Choose your partner: Chromosome pairing in yeast meiosis

Premeiotic association of homologous chromosomes in the yeast, Saccharomyces cerevisiae has been shown, by means of fluorescent in situ hybridization (FISH)^(1,2). Time course and mutant studies show that the premeiotic associations are disrupted upon entry into meiosis, to be reestablished shortly before synapsis. The data are consistent with a model in which multiple, unstable interactions bring homologues together, prior to stable joining by recombination⁽³⁾.     

Precursors of both conventional and Ly-1 B cells can escape feedback inhibition of Ig gene rearrangement

Experiments with transgenic mice carrying rearranged Ig transgenes have shown that membrane bound Ig molecules cause feedback inhibition of endogenous Ig gene rearrangement. However, this inhibition is never complete. It has been postulated that escape from feedback may be a property of the Ly-1 B cell subset, whereas rearrangement of endogenous Ig genes may be completely inhibited in conventional B cells. This possibility was investigated in transgenic mice carrying a lambda transgene under the control of the H chain enhancer. It was found that kappa producing B cells in these lambda transgenic mice were for the most part, although not exclusively, of the conventional B cell phenotype. Examination of peritoneal exudate cells revealed that a large proportion of Ly-1 B cells also express kappa. Adoptive transfer of bone marrow from adult lambda transgenic mice, a source of conventional B cell precursors, resulted in the production of relatively high levels of serum kappa 2 to 3 mo after transfer into recipient SCID mice. A high proportion of donor B cells in the spleen produced endogenous kappa protein with or without co-production of lambda. It is concluded that precursors of both conventional and Ly-1 B cells can escape feedback inhibition of L chain gene rearrangement.     

Determination of antithrombocyte antibodies in the blood serum of patients with idiopathic thrombocytopenic purpura by an immunoenzyme method

Enzyme-linked immunosorbent assay (ELISA) was developed for determination of serum antiplatelet antibodies. Platelets obtained from healthy donors of blood group 0(1) were washed off plasma and sedimented on the bottom of microtest wells. After washing off unattached platelets and blocking of plastic with albumin platelets were incubated with sera under investigation and binding of serum antibodies was detected using antihuman immunoglobulin antibodies conjugated with peroxidase. Ten patients with idiopathic thrombocytopenic purpura (ITP). 1 patient with systemic lupus erythematosus. 1 patient with red blood cell aplasia and 9 healthy donors (negative control) were studied by ELISA. Serum antibodies which effectively bound to platelets were detected in 5 patients with ITP, in patient with lupus erythematosus and in patient with red blood cell aplasia.     

Enzymes Associated with Defence against Toxic Oxygen Species in PCNB-Tolerant and Sensitive Soil Fungi

The specific activities of enzymes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPOX) and glutathione reductase (GR), which are involved in protection against toxic species of oxygen, were determined in mycelia extracts of pentachloronitrobenzene (PCNB)-tolerant and susceptible soil fungi. The organisms assayed were the highly PCNB-sensitive Rhizoctonia solani and Rhizopus arrhizus; Sclerotium rolfsii and Trichoderma harzianum, which are moderately susceptible to PCNB, and the fungicide-tolerant Fusarium oxysporum f. sp. melonis and Pythium aphanidermatum. No GPOX activity was detected in the six examined fungi. Significant differences in the specific activities of the other enzyme systems among the fungi were evident. Remarkably low levels of CAT activities were measured in R. solani. Except for T. harzianum, no meaningful differences regarding SOD, CAT and GR activities with age of the fungi cultures were observed. The electrophoretic patterns of SOD and CAT displayed dissimilarities among the fungi under study. P. aphanidermatum is more polymorphic with respect to both SOD and CAT enzyme systems as compared to the other fungi. The SOD of F. oxysporum f. sp. melonis, R. arrhizus and T. harzianum is a cuprozinc enzyme, while the mangano-SOD species was detected in S. rolfsii, R. solani and T. harzianum.     

Retrovirus-mediated gene transfer into CD4+ and CD8+ human T cell subsets derived from tumor-infiltrating lymphocytes and peripheral blood mononuclear cells

Summary Studies were undertaken to test the susceptibility of individual T cell subpopulations to retroviral-mediated gene transduction. Gene transfer into human tumor-infiltrating lymphocytes (TIL) or peripheral blood mononuclear cells (PBMC) was carried out by transduction with an amphotropic murine retroviral vector (LNL6 or N2) containing the bacterialneo ^R gene. The presence of theneo ^R gene in the TIL population was demonstrated by Southern blot analysis, detection of the enzymatic activity of the gene product and by the ability of transduced TIL to proliferate in high concentrations of G418, a neomycin analog that is toxic to eukaryotic cells. The presence of theneo ^R gene in TIL did not alter their proliferation or interleukin-2 dependence compared to nontransduced TIL. The differential susceptibility of CD4⁺ and CD8⁺ lymphoid cells to the retro-virus-mediated gene transfer was then tested. Transduction of heterogeneous TIL cultures containing both CD4⁺ and CD8⁺ cells resulted in gene insertion into both T cell subsets with no preferential transduction frequency into either CD4⁺ or CD8⁺ cells. In other experiments highly purified CD4⁺ and CD8⁺ T cell subpopulations from either TIL or PBMC could be successfully transduced with theneo ^R gene as demonstrated by Southern blot analysis and detection of the gene product neophosphotransferase activity. No such activity or vector DNA could be detected in controls of nontransduced cells. In these highly purified cell subsets the distinctive T cell phenotypic markers were continually expressed after transduction, G418 selection and long-term growth. Clinical trials have begun in patients with advanced cancer using heterogeneous populations of CD4⁺ and CD8⁺ gene-modified TIL. Current address: Bone Marrow Transplantation, Hadassah University Hospital, 91120 Jerusalem, Israel     

Effect of Nitrogen on Polysaccharide Production in a Porphyridium sp

Porphyridium cultures grown on either nitrate or ammonium as the nitrogen source showed similar patterns of growth and cell wall polysaccharide production. The effect of nitrogen on growth and cell wall polysaccharide production was studied by applying three regimens of supply: batch mode, in which nitrate was supplied at the beginning of the experiment and became depleted at day 6; continual mode, in which nitrate was added daily; and deficient mode, in which the cells were cultured in a nitrate-free medium. Growth was similar in the batch- and continual-mode cultures, whereas it was totally inhibited in the deficient-mode culture. Polysaccharide content (per volume) was highest in the batch-mode culture and lowest in the deficient-mode culture. However, polysaccharide production per cell was similar in the continual- and deficient-mode cultures, the highest value being found in the batch-mode culture. In addition to its effect on polysaccharide content, nitrogen affected the polysaccharide distribution between soluble and bound polysaccharides. In the deficientmode culture, most of the cell wall polysaccharide was dissolved in the medium.     

The potential of production of sulfated polysaccharides from Porphyridium

Summary The environmental conditions prevailing in Israel make marine algae an attractive crop for the production of valuable chemicals. A marine species of Porphyridium seems to fit this purpose.The unicellular red alga Porphyridium is encapsulated by a polysaccharide envelope that is present in the gel state. This polysaccharide is an acidic heteropolymer composed of sulfated sugars. It forms ionic bridges through divalent cations, thus reaching a very high molecular weight. The thickness of the polysaccharide capsule varies according to the phase of growth and the growth conditions. Its outer part dissolves in the growth medium, which becomes progressively more viscous. Sulfated polysaccharides form theramlly reversible gels similar to agar and carrageenan, which are usually extracted from marine macroalgae. These gels have been finding increasing use in commercial applications as gelling agents, thickeners, stabilizers, and emulsifiers.We have done experiments on the cultivation of a marine species of Porphyridium for the production of polysaccharides. This unicellular alga has an advantage over the macroalgae due to its relatively faster growth rate and the possibility to regulate its growth. The potential for production of the polysaccharide, both that dissolved in the external medium and that attached to the cell (including an intracellular fraction), and the effects of growth conditions on productivity were suudied in the laboratory. Porphyridium was also cultivated outdoors in seawater in 1-m² ponds and its growth potential investigated.     

The parasitoids of the aleyrodid Bemisia tabaci in Israel: development, host preference and discrimination of the aphelinid wasp Eretmocerus mundus

Developmental history and behavior of Eretmocerus mundus Mercet, a parasitoid of Bemisia tabaci was studied at 25°C. The eggs may be laid under all four nymphal instars but not under the pupa. Yet the second and third instars are preferred. The egg hatches only under the fourth instar or the pupa. Developmental medians at 25°C are: Instar I-2.5, II-4, II-4, prepupa-2 and pupa 8 days. When ovipositing, the female stands at an angle of 90° to the host, with wings raised and inserts the ovipositor under the whitefly nymph. The egg is laid close to the insertion point of the whitefly's proboscis into the leaf. After oviposition, the female apparently marks the host while drumming on it with her hind legs. She distinguishes already parasitized hosts from unparasitized ones and refrains from laying under the former. Discrimination is accomplished after antennal drumming only.

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Les parasitoïdes de Bemisia tabaci (aleyrodidae) en Israel: développement, ponte et sélection des hôtes ches Eretmocerus mundus (aphelinidae)

Résumé Le développment et le comportement de E. mundus, parasitoïde de B. tabaci, ