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Y H Edwards

Publications and source records attributed to Y H Edwards.

At least 55 records · Page 3Linked to original sources
PubMed1993

Expression of the dystrophin-related protein (utrophin) gene during mouse embryogenesis.

The utrophin (UTRN) locus is the autosomal homologue of the DMD (Duchenne muscular dystrophy) gene and encodes a protein, utrophin which is thought to be upregulated in the absence of dystrophin. In this study the spatial and temporal expression of the UTRN gene has been examined during mouse embryogenesis and compared with that of the DMD gene. The patterns of expression of these two genes are very different. Whilst DMD is expressed largely in mesodermal derivatives such as cardiac and striated muscle, UTRN shows a more widespread distribution and is expressed in neural tube, tissues which originate from neural crest and a variety of other sites of non-neural origin. In early embryos UTRN transcripts initially accumulate in the mid-neural plate and thereafter in the caudal neural tube. UTRN mRNA then becomes abundant in a subset of neural crest cell derived tissues, in particular the spinal and facial ganglia and ossifying facial cartilages. UTRN is also expressed in a variety of other sites and organs such as the tendon primordia in the digits, the pituitary, thyroid and adrenal glands, cardiac muscle, kidney and lung, follicles of the vibrissae and the outflow tract of the heart. Several patterns of UTRN expression are apparent and we discuss the possibility that these can be ascribed to a family of mRNAs transcribed from the UTRN gene using alternative promoters.

Animals↗
PubMed1993

A cosmid library specific for human chromosome regions 6p21.3 and 6q27.

We have explored the potential of irradiation-fusion gene transfer (IFGT) hybrids as a source of well-defined human chromosome fragments from which probes can be derived. Extensive characterization of the IFGT hybrid 4J4 with a full panel of markers from Chromosome (Chr) 6 showed that the human DNA content derives largely from 6p21.3 and 6q27. A cosmid library has been constructed from 4J4 DNA, and 370 recombinants containing human DNA have been isolated and overlapping clones ordered into 20 contigs. Regional localization of representative clones from each contig, determined by fluorescent in situ hybridization (FISH), places 13 contigs in 6q27 and 6 in 6p21.3. Preliminary screening of cDNA libraries with selected cosmids has identified two expressed sequences. Since there are a number of medically important genes in both these regions of human Chr 6 with several disease loci linked to the HLA-A region in 6p21.3 and various tumor suppressor genes to 6q27, this library will provide a valuable resource to aid the isolation of candidate genes for these diseases. In addition, unique markers for detailed physical and genetic mapping of these regions of human Chr 6 can be easily obtained.

Base Sequence↗
PubMed1993

Genetic polymorphism in the 3' untranslated region of human phosphoglucomutase-1.

A 317-bp segment of DNA from the 3' region of the human phosphoglucomutase-1 (PGM1) gene has been examined by a non-radioactive technique for the occurrence of single-strand conformation polymorphism (SSCP). Eight phenotypes were detected and attributed to the presence of four alleles. Genetic analysis of 75 unrelated individuals and six CEPH families whose PGM1 protein phenotypes were known revealed strong association between the PGM1 '+' and '-' isozyme phenotypes and the variation detected in this region, but no association with the PGM1 1 and PGM1 2 isozyme phenotypes. DNA sequence analysis demonstrated the presence of three nucleotide substitutions underlying the alleles, which were located in the untranslated region of the PGM1 gene. There was complete correlation between the nucleotide sequence and the phenotype detected by SSCP analysis. This study provides support for the model that the PGM1 isozyme polymorphism is determined at two distinct sites in the coding sequence, one coding for the '1' and '2' alleles and the other coding for the '+' and '-' alleles, separated by a region where intragenic recombination occurs.

Base Sequence↗
PubMed1992

Erythroid expression and DNAaseI-hypersensitive sites of the carbonic anhydrase 1 gene.

