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Biomedical subjects

J Yon

Publications and source records attributed to J Yon.

At least 19 recordsLinked to original sources
PubMed1999

Enovin, a member of the glial cell-line-derived neurotrophic factor (GDNF) family with growth promoting activity on neuronal cells. Existence and tissue-specific expression of different splice variants.

Glial cell-line-derived neurotrophic factor (GDNF), neurturin and persephin are neurotrophic factors involved in neuroneal differentiation, development and maintenance. They act on different types of neuroneal cells and signal through a receptor complex composed of a specific ligand-binding subunit of the GDNF family receptor alpha (GFRalpha) family together with a common signaling partner, the cRET protein tyrosine kinase. We describe the molecular cloning, expression, chromosomal localization and functional characterization of enovin, a fourth GDNF family member almost identical to the recently described artemin. We show the occurence in most tissues of several differently spliced mRNA variants for enovin, of which only two are able to translate into functional enovin protein. Some tissues seem to express only nonfunctional transcripts. These observations may underlie a complex transcriptional regulation pattern. Enovin mRNA expression is detectable in all adult and fetal human tissues examined, but expression levels are highest in peripheral tissues including prostate, placenta, pancreas, heart and kidney. This tissue distribution pattern is in accordance with that of GFRalpha-3, which here is shown to be the preferred ligand-binding receptor for enovin (Kd = 3.1 nM). The human enovin gene is localized on chromosome 1, region p31.3-p32. In vitro, enovin stimulates neurite outgrowth and counteracts taxol-induced neurotoxicity in staurosporine-differentiated SH-SY5Y human neuroblastoma cells. The peripheral expression pattern of enovin and its receptor together with its effects on neuroneal cells suggest that enovin might be useful for the treatment of neurodegenerative diseases in general and peripheral neuropathies in particular.

Adult↗
PubMed1998

A human homologue of the Schizosaccharomyces pombe rad1+ checkpoint gene encodes an exonuclease.

In the fission yeast Schizosaccharomyces pombe the rad1(+) gene is required for both the DNA damage-dependent and the DNA replication-dependent cell cycle checkpoints. We have identified a human homologue of the S. pombe rad1(+) gene, designated Hrad1, as well as a mouse homologue: Mrad1. Two Hrad1 alternative splice variants with different open reading frames have been identified; one codes for a long form, Hrad1A, and the other encodes a short form because of N-terminal truncation, Hrad1B. Hrad1A has 60% identity to the S. pombe rad1+ sequence at the DNA level and 49% identity and 72% similarity at the amino acid level. Northern blot analysis indicates elevated levels of expression in testis and cancer cell lines. Chromosomal localization by fluorescence in situ hybridization indicates that Hrad1 is located on chromosome 5p13. 2-13.3. This region is subject to loss of heterozygosity in several human cancers. Hrad1 also shares homology with the Saccharomyces cerevisiae RAD17 and Ustilago maydis REC1 proteins. REC1 has previously been characterized as a 3' --> 5' exonuclease with a C-terminal domain essential for cell cycle checkpoint function. We have expressed and purified polyhistidine-tagged fusions of Hrad1A and Hrad1B and show that HisHrad1A has 3' --> 5' exonuclease activity, whereas HisHrad1B lacks such activity. The biological functions of the two proteins remain to be determined.

Alternative Splicing↗
PubMed1994

Molecular and biological characterization of simian immunodeficiency virus macaque strain 32H proviral clones containing nef size variants.

The proviral genome of the 32H reisolate of simian immunodeficiency of macaques (SIVmac32H) has been cloned and sequenced. Including both long terminal repeats, it is 10277 base pairs in length and contains open reading frames for all known SIV genes (gag, pol, vif, vpx, vpr, tat, rev, env and nef). This is the first report of an infectious SIVmac molecular clone which contains no premature termination codons. Three molecular clones of SIVmac32H have been constructed differing in sequence only within their last 1.2 kb. Two of the molecular clones, SIVmac32H(pJ5) and SIVmac32H (pC8), differ in the nef coding region by an in-frame deletion of four amino acids in pC8 and two conservative amino acid changes; other nucleotide changes in the 3' LTR were not associated with known functionally critical motifs. The third clone, SIVmac32H(pB1), contains the last 1.2 kb of the SIVmac251 clone pBK28. The biological properties of virus produced after electroporation of these clones into C8166 cells has been assessed by infection of rhesus and cynomolgus macaques, time to seroconversion and by induction of cytopathic effects upon co-cultivation of infected rhesus peripheral blood lymphocytes with C8166 cells. The viruses obtained from these clones have identical growth kinetics in vitro but differ in their ability to persist in macaques. Macaques infected with pJ5 derived virus remain viraemic longer than macaques infected with pC8-derived virus. PCR analysis of circulating provirus indicates that the nef gene evolved over time in pJ5 virus-infected macaques, whereas late in infection in pC8 virus-infected macaques the nef gene remained invariant in sequence. These results support the observation that a nef deletion mutant of SIVmac239 lost its pathogenic potential and resulted in low-level viraemia when rhesus macaques were infected. Virus challenge pools for vaccine studies have been prepared for pJ5 using both human and monkey cell substrates and these stocks have been titrated both in vitro and in vivo. Virus has also been prepared from pC8 and titrated in vitro. This virus pool is being assessed as an attenuated live-virus vaccine in macaques. Since only virus originating from the SIVmac239 molecular clone is known to cause AIDS-like symptoms in rhesus macaques consistently, the SIVmac32H molecular clones should tell us more about which viral sequence features are important for the pathogenesis of AIDS.

