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R Nash

Publications and source records attributed to R Nash.

At least 19 recordsLinked to original sources
PubMed1992

The Cln3-Cdc28 kinase complex of S. cerevisiae is regulated by proteolysis and phosphorylation.

In Saccharomyces cerevisiae, several of the proteins involved in the Start decision have been identified; these include the Cdc28 protein kinase and three cyclin-like proteins, Cln1, Cln2 and Cln3. We find that Cln3 is a very unstable, low abundance protein. In contrast, the truncated Cln3-1 protein is stable, suggesting that the PEST-rich C-terminal third of Cln3 is necessary for rapid turnover. Cln3 associates with Cdc28 to form an active kinase complex that phosphorylates Cln3 itself and a co-precipitated substrate of 45 kDa. The cdc34-2 allele, which encodes a defective ubiquitin conjugating enzyme, dramatically increases the kinase activity associated with Cln3, but does not affect the half-life of Cln3. The Cln--Cdc28 complex is inactivated by treatment with non-specific phosphatases; prolonged incubation with ATP restores kinase activity to the dephosphorylated kinase complex. It is thus possible that phosphate residues essential for Cln-Cdc28 kinase activity are added autocatalytically. The multiple post-translational controls on Cln3 activity may help Cln3 tether division to growth.

Base Sequence↗
PubMed1992

Isoelectric focusing patterns of beta-lactamases in the rapidly growing mycobacteria.

beta-lactamases from 259 strains of rapidly growing mycobacteria that included the third biovariant complex of Mycobacterium fortuitum, M. peregrinum, M. abscessus, M. chelonae, the M. chelonae-like organisms (MCLO), and M. smegmatis were analyzed by isoelectric focusing (IEF). All isolates produced acidic beta-lactamases with major band isoelectric points (pIs) between 4.4 and 6.0. Each of the 6 taxonomic groups exhibited 1 or 2 characteristic beta-lactamase IEF patterns. Heterogeneity among IEF patterns was evident in 5 of the 6 groups, however, and was greatest among the third biovariant complex of M. fortuitum. beta-lactamase patterns correlated with previously identified taxonomic subgroups of M. smegmatis and the third biovariant complex of M. fortuitum. beta-lactamase IEF analysis of MCLO strains isolated from two outbreaks demonstrated its possible usefulness for epidemiologic evaluation.

Animals↗
PubMed1992

Effect of endotoxin on serum granulocyte and granulocyte-macrophage colony-stimulating factor levels in dogs.

The biologic effects of endotoxin are attributed to the release of several cytokines, including interleukin-1, interleukin-6, tumor necrosis factor, and the colony-stimulating factors. To investigate the mechanism of endotoxin-induced neutrophilia in dogs, several cell lines known to proliferate selectively in response to recombinant human colony-stimulating factors were examined to determine their responses to recombinant canine granulocyte colony-stimulating factor (rcG-CSF) or recombinant canine granulocyte-macrophage colony-stimulating factor (rcGM-CSF). The murine cell line NFS-60 was found to respond well to rcG-CSF and the human cell line TALL-101 to rcGM-CSF, and these responses were neutralized by antibodies to these recombinant proteins. These bioassays were then used to determine G-CSF and GM-CSF levels in dogs after intravenous endotoxin administration. G-CSF levels increased by 2 h, peaked at 4 h, and had not returned to normal by 24 h after endotoxin. In contrast, GM-CSF was not detectible before or after endotoxin administration.

Animals↗
PubMed1992

[Prevalence of hepatitis C virus antibody in chronic HBsAg-negative non alcoholic hepatopathy].