The carbonic anhydrase 1 gene is expressed in adult human and mouse erythroid cells and colon epithelia from two distinct promoters. We have explored the erythroid promoter for cis-acting sequences involved in transcription using DNAaseI as a probe. Two DNAaseI-hypersensitive sites (DHS-1 and DHS-2) have been identified in the distal erythroid promoter in CA1-expressing erythroleukaemic cells. These sites are present at low levels in K562 cells, which have a foetal/embryonic phenotype and do not express CA1. DHS-1 and DHS-2 are not present in non-erythroid cells, including colon cells, which express CA1 from the proximal colon promoter. DHS-1 and DHS-2 were also generated in an heterologous CA1 gene containing 5 kb of erythroid promoter sequence after transfection into erythroid cells, including K562 cells. These transfection studies showed that both this fragment, and an abbreviated 817 bp promoter fragment which contains only DHS-1, were sufficient to confer erythroid-specific expression to a reporter gene. These promoters were active in cell lines expressing CA1 and in K562 cells. This latter observation implies that a developmental repressor factor is both present in K562 cells and binds to a cis-acting sequence that is absent from the sequence 5 kb upstream of the erythroid transcription start site.

Base Sequence↗
PubMed1992

Phosphoglucomutase 1: complete human and rabbit mRNA sequences and direct mapping of this highly polymorphic marker on human chromosome 1.

A cDNA clone encoding the mRNA for the highly polymorphic human enzyme phosphoglucomutase 1 (PGM1; EC 5.4.2.2) has been isolated and characterized. This was achieved indirectly by first isolating a rabbit cDNA from an expression library using anti-rabbit PGM antibodies. A comparison of the nucleotide sequences shows that the homologies between human and rabbit PGM1 mRNAs are 92% and 97% for the coding nucleotide sequence and the amino acid sequence, respectively. The derived rabbit amino acid sequence is in complete agreement with the published protein sequence for rabbit muscle PGM. A physical localization of the human PGM1 gene to chromosome 1p31 has been determined by in situ hybridization. Analysis of DNA from a wide variety of vertebrates indicates a high level of PGM1 sequence conservation during evolution.

Animals↗
PubMed1992

Characterization of a 4.8kb transcript from the Duchenne muscular dystrophy locus expressed in Schwannoma cells.

The 14kb dystrophin transcript from the Duchenne muscular dystrophy (DMD) locus, which encodes a 427kDa protein, is differentially spliced at the amino terminal end giving rise to alternative transcripts expressed in muscle and brain. Here we present evidence for a 4.8kb transcript from the DMD locus which is ubiquitously expressed but is particularly abundant in Schwannoma cells where dystrophin could not be detected. The hybridisation of Western blots with dystrophin antibodies also identifies a protein of approximately 80kDa of variable abundance in different human and mdx tissues. Immunocytochemistry studies confirm the expression of this protein in nerve cells, a tissue in which full length dystrophin is not detected. Sequencing of the 5' end of a clone isolated from a rat Schwannoma cDNA library, shows that the 4.8kb transcript shares exons with the carboxy terminal end of dystrophin but the 5' untranslated region is not contained within the dystrophin transcript. We propose that the 4.8kb gene product be called apodystrophin-1 as its expression is distinct from the dystrophin 14kb mRNA but it is transcribed from the same locus.

Amino Acid Sequence↗
PubMed1992

Carbonic anhydrase 3 (CA3), a mesodermal marker.

Carbonic anhydrase 3 (CA3) is an abundant muscle protein characteristic of adult type 1, slow-twitch, fibres. The protein plays an important role in facilitated CO2 diffusion and diverse processes involving H+ and HCO-3 transport. Nucleotide sequence comparisons have identified putative promoter and enhancer regions in the 5' flanking sequences of the CA3 gene. Functional assays show that 2.8kb of 5' flanking sequence efficiently promotes transcription of a reporter gene in a muscle specific manner. Removal of sequences 5' to -722bp leads to a major loss of activity and this result implies that the proximal promoter region which includes a GArG box and four potential MyoD1 binding sites is not adequate for maximal transcription. The longest CA3 promoter construct is also active in 10T1/2 cells, which are precursor mesodermal cells and do not normally express CA3. In situ hybridization to mRNA in developing mouse embryos reveals a pattern of expression in myotomes and pre-muscles masses of the limb buds which is consistent with the regulation of CA3 by myogenic determination factors. These studies also showed that CA3 expression is not confined to cells of the muscle lineage since it is expressed in primitive mesoderm prior to the onset of myogenesis. Later in embryogenesis CA3 defines a subset of mesodermal cell types which includes not only skeletal muscle but also notochord and adipocytes.

Animals↗
PubMed1991

Tissue distribution of the dystrophin-related gene product and expression in the mdx and dy mouse.

We have previously reported a dystrophin-related locus (DMDL for Duchenne muscular dystrophy-like) on human chromosome 6 that maps close to the dy mutation on mouse chromosome 10. Here we show that this gene is expressed in a wide range of tissues at varying levels. The transcript is particularly abundant in several human fetal tissues, including heart, placenta, and intestine. Studies with antisera raised against a DMDL fusion protein identify a 400,000 Mr protein in all mouse tissues tested, including those of mdx and dy mice. Unlike the dystrophin gene, the DMDL gene transcript is not differentially spliced at the 3' end in either fetal muscle or brain.

Animals↗
PubMed1991

CAIII a marker for early myogenesis: analysis of expression in cultured myogenic cells.

Carbonic anhydrase III (CAIII) is an abundant muscle protein characteristic of adult type-1, slow-twitch, muscle fibers. We demonstrate that CAIII is not confined to mature muscle but is also expressed in cultured myogenic cells that were originally derived from adult and fetal limb muscle (G8 and C2C12) and by azacytidine treatment of 10T1/2 fibroblasts (23A2). Transcripts may accumulate in these cells to levels that correspond to 6.5% of that found in mature muscle. CAIII is expressed in mononucleate myoblasts and is abundant in those that preferentially fuse to form myotubes, and these findings contrast with those for many other muscle genes whose transcripts only accumulate on or after terminal differentiation. Preliminary promoter-function assays by transfection shows that 2.8 kb of sequence flanking the 5' end of the human CAIII gene efficiently promotes transcription of the bacterial chloramphenicol acetyltransferase gene in myogenic cells. However, none of the sequences within this region are sufficient to confer muscle-specific expression. Removal of sequences 5' to -715 bp leads to a major loss of transcriptional activity of the CAIII promoter. These results imply that the proximal CAIII promoter, which includes a putative CArG box and four potential MyoD binding sites, is not adequate for either myoblast-specific or maximal transcription.

Animals↗
PubMed1991

Physical mapping of the human carbonic anhydrase gene cluster on chromosome 8.

A cluster of genes encoding the three cytoplasmic carbonic anhydrase isozymes CAI, CAII, and CAIII lie on the long arm of chromosome 8 (8q22) in humans. These genes have been mapped using pulsed-field gel electrophoresis. The genes lie in the order CA2, CA3, CA1. CA2 and CA3 are separated by 20 kb and are transcribed in the same direction, away from CA1. CA1 is separated from CA3 by over 80 kb and is transcribed in the direction opposite to CA2 and CA3. The arrangement of the genes is consistent with proposals that the duplication event which gave rise to CA1 predated the duplication which gave rise to CA2 and CA3. The order of these three genes differs from that suggested for the mouse based on recombination frequency.

Blotting, Southern↗
PubMed1991

The gene for human neurone specific ubiquitin C-terminal hydrolase (UCHL1, PGP9.5) maps to chromosome 4p14.

Ubiquitin carboxy terminal hydrolase 1, UCHL1, is a neurone-specific protein involved in the ubiquitin-mediated proteolytic pathway. The gene for human UCHL1 has been mapped to chromosome 4 using the polymerase chain reaction to amplify specifically the human UCHL1 sequences in rodent/human somatic cell hybrid DNA. A regional assignment of this locus to 4p14 has been made by in situ hybridization to metaphase chromosomes using both tritium and fluorescently labelled probes.

Base Sequence↗
PubMed1991

Carbonic anhydrase III, an early mesodermal marker, is expressed in embryonic mouse skeletal muscle and notochord.

Carbonic anhydrase III (CAIII) is an abundant soluble protein in adult mammalian slow twitch skeletal muscle fibers. It is thought to be an early marker for myogenesis based upon its high level of expression in myoblasts in vitro prior to fusion. Using in situ hybridization, we have studied the in vivo distribution of CAIII gene transcripts in mouse embryos and fetuses from 7.25 days to 17.5 days post coitum (p.c.). CAIII mRNAs are first detected in the myotomes of somites between 9.5 and 10.5 days p.c. (20-30 somites). At 15.5 days p.c., CAIII begins to be restricted to developing slow muscle fibers. By two weeks post partum (p.p.), CAIII mRNAs are detected mainly in slow muscle fibers. CAIII transcripts are detected at an earlier stage (7.25 days p.c.) in the developing notochord. CAIII is expressed at a much higher level in the notochord than it is in developing skeletal muscle. As the notochord forms the nucleus pulposus in fetal mice, CAIII mRNA levels remain very high. Expression of CAIII in the notochord is of interest in the context of skeletal myogenesis because the notochord is thought to play an important role in somite formation. In addition to the notochord, CAIII transcripts are detected prenatally in several other non-muscle tissues: in cells of the choroid plexus, endocardial cushion and ureter, and in adipocytes.