Amino Acid Sequence↗
PubMed1993

The organization and conservation of the human Surfeit gene cluster and its localization telomeric to the c-abl and can proto-oncogenes at chromosome band 9q34.1.

The mouse Surfeit locus contains an unusually tight cluster of six housekeeping genes (Surf-1 to -6) which are unrelated by sequence homology. Using a mouse Surfeit locus probe, a 16 kb clone has been isolated which contains the human Surf-1 and Surf-3 genes and regions of the human Surf-2 and Surf-5 genes. The organization and juxtaposition of these human Surfeit locus genes are the same as found in the mouse. Using the human clone as a biotinylated probe for fluorescence in situ hybridization (FISH) we have confirmed the location of the human Surfeit locus to chromosome band 9q34. Metaphase spreads of human chronic myeloid leukemic cells containing the t(9;22)(q34;q11) translocation involving The c-abl gene at 9q34.1 an acute nonlymphocytic leukemic cells containing the t(6;9)(q34;p23) translocation involving the can gene at 9q34.1 were analyzed by FISH using the human Surfeit clone as a probe. These analyses locate the human Surfeit locus telomeric to the c-abl and can genes at chromosome band 9q34.1.

Animals↗
PubMed1992

Conservation of the organization of five tightly clustered genes over 600 million years of divergent evolution.

The organization of the mouse surfeit locus is unusual in that it contains six housekeeping genes (Surf-1-Surf-6), which are unrelated by sequence homology, in the tightest mammalian gene cluster thus far described. A maximum of only 73 base pairs separates any two of the four well-characterized genes, and two of the genes overlap at their 3' ends. The direction of transcription of each of the five surfeit genes, Surf-1-Surf-5, alternates with respect to that of its neighbor, suggesting cis-interaction or coregulation between the genes by mechanisms such as the sharing of regulatory elements and/or antisense regulation. The Surf-3 gene has been identified as encoding the ribosomal protein L7a (Rpl7a). We have used the high conservation of the Rpl7a gene to clone the chicken gene and surrounding genomic DNA. The tight clustering and juxtaposition of at least five of the surfeit genes (Surf-1-Surf-5) and their associated CpG-rich islands have been found to be conserved over the 600 million years of divergent evolution that separates birds and mammals. This strongly suggests that the surfeit locus represents a different form of gene cluster in which gene organization may play both a positive and negative regulatory role in gene expression possibly via cis-interactions between the closely spaced genes.

Amino Acid Sequence↗
PubMed1992

Stimulation of specific immune responses to simian immunodeficiency virus using chimeric hepatitis B core antigen particles.

Subunit approaches to vaccines against viral diseases have resulted in the development of a number of methods for presentation of defined epitopes to the immune system. We have exploited a highly immunogenic presentation system based on hepatitis B core antigen (HBcAg) particles to produce a number of candidate vaccines against simian immunodeficiency virus (SIV). Recombinant particles have been produced in bacteria which carry multiple copies of defined or predicted neutralizing epitopes of SIV at a number of different sites within the particle. In parallel, a number of synthetic peptide-based SIV vaccines have been produced based on homology to reported neutralizing epitopes in human immunodeficiency virus. Although potent immune responses were elicited against both particulate and peptide forms of the antigen, neutralizing antibodies were not induced as judged by available assays.

Acquired Immunodeficiency Syndrome↗
PubMed1991

The organization and expression of the human L7a ribosomal protein gene.

The genomic structure and sequence of the human ribosomal protein L7a (rpL7a) and its transcriptional start sites have been determined. The gene contains eight exons and seven introns spread over 3179 bp. The human rpL7a gene is similar to other mammalian ribosomal protein genes in containing a short first exon, a short 5' untranslated leader and its transcriptional start sites at C residues embedded in a polypyrimidine tract. The 25 bp polypyrimidine tract containing the two human rpL7a start sites is the longest polypyrimide tract so far observed for a mammalian ribosomal protein gene.

Base Sequence↗
PubMed1991

Structural and functional characterization of Factor VIII-delta II, a new recombinant Factor VIII lacking most of the B-domain.

A recombinant Factor VIII (Factor VIII-delta II) consists of a unique polypeptide chain of 165 kDa deleted from the major part of the B-domain and from the cleavage site at Arg-1648-Glu-1649 found in plasma-derived Factor VIII. It was expressed in mammalian cells in serum-free medium containing von Willebrand factor and purified by a one-step immunopurification. The recombinant Factor VIII was characterized as a single active peak when subjected to f.p.l.c., in contrast with the plasma-derived molecule. Its coagulant activity was decreased in the presence of EDTA, suggesting that a bivalent ion is required, as for plasma-derived Factor VIII. The activation by thrombin and the inactivation by activated protein C were studied and the resulting molecular forms were analysed by f.p.l.c. and SDS/PAGE. The results clearly demonstrate that, despite the structural differences between plasma-derived and recombinant Factor VIII, activation and inactivation of Factor VIII-delta II generate proteolysed complexes similar to that described for plasma-derived Factor VIII. Thus this deleted recombinant Factor VIII, which is processed similarly to plasma-derived Factor VIII, should be normally integrated in the regulation system of Factor X activation in the blood-coagulation cascade.

Amino Acid Sequence↗
PubMed1991

The organization and expression of the Saccharomyces cerevisiae L4 ribosomal protein genes and their identification as the homologues of the mammalian ribosomal protein gene L7a.

A cDNA for the mouse ribosomal protein (rp) L7a, formerly called Surf-3, was used as a probe to isolate two homologous genes from Saccharomyces cerevisiae. The two yeast genes (L4-1 and L4-2) were identified as encoding S. cerevisiae L4 by 2D gel analysis of the product of the in vitro translation of hybrid-selected mRNA and additionally by direct amino acid sequencing. The DNA sequences of the two yeast genes were highly homologous (95%) over the 771 bp that encode the 256 amino acids of the coding regions but showed little homology outside the coding region. L4-1 differed from L4-2 by 7 out of the 256 amino acids in the coding region, which is the greatest divergence between the products of any two duplicated yeast ribosomal protein genes so far reported. There is strong homology between the mouse rpL7a/Surf-3 and the yeast L4 genes -57% at the nucleic acid level and also 57% at the amino acid level (though some regions reach as much as 80-90% homology). While most yeast ribosomal protein genes contain an intron in their 5' region both L4-1 and L4-2 are intronless. The mRNAs derived from each yeast gene contained heterogenous 5' and 3' ends but in each case the untranslated leaders were short. The L4-1 mRNA was found to be much more abundant than the L4-2 mRNA as assessed by cDNA and transcription analyses. Yeast cells containing a disruption of the L4-1 gene formed much smaller colonies than either wild-type or disrupted L4-2 strains. Disruption of both L4 genes is a lethal event, probably due to an inability to produce functional ribosomes.

Amino Acid Sequence↗
PubMed1989

Ribosomal protein L7a is encoded by a gene (Surf-3) within the tightly clustered mouse surfeit locus.

The mouse Surfeit locus, which contains a cluster of at least four genes (Surf-1 to Surf-4), is unusual in that adjacent genes are separated by no more than 73 base pairs (bp). The heterogeneous 5' ends of Surf-1 and Surf-2 are separated by only 15 to 73 bp, the 3' ends of Surf-1 and Surf-3 are only 70 bp apart, and the 3' ends of Surf-2 and Surf-4 overlap by 133 bp. This very tight clustering suggests a cis interaction between adjacent Surfeit genes. The Surf-3 gene (which could code for a basic polypeptide of 266 amino acids) is a highly expressed member of a pseudogene-containing multigene family. By use of an anti-peptide serum (against the C-terminal nine amino acids of the putative Surf-3 protein) for immunofluorescence and immunoblotting of mouse cell components and by in vitro translation of Surf-3 cDNA hybrid-selected mRNA, the Surf-3 gene product was identified as a 32-kilodalton ribosomal protein located in the 60S ribosomal subunit. From its subunit location, gel migration, and homology with a limited rat ribosomal peptide sequence, the Surf-3 gene was shown to encode the mouse L7a ribosomal protein. The Surf-3 gene is highly conserved through evolution and was detected by nucleic acid hybridization as existing in multiple copies (multigene families) in other mammals and as one or a few copies in birds, Xenopus, Drosophila, and Schizosaccharomyces pombe. The Surf-3 C-terminal anti-peptide serum detects a 32-kilodalton protein in other mammals, birds, and Xenopus but not in Drosophila and S. pombe. The possible effect of interaction of the Surf-3 ribosomal protein gene with adjacent genes in the Surfeit locus at the transcriptional or posttranscriptional level or both levels is discussed.