The presence of antibody to hepatitis C virus was determined in 316 HBsAg-negative patients with non-alcoholic chronic hepatitis who did not receive any blood transfusion once the diagnosis was made. A titre of antinuclear antibodies of 1/40 or lower was found in 18 patients. Persistent chronic hepatitis was present in 21 patients, active chronic hepatitis in 145, hepatic cirrhosis in 128, and hepatocarcinoma in 22 patients. One hundred and three patients had previously received blood transfusion, 76 had undergone previous surgery without transfusion, a clinical episode of hepatitis could be traced in 14, 13 patients were drug addicts (all of them HIV negative), 1 patient had received multiples injections, another had been treated with acupuncture, and 108 patients were free of any of the above. Anti-HCV was present in 76.6% of patients; a significantly higher proportion (87.4%) was found among patients who had received blood transfusion than in patients with previous surgery (72.4%) (p = 0.012), clinical hepatitis (57.1%), or without previous hepatic disease (70.3%) (p = 0.003). The incidence of anti-HCV was lower among cirrhotics (70.3%) than in patients with active chronic hepatitis (84.1%) (p = 0.006); in contrast, previous blood transfusion was significantly higher (p = 0.001) among the latter (40.7%) than in cirrhotics (21.9%). The incidence of anti-HCV was similar among patients with (78.6%) and without (75.8%) type B infection. Our results suggest that infection with virus C may account for a high proportion of non-alcoholic non-B chronic hepatitis.

Adolescent↗
PubMed1990

Serial cytogenetic studies showing persistence of original clone in Hodgkin's disease.

A case of Hodgkin's disease is described in which cytogenetic studies were performed at intervals for 5 years. Clonally related chromosomally rearranged cells persisted during that time, with emergence of more complex karyotypes in the terminal phase of the disease. The chromosome findings are discussed in relation to the limited data available on banded karyotypes in Hodgkin's disease.

Bone Marrow Cells↗
PubMed1989

Estimating the size of the dividing stem cell pool after allogeneic bone marrow transplantation.

Currently the problem of estimation of the number of pluripotent stem cells reconstituting marrow grafts following bone marrow transplantation, or studies looking at questions of clonal dominance in hematopoietic cell populations, rely on indirect measurement and a simple application of the formula for the sampling variation of a binomial proportion. This approach, from a statistical viewpoint, can be seen to be flawed. It is very easily remedied though and only requires appropriate use of variance stabilizing transformations. These lead to a very simple estimator for the number of hematopoietic stem cells involved in repopulating the marrow and require little in the way of additional calculation. We give the distribution theory for this estimator as well as simple approximations for practical application. As an illustration we rework data recently gathered to address the question as to whether or not reconstitution of marrow grafts in the clinical setting is oligoclonal.

Bone Marrow Transplantation↗
PubMed1989

Stimulation of canine hematopoiesis by recombinant human granulocyte-macrophage colony-stimulating factor.

The effect of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) on canine hematopoiesis was evaluated. rhGM-CSF stimulated granulocyte-macrophage colony formation of canine marrow depleted of accessory cells up to tenfold. Stimulation of colony formation was abrogated by anti-rhGM-CSF antiserum or heat inactivation. rhGM-CSF also stimulated in vivo canine hematopoiesis both when given as continuous i.v. infusion and as intermittent s.c. injections. Neutrophil, monocyte, and lymphocyte counts were increased three- to eightfold above controls, whereas values for eosinophils, reticulocytes, and hematocrits were not changed. Bone marrow histology after 2 weeks of treatment with rhGM-CSF showed hypercellularity with myeloid hyperplasia and left-shifted granulocytopoiesis. After discontinuation of rhGM-CSF, peripheral leukocyte counts returned to control level within 3-7 days. Platelet counts decreased rapidly after starting rhGM-CSF, to 5000-15,000 platelets/mm3, and increased within 24 h after stopping rhGM-CSF treatment, whereas marrow histology after 2 weeks of rhGM-CSF application showed the normal number and morphology of megakaryocytes.

Animals↗
PubMed1989

Improved retroviral transfer of genes into canine hematopoietic progenitor cells kept in long-term marrow culture.

Amphotropic helper-free retroviral vectors containing either the bacterial neomycin phosphotransferase gene (NEO) or a mutant dihydrofolate reductase gene (DHFR*) were used to infect canine hematopoietic progenitor cells. In previous experiments, successful transfer and expression of both genes in canine CFU-GM were achieved after 24-hour cocultivation with virus-producing cells. The average rate of gene expression was 10% (6% to 16%) as measured by the number of CFU-GM resistant to either the aminoglycoside G418 or methotrexate. In an attempt to increase the efficiency of gene transfer, marrow was cocultured for 24 hours with either NEO or DHFR* virus-producing packaging cells and then kept in long-term marrow culture fed three times with virus-containing supernatant (2 to 5 x 10(6) CFU/mL). After six days, cells were harvested and cultured in CFU-GM assay with and without a selective agent. The average rate of gene expression in CFU-GM in five independent experiments was 46% and ranged from 19% to 87%. In conclusion, the efficiency of gene transfer into canine hematopoietic progenitor cells has been increased fourfold by combining cocultivation with long-term marrow culture as compared with results obtained with cocultivation only.

Animals↗
PubMed1988

The WHI1+ gene of Saccharomyces cerevisiae tethers cell division to cell size and is a cyclin homolog.

WHI1-1 is a dominant mutation that reduces cell volume by allowing cells to commit to division at abnormally small sizes, shortening the G1 phase of the cell cycle. The gene was cloned, and dosage studies indicated that the normal gene activated commitment to division in a dose-dependent manner, and that the mutant gene had a hyperactive but qualitatively similar function. Mild over-expression of the mutant gene eliminated G1 phase, apparently entirely relaxing the normal G1 size control, but revealing hitherto cryptic controls. Sequence analysis showed that the hyperactivity of the mutant was caused by the loss of the C-terminal third of the wild-type protein. This portion of the protein contained PEST regions, which may be signals for protein degradation. The WHI1 protein had sequence similarity to clam cyclin A, to sea urchin cyclin and to Schizosaccharomyces pombe cdc13, a cyclin homolog. Since cyclins are inducers of mitosis, WHI1 may be a direct regulator of commitment to division. A probable accessory function of the WHI1 activator is to assist recovery from alpha factor arrest; WHI1-1 mutant cells could not be permanently arrested by pheromone, consistent with a hyperactivation of division.

Amino Acid Sequence↗
PubMed1988

Retroviral transfer of genes into canine hematopoietic progenitor cells.

Amphotropic retroviral vectors containing either the bacterial neomycin phosphotransferase gene or a mutant dihydrofolate reductase gene (DHFR*) were used to infect canine hematopoietic progenitor cells. Successful transfer and expression of both genes in canine hematopoietic progenitor cells has been achieved as measured by the ability of the viruses to confer resistance to either methotrexate (MTX) or the aminoglycoside G418, respectively. Gene transfer was achieved using helper-free retroviral vectors. The rate of gene expression in canine granulocyte/macrophage colony-forming units (CFU-GM) after cocultivation for 24 hours with virus-producing packaging cells ranged from 6-25%. Autologous marrow cocultivated for 24 hours with virus-producing packaging cells was transplanted into six dogs after lethal total body irradiation. All dogs showed engraftment within two weeks and four dogs survived for 5-7 months without adverse effects. One dog that had been given marrow infected with a DHFR* virus and that received MTX as in vivo selection after marrow transplantation and survived, showed 0.1 and 0.03% MTX-resistant CFU-GM at weeks 3 and 5. The efficiency of gene transfer into canine CFU-GM has been increased threefold by culturing marrow cells for six days in long-term marrow culture after 24 hour cocultivation with virus producing packaging cells.

Animals↗
PubMed1988

Polyclonal reconstitution of human marrow after allogeneic bone marrow transplantation.

Clonal dominance suggestive of reconstitution of marrow from small numbers of pluripotent hematopoietic stem cells has been noted in different experimental and clinical situations. Recipients of human allogeneic marrow transplants have not been previously studied to determine if clonal dominance occurs in this clinical setting. Clonal analysis of 20 allogeneic marrow transplant recipients was performed on DNA from peripheral blood neutrophils using restriction fragment length polymorphisms on the X chromosome. Similar studies were performed on 16 of the donors. To analyze the results further, recipients were paired with their respective donors. There was no evidence of shifts in cell populations contributing to the X chromosome inactivation patterns in recipient marrow grafts when compared with their respective donors. A mathematical model based on binomial statistics was adapted to estimate the numbers of reconstituting pluripotent hematopoietic stem cells. There was no evidence of clonal dominance suggestive of oligoclonal reconstitution in marrow grafts after allogeneic marrow transplantation. This does not preclude the possibility of oligoclonal reconstitution in other marrow transplant settings such as autologous transplantation.

Bone Marrow Transplantation↗