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Biomedical subjects

E Hogan

Publications and source records attributed to E Hogan.

18 recordsLinked to original sources
PubMed2006

Safety and efficacy of an infectious bronchitis virus used for chicken embryo vaccination.

Commercial vaccines for in ovo vaccination have not yet been developed for infectious bronchitis virus (IBV), the major coronavirus in the poultry industry. Recombinant IBVs based on the Beaudette strain expressing the Beaudette spike protein (Beau-R) or that from the virulent M41 strain (BeauR-M41(S)) were assessed for their potential as prototype vaccines for application to 18-day-old embryos. Pathogenicity was assessed by observing the effect on hatchability, and/or the production of nasal discharge and/or the effects on ciliary activity in the trachea at various time points post hatch. In contrast to commercial IBV vaccines given in ovo, the Beau-R and BeauR-M41(S) strains did not reduce hatchability or cause nasal discharge, and caused minimal damage to the ciliated epithelium of the trachea. The presence of the spike protein from a virulent virus did not increase the pathogenicity of the virus according to the criteria used. Assessment of the BeauR-M41(S) strain for efficacy showed that it protected up to 90% of chicks against challenge with virulent IB virus (M41) in a dose dependent manner. Further egg passage of the BeauR-M41(S) strain (BeauR-M41(S) EP10) did not increase its pathogenicity though it did improve its efficacy, based on serology and protection against a virulent challenge. BeauR-M41(S) EP10 was more efficacious than BeauR-M41(S) protecting more birds against virulent challenge and providing a better serological antibody response. BeauR-M41(S) EP10 induced a serological response similar to that of a commercial vaccine given at day-old though the commercial vaccine provided slightly higher efficacy. These results are promising for the development of embryo safe efficacious IBV vaccines for in ovo application.

Animals↗
PubMed1997

Centromere position in budding yeast: evidence for anaphase A.

Although general features of chromosome movement during the cell cycle are conserved among all eukaryotic cells, particular aspects vary between organisms. Understanding the basis for these variations should provide significant insight into the mechanism of chromosome movement. In this context, establishing the types of chromosome movement in the budding yeast Saccharomyces cerevisiae is important since the complexes that mediate chromosome movement (microtubule organizing centers, spindles, and kinetochores) appear much simpler in this organism than in many other eukaryotic cells. We have used fluorescence in situ hybridization to begin an analysis of chromosome movement in budding yeast. Our results demonstrate that the position of yeast centromeres changes as a function of the cell cycle in a manner similar to other eukaryotes. Centromeres are skewed to the side of the nucleus containing the spindle pole in G1; away from the poles in mid-M and clustered near the poles in anaphase and telophase. The change in position of the centromeres relative to the spindle poles supports the existence of anaphase A in budding yeast. In addition, an anaphase A-like activity independent of anaphase B was demonstrated by following the change in centromere position in telophase-arrested cells upon depolymerization and subsequent repolymerization of microtubules. The roles of anaphase A activity and G1 centromere positioning in the segregation of budding yeast chromosomes are discussed. The fluorescence in situ hybridization methodology and experimental strategies described in this study provide powerful new tools to analyze mutants defective in specific kinesin-like molecules, spindle components, and centromere factors, thereby elucidating the mechanism of chromosome movement.

Anaphase↗
PubMed1995

SMC2, a Saccharomyces cerevisiae gene essential for chromosome segregation and condensation, defines a subgroup within the SMC family.

We characterized the SMC2 (structural maintenance of chromosomes) gene that encodes a new Saccharomyces cerevisiae member of the growing family of SMC proteins. This family of evolutionary conserved proteins was introduced with identification of SMC1, a gene essential for chromosome segregation in budding yeast. The analysis of the putative structure of the Smc2 protein (Smc2p) suggests that it defines a distinct subgroup within the SMC family. This subgroup includes the ScII, XCAPE, and cut14 proteins characterized concurrently. Smc2p is a nuclear, 135-kD protein that is essential for vegetative growth. The temperature-sensitive mutation, smc2-6, confers a defect in chromosome segregation and causes partial chromosome decondensation in cells arrested in mitosis. The Smc2p molecules are able to form complexes in vivo both with Smc1p and with themselves, suggesting that they can assemble into a multimeric structure. In this study we present the first evidence that two proteins belonging to two different subgroups within the SMC family carry nonredundant biological functions. Based on genetic, biochemical, and evolutionary data we propose that the SMC family is a group of prokaryotic and eukaryotic chromosomal proteins that are likely to be one of the key components in establishing the ordered structure of chromosomes.

Amino Acid Sequence↗
PubMed1994

Chromosome condensation and sister chromatid pairing in budding yeast.

We have developed a fluorescent in situ hybridization (FISH) method to examine the structure of both natural chromosomes and small artificial chromosomes during the mitotic cycle of budding yeast. Our results suggest that the pairing of sister chromatids: (a) occurs near the centromere and at multiple places along the chromosome arm as has been observed in other eukaryotic cells; (b) is maintained in the absence of catenation between sister DNA molecules; and (c) is independent of large blocks of repetitive DNA commonly associated with heterochromatin. Condensation of a unique region of chromosome XVI and the highly repetitive ribosomal DNA (rDNA) cluster from chromosome XII were also examined in budding yeast. Interphase chromosomes were condensed 80-fold relative to B form DNA, similar to what has been observed in other eukaryotes, suggesting that the structure of interphase chromosomes may be conserved among eukaryotes. While additional condensation of budding yeast chromosomes were observed during mitosis, the level of condensation was less than that observed for human mitotic chromosomes. At most stages of the cell cycle, both unique and repetitive sequences were either condensed or decondensed. However, in cells arrested in late mitosis (M) by a cdc15 mutation, the unique DNA appeared decondensed while the repetitive rDNA region appeared condensed, suggesting that the condensation state of separate regions of the genome may be regulated differently. The ability to monitor the pairing and condensation of sister chromatids in budding yeast should facilitate the molecular analysis of these processes as well as provide two new landmarks for evaluating the function of important cell cycle regulators like p34 kinases and cyclins. Finally our FISH method provides a new tool to analyze centromeres, telomeres, and gene expression in budding yeast.

Chromatids↗
PubMed1992

Addition of extra origins of replication to a minichromosome suppresses its mitotic loss in cdc6 and cdc14 mutants of Saccharomyces cerevisiae.

Many cell division cycle (cdc) mutants of Saccharomyces cerevisiae exhibit elevated mitotic loss of pDK243, a 14-kilobase minichromosome with a centromere and one autonomous replicating sequence (ARS). Tandem copies of different ARSs were added to pDK243. The addition of these ARS clusters to pDK243 had no effect on its mitotic loss in cdc7 (protein kinase), cdc9 (DNA ligase), or cdc16 or cdc17 (DNA polymerase) mutants. However, in cdc6 and cdc14 mutants, the mitotic loss of pDK243 with an ARS cluster was suppressed by a factor of 6-8 compared to pDK243 without the cluster. This suppression was dependent upon the number of ARSs in the cluster and the integrity of the ARS consensus sequence in each ARS of the cluster. ARSs are known to be DNA replication origins. Therefore, the suppression of mini-chromosome loss by ARSs in cdc6 and cdc14 mutants suggests that these mutants are defective in the initiation of DNA replication. Since the CDC6 protein appears to act at the G1/S phase transition, the CDC6 protein may be a factor required at the beginning of S phase to initiate DNA replication at origins. In contrast, the CDC14 protein acts after mitosis. We suggest that the CDC14 protein performs a function late in the cell cycle that may be required for efficient initiation of DNA replication during S phase of the next cell cycle.

Cell Cycle↗
PubMed1991

The CDC20 gene product of Saccharomyces cerevisiae, a beta-transducin homolog, is required for a subset of microtubule-dependent cellular processes.