Adipose Tissue, Brown↗
PubMed1990

CpG islands in genes showing tissue-specific expression.

Patterns of DNA methylation at CpG dinucleotides and their relations with gene expression are complex. Methylation-free CpG clusters, so-called HTF islands, are most often associated with the promoter regions of housekeeping genes, whereas genes expressed in a single-cell type are usually deficient in these sequences. However, in the human carbonic anhydrase (CA) gene family, both the ubiquitously expressed CAII and the muscle specific CAIII appear to have such CpG islands although erythrocyte-specific CAI does not. The CAII island is quantitatively more CpG rich than that of CAIII, with a CpG:GpC ratio of 0.94 compared with 0.82 for CAIII. Estimation of CpG:GpC ratios in the proximal-promoter regions of 44 vertebrate genes suggest that 40% of genes with tissue-specific or limited tissue distribution may show methylation-free CpG clusters in their promoter regions. In many cases the CpG:GpC ratio is less than that found in housekeeping genes and this may reflect variation in the interaction of CpG clusters with regulatory factors that define different patterns of tissue expression.

Animals↗
PubMed1989

An autosomal transcript in skeletal muscle with homology to dystrophin.

The Duchenne muscular dystrophy (DMD) gene has been localized to chromosome Xp21 and codes for a 14-kilobase (kb) transcript and a protein called dystrophin, of relative molecular mass 427,000. Dystrophin is associated with the cytoplasmic face of muscle fibre membranes and its C-terminal domain is thought to mediate membrane attachment. Although N-terminal and central domain structures share common features with other cytoskeletal components, no significant sequence similarity between the C-terminal region of dystrophin and other previously characterized proteins has been described. Here we report that fragments from the C-terminal domain of the DMD complementary DNA detect a closely related sequence which exhibits nucleic-acid and predicted amino-acid identities with dystrophin of approximately 65 and 80%, respectively. The dystrophin-related sequence identifies a 13-kb transcript in human fetal muscle and maps to chromosome 6. Thus, dystrophin may be a member of a family of functionally related large structural proteins in muscle.

Base Sequence↗
PubMed1989

Isozyme and DNA analysis of human S-adenosyl-L-homocysteine hydrolase (AHCY).

Erythrocyte and tissue isozymes of human AHCY have been studied by starch gel electrophoresis, cellulose acetate electrophoresis, isoelectric focusing and Na dodecyl sulphate electrophoresis. The same isozyme was observed in all the tissues studied, suggesting that human AHCY is encoded by a single structural locus. Two variant alleles were identified in erythrocyte AHCY using starch gel electrophoresis in a sample of 166 unrelated individuals from the British population. The gene frequencies were 0.024 for AHCY*2 and 0.006 for AHCY*3. The variant isozyme patterns could not be distinguished by isoelectric focusing. Using the homologous rat cDNA AHCY probe, human AHCY cDNA recombinants were isolated from a placental cDNA library. The human and rat sequences show considerable homology in the coding region of the gene and also, but to a lesser extent, in the distal part of the 3' untranslated region. Preliminary observations suggest the occurrence of a high frequency PvuII site RFLP identified with the human AHCY probe.

Adenosylhomocysteinase↗
PubMed1988

A non-methylated CpG-rich island associated with the human muscle-specific carbonic anhydrase III gene.

A cluster of CpG dinucleotides immediately upstream from exon 1 in the muscle-specific carbonic anhydrase III gene (CAIII) resembles the 'HpaII tiny fragment' (HTF) islands characteristic of mammalian 'housekeeping' genes. Since this CAIII gene shows tissue-specific expression we have carried out a detailed examination of methylation status within the CpG cluster using a polyacrylamide gel/electroblot procedure to extend the range of conventional Southern blotting. None of the clustered CpGs are methylated in DNA from muscle or other somatic tissues or in DNA from spermatozoa although flanking CpGs are methylated. Comparison with a candidate HTF island from the more ubiquitously expressed carbonic anhydrase II gene (CAII) shows that the CAII CpG cluster is markedly more CpG-rich than that from the strictly tissue-specific CAIII gene.

Carbonic Anhydrases↗