Animals↗
PubMed1988

The mouse surfeit locus contains a very tight cluster of four "housekeeping" genes that is conserved through evolution.

The four identified alternating transcription units (Surf-1 to Surf-4) in the mouse surfeit locus are very tightly clustered, no two neighboring units being separated by more than 73 base pairs and the Surf-2 and Surf-4 transcription units overlapping by 133 base pairs at their 3' ends. All four surfeit genes, which are unrelated by sequence similarity, were found to have the properties of "housekeeping" genes, being expressed in a variety of differentiated mouse cell lines and containing unmethylated CpG-rich islands in the vicinity of their 5' ends. The unusual organization of the four surfeit genes was found not to be unique to the mouse: the same juxtaposition of the genes was found to be conserved in a number of different mammals, including humans. The four human surfeit genes were also found to be transcriptionally active.

Animals↗
PubMed1976

Binding of nucleotides ATP and ADP to sarcoplasmic reticulm: study by rate of dialysis.

Binding of the nucleotides ATP and ADP by preparations of sarcoplasmic reticulum was investigated by the method of flow dialysis. For ATP, experimental data could not be analyzed directly in terms of binding since a significant though small amount of hydrolysis could be observed even in presence of EDTA. ADP binding could be analyzed and gave a dissociation constant of 10-20 muM at neutral pH, and a stoichiometry of 0.35 - 0.45 per mole ATPase. The possible significance of this stoichiometry is discussed. Similar experiments were performed after ethoxyformylation of sarcoplasmic reticulum which inhibits the Ca2+ dependnet ATPase activity. The results confirmed the inhibition of ATP hydrolysis and pointed to a considerably reduced affinity for nucleotides. The method based on the measurement of dialysis rates is convenient, and accurate enough to detect the effects of chemical modification on sarcoplasmic reticulum membranes.

Adenosine Diphosphate↗
PubMed1975

Resonance Raman spectroscopic studies of the interactions between trypsin and a competitive inhibitor.

Raman spectroscopy was used to study the interactions between bovine trypsin and a competitive inhibitor. For this purpose, a chromophoric substrate analogue, 4-amidino-4'-dimethylamine azobenzene, was synthesized. This compound competitively inhibits the enzyme with a 1:1 stoichiometry and an inhibition constant Ki of 2.3 muM at pH 6.08 and 15 degrees. Resonance Raman spectra in aqueous solution of free or enzyme-bound inhibitor were analyzed. The main spectral changes observed upon enzyme-inhibitor complex formation were changes in the relative intensities of four bands (1171, 1206, 1315, 1608 cm-1) while no large frequency shifts occurred. The binding of the inhibitor molecule to the enzyme did not induce a twisting of the phenyl groups around the N=N bond. Some modifications of the band widths are interpreted in terms of a restriction of rotational motions in the inhibitor molecule. The possible involvement of specific interactions between trypsin and the benzamidinium ion part of the inhibitor molecule is discussed.

Azo Compounds↗
PubMed1975

An uncontrolled comparison of treatments for achalasia.

A series of 78 patients with achalasia, seen during a 10-year period, was like those reported by others with regard to age at onset, nature of symptoms, and duration of symptoms. Analysis of the results of 5 different treatment modalities administered allows an uncontrolled comparison heretofore not available from an institution at which no one treatment was favored over all others. Those who were not treated or who received only anticholinergic medication did not become asymptomatic. Those treated by single or repeated bougienage were not improved for more than a few weeks and suffered a 6% incidence of esophageal perforation. Forty-six per cent of those treated by a single pneumatic dilatation were asymptomatic for 1 year or more. Although esophagocardiomyotomy provided symptomatic relief for 1 year or more in 85%, there was a 25% incidence of gastroesophageal reflux.

Adolescent↗