained cells in which ATM1 mRNA is also delocalized, and presenting a respiratory dysfunction. This demonstrates that mRNA localization to the vicinity of mitochondria plays a critical role in organelle biogenesis.

3' Untranslated Regions↗
PubMed1994

Identification of allele-specific p22-phox mutations in a compound heterozygous patient with chronic granulomatous disease by mismatch PCR and restriction enzyme analysis.

A rare subgroup (approx. 5%) of all chronic granulomatous disease (CGD) patients suffers from mutations in the gene encoding the small p22-phox subunit of the flavocytochrome b558 heterodimer, the terminal redox component of the phagocyte NADPH oxidase. A male CGD patient with neutrophil granulocytes devoid of any spectrometrically detectable cytochrome b558 owing to autosomally inherited p22-phox deficiency (phenotype, A22-) is reported. The patient was identified as being compound heterozygous for two independent mutations of his p22-phox alleles. On the maternal allele a single base substitution (A186 to T) was found that predicts a nonconservative replacement of Glu 53 by Val. On his paternal p22-phox allele a G was found to be added to a G stretch between nucleotides G195 and G199 in the cDNA sequence. The resulting frame shift predicts an aberrant open reading frame, 16 amino acids longer than the normal p22-phox polypeptide. Genomic DNA was tested for the presence of the mutant allele by mismatch PCR (polymerase chain reaction). For this purpose, a single base mismatch was introduced at nucleotide position 189, leading to digestion of the normal allele by the restriction enzyme HinfI. The maternal allele was found to be present in 50% of the patient's DNA and in 50% of the DNA from his mother. The same mismatch PCR analysis with control DNA from 35 healthy individuals ruled out the possibility that the single base substitution (A186 to T) represents a common polymorphism. Inheritance of the second allelic mutation (G insertion) was verified by restriction enzyme analysis using BslI [CC(N)7GG] to digest PCR-amplified genomic DNA at the mutation site. PCR in combination with restriction enzyme analysis proved to be a powerful tool for verification of point mutations in the compound heterozygous CGD patient analyzed and may be used for prenatal diagnosis in this family.

Alleles↗
PubMed2000

Identification of two novel human CD1E alleles.

CD1 is a family of proteins structurally related to major histocompatibility complex (MHC) molecules and specialized in presenting lipids or glycolipids to T cells. In humans, there are five CD1 genes (CD1A to CD1E). It has been shown that, in contrast with classical MHC genes, CD1 loci display a very limited polymorphism. In the present work we describe two novel CD1E alleles found in two healthy Caucasian individuals. One allele differs from the wild-type by a point mutation resulting in a replacement of arginine at position 154 by a tryptophan. In the second allele we found a substitution of the leucine 184 by a proline.

Alleles↗
PubMed1996

Mutation in the thyroid hormone receptor (TR) beta gene (M313T) not previously reported in two unrelated families with resistance to thyroid hormone (RTH).

Two families expressing the RTH phenotype and harboring the same mutation in the TRbeta gene are described. Five and four affected members in each family were investigated as well as a total of 12 unaffected relatives. The mutation, a T to C transition of nucleotide 1223 in one allele of affected individuals, results in the replacement of the normal Met for a Thr. Haplotyping revealed that the same mutation developed in each family independently. Whereas attention deficit hyperactivity disorder was associated with RTH in 7 of the 9 affected individuals, it was also present in 2 family members without RTH.

Alleles↗