Previous analysis of cdc20 mutants of the yeast Saccharomyces cerevisiae suggests that the CDC20 gene product (Cdc20p) is required for two microtubule-dependent processes, nuclear movements prior to anaphase and chromosome separation. Here we report that cdc20 mutants are defective for a third microtubule-mediated event, nuclear fusion during mating of G1 cells, but appear normal for a fourth microtubule-dependent process, nuclear migration after DNA replication. Therefore, Cdc20p is required for a subset of microtubule-dependent processes and functions at multiple stages in the life cycle. Consistent with this interpretation, we find that cdc20 cells arrested by alpha-factor or at the restrictive temperature accumulate anomalous microtubule structures, as detected by indirect immunofluorescence. The anomalous microtubule staining patterns are due to cdc20 because intragenic revertants that revert the temperature sensitivity have normal microtubule morphologies. cdc20 mutants have a sevenfold increase in the intensity of antitubulin fluorescence in intranuclear spindles compared with spindles from wild-type cells, yet the total amount of tubulin is indistinguishable by Western immunoblot analysis. This result suggests that Cdc20p modulates microtubule structure in wild-type cells either by promoting microtubule disassembly or by altering the surface of the microtubules. Finally, we cloned and sequenced CDC20 and show that it encodes a member of a family of proteins that share homology to the beta subunit of transducin.

Amino Acid Sequence↗
PubMed1990

Mitotic transmission of artificial chromosomes in cdc mutants of the yeast, Saccharomyces cerevisiae.

In the yeast, Saccharomyces cerevisiae, cell division cycle (CDC) genes have been identified whose products are required for the execution of different steps in the cell cycle. In this study, the fidelity of transmission of a 14-kb circular minichromosome and a 155-kb linear chromosome fragment was examined in cell divisions where specific CDC products were temporarily inactivated with either inhibitors, or temperature sensitive mutations in the appropriate CDC gene. All of the cdc mutants previously shown to induce loss of endogenous linear chromosomes also induced loss of a circular minichromosome and a large linear chromosome fragment in our study (either 1:0 or 2:0 loss events). Therefore, the efficient transmission of these artificial chromosomes depends upon the same trans factors that are required for the efficient transmission of endogenous chromosomes. In a subset of cdc mutants (cdc6, cdc7 and cdc16), the rate of minichromosome loss was significantly greater than the rate of loss of the linear chromosome fragment, suggesting that a structural feature of the minichromosome (nucleotide content, length or topology) makes the minichromosome hypersensitive to the level of function of these CDC gene products. In another subset of cdc mutants (cdc7 and cdc17), the relative rate of 1:0 events to 2:0 events differed for the minichromosome and chromosome fragment, suggesting that the type of chromosome loss event observed in these mutants was dependent upon chromosome structure. Finally, we show that 2:0 events for the minichromosome can occur by both a RAD52 dependent and RAD52 independent mechanism. These results are discussed in the context of the molecular functions of the CDC products.

Cell Division↗
PubMed1988

pH-sensitive activation of the intracellular-pH regulation system in squid axons by ATP-gamma-S.

The regulation of intracellular pH (pHi) is essential for normal cell function, and controlled changes in pHi may play a central role in cell activation. Sodium-dependent Cl-HCO3 exchange is the dominant mechanism of pHi regulation in the invertebrate cells examined, and also occurs in mammalian cells. The transporter extrudes acid from the cell by exchanging extracellular Na+ and HCO3- (ref. 9) (or a related species) for intracellular Cl- (refs 3, 4). It is blocked by the stilbene derivatives DIDS (4,4'-diisothiocyano-stilbene-2,2'-disulphonate, ref. 10) and SITS (4-acetamido-4'-isothiocyano-stilbene-2,2'-disulphonate, ref. 3), and has a stoichiometry of two intracellular H+ neutralized for each Na+ taken up and each Cl- extruded by the axon. Because the inwardly-directed Na+ concentration gradient is sufficiently large to energize both the HCO3- influx and Cl- efflux, this electroneutral exchanger could be a classic secondary active transporter, thermodynamically independent of ATP hydrolysis. However, at least in the squid axon, the exchanger has an absolute requirement for ATP (ref. 3). Thus, a major unresolved issue is whether this Na-dependent Cl-HCO3 exchanger stoichiometrically hydrolyses ATP (the pump hypothesis), or whether ATP activates the transporter by a mechanism such as phosphorylation or simple binding (the activation hypothesis). We have now explored the role of ATP in pHi regulation by dialysing axons with the ATP analogue ATP-gamma-S. In many systems, ATP-gamma-S is an acceptable substrate for protein kinases, whereas the resulting thiophosphorylated proteins are not as readily hydrolysed by phosphatases as are phosphorylated proteins. Our results rule out the pump hypothesis, and show that the basis of the axon's ATP requirement is the pH-dependent activation (by, for instance, phosphorylation or ATP binding) of the exchanger itself, or of an essential activator.

Adenosine Triphosphate↗
PubMed1988

Genomic organization of major sperm protein genes and pseudogenes in the nematode Caenorhabditis elegans.

The major sperm proteins (MSPs) are a family of closely related, small, basic proteins comprising 15% of the protein in Caenorhabditis elegans sperm. They are encoded by a multigene family of more than 50 genes, including many pseudogenes. MSP gene transcription occurs only in late primary spermatocytes. In order to study the genomic organization of transcribed MSP genes, probes specific for the 3' untranslated regions of sequenced cDNA clones were used to isolate transcribed genes from genomic libraries. These and other clones of MSP genes were located in overlapping cosmid clones by DNA fingerprinting. These cosmids were aligned with the genetic map by overlap with known genes or in-situ hybridization to chromosomes. Of 40 MSP genes identified, 37, including all those known to be transcribed, are organized into six clusters composed of 3 to 13 genes each. Within each cluster, MSP genes are not in tandem but are separated by at least several thousand bases of DNA. Pseudogenes are interspersed among functional genes. Genes with similar 3' untranslated sequences are in the same cluster. The six MSP clusters are confined to only three chromosomal loci; one on the left arm of chromosome II and two near the middle of chromosome IV. Additional sperm-specific genes are located in one cluster of MSP genes on chromosome IV. The multiplicity of MSP genes appears to be a mechanism for enhancing MSP synthesis in spermatocytes, and the loose clustering of genes could be a result of the mechanism of gene duplication or could play a role in regulation.

Animals↗
PubMed1987

Proton magnetic resonance spectroscopy of leech muscle and nervous system.

1. Proton nuclear magnetic resonance spectroscopy (1H NMR) was used to measure the major intracellular metabolites in perchloric acid extracts of the Macrobdella decora muscle and nervous systems and the Oryctolagus cuniculus cerebrum. 2. Acetate, alanine, choline, glutamate, inositol, and lactate were assigned in the spectrum of leech ventral cord, leech muscle, and rabbit cerebrum. 3. Hirudonine and propionate were clearly observed only in the spectrum of leech muscle. 4. Creatine, N-acetyl aspartate, gamma aminobutyric acid, aspartate, and taurine, distinctive components of spectra of the mammalian cerebrum, were not seen in the invertebrate spectra. 5. 1H NMR spectroscopy provides a simple and rapid means of characterizing the major organic metabolites found in leech muscle and nervous tissues.

3-Hydroxybutyric Acid↗
PubMed1986

Monoclonal antibodies that recognize a polypeptide antigenic determinant shared by multiple Caenorhabditis elegans sperm-specific proteins.

Four monoclonal antibodies that are directed against antigens present in sperm and absent from other worm tissues were characterized. Antibody TR20 is directed against the major sperm proteins, a family of small, abundant, cytoplasmic proteins that have been previously described (Klass, M. R., and D. Hirsh, 1981, Dev. Biol., 84:299-312; Burke, D. J., and S. Ward, 1983, J. Mol. Biol., 171:1-29). Three other antibodies, SP56, SP150, and TR11, are all directed against the same set of minor sperm polypeptides that range in size from 29 to 215 kD. More than eight different sperm polypeptides are antigenic by both immunotransfer and immunoprecipitation assays. The three antibodies are different immunoglobulin subclasses, yet they compete with each other for antigen binding so they are directed against the same antigenic determinant on the multiple sperm proteins. This antigenic determinant is sensitive to any of six different proteases, is insensitive to periodate oxidation or N-glycanase digestion, and is detectable on a polypeptide synthesized in vitro. Therefore, the antigenic determinant resides in the polypeptide chain. However, peptide fragments of the proteins are not antigenic, thus the determinant is likely to be dependent on polypeptide conformation. The antigenic determinant shared by these proteins could represent a common structural feature of importance to the localization or cellular specificity of these proteins.

Animals↗
PubMed1983

The initiation of spermiogenesis in the nematode Caenorhabditis elegans.

Spermiogenesis in nematodes involves the activation of sessile spherical spermatids to motile bipolar amoeboid spermatozoa. In Caenorhabditis elegans males spermiogenesis is normally induced by copulation. Spermatids transferred to hermaphrodites as well as some of those left behind in the male become spermatozoa a few minutes after mating. Spermiogenesis can also be induced in vitro by the ionophore monensin (G.A. Nelson and S. Ward, 1980, Cell 19, 457-464) and by weak bases such as triethanolamine. Both triethanolamine and monensin cause a rapid increase in intracellular pH from 7.1 to 7.5 or 8.0. This pH increase precedes the subsequent morphological events of spermiogenesis. Triethanolamine or monensin must be present throughout spermiogenesis for all cells to form pseudopods, but once pseudopods are formed the inducers are unnecessary for subsequent motility. The pH induced spermiogenesis is inhibited by drugs that block mitochondria or glycolysis. Protease treatment can also induce spermiogenesis without increasing intracellular pH, apparently bypassing the pH-dependent steps in activation and the requirement for glycolysis. These results show that the initiation of spermiogenesis in C. elegans, like some steps in egg activation and the initiation of sea urchin sperm motility, can be induced by an increase in intracellular pH, but this pH change can be bypassed by proteolysis.

Animals↗
PubMed1980

Utility of an on-line computer system in a clinical setting for nutrient intake analyses.

The present report describes a four-week trial at the Victoria General Hospital, Halifax, to determine the effect of access to a computer system on nutrient intake analyses, the problems in implementation of the system, and the response of the staff dietitians. An on-line system was obtained from the Ohio State University Hospitals and modified to reflect Canadian food and nutrient content, and to include some additional program features. Twelve hundred and eighty-six foods were characterized according to 15 food components, the system was responsive to nine interactive commands, and there were nine possible print-out formats of calculations. The results of the trial reinforced the expectation of greatly increased work efficiency, however, the system was not utilized to its potential. Some technical problems were largely overcome within the first week of operation. However, needs relating to the extensiveness of the food and nutrient data base, and the methodologies and time for obtaining and evaluating individual dietary intake information prevented maximum efficiency. The dietitians liked the comprehensiveness and detail of the system, however, routine dietary intake collections were too time-consuming for their present work schedules.

Adult↗
PubMed1978

Same/different concept learning in the pigeon: the effect of negative instances and prior adaptation to transfer stimuli.

Pigeons were trained on a matching-to-sample or oddity-from-sample task with shapes (circle and plus). Half of each group was exposed to "negative instance" trials i.e., for matching birds, neither comparison key matched the sample, and for oddity birds both comparison keys matched the sample. When all birds were transferred to a new task involving colors (red and green), nonshifted birds (transferred from matching to matching, or oddity to oddity) performed significantly better than shifted birds (transferred from matching to oddity, or oddity to matching), but only if they had experienced negative instances of the training concept. When all birds were exposed to negative instances of the transfer task and then transferred to a new color task (yellow and blue), dramatic transfer effects were observed. The effect of pre-exposure to the yellow and blue colors, in order to reduce transfer-stimulus novelty, had a minor effect on transfer.

Journal Article↗
PubMed1977

Evidence for defective incorporation of proteins in myelin of the quaking mutant mouse.

The defect in myelinogenesis present in the Quaking mutant mouse was investigated using a double radioisotope technique for comparing the incorporation of amino acid into myelin proteins of normal and mutant mice. Quaking mice and littermate controls received intracranial injections of 150 muCi [3H]glycine and 25 muCi of [14C]glycine respectively. After 2 h their brains were combined and jointly processed to obtain subcellular fractions. The 3H/14C ratio for the myelin subfraction was 1.88 as compared to a 3H/14C ratio of 3.0 for the other subfractions, indicating a 40% decrease in glycine incorporation into myelin of Quaking mice. Myelin proteins were separated by discontinuous gel electrophoresis in the presence of sodium dodecyl sulfate (SDS) and the 3H/14C ratios determined in each gel slice. In contrast to the microsomal subfractions which gave a 3H/14C ratio of 2.6 across the gel, the 3H/14C ratio of myelin showed large variations with values ranging from 0.54 for proteolipid protein to 2.0 for some of the high molecular weight proteins. During development, the Quaking mutant exhibited a preferential depression in glycine incorporation into proteolipid protein in 18-day-old mice, while in older animals (32-54 days) the fast migrating basic protein, as well as the proteolipid protein, was labeled to a significantly lesser extent.

Age Factors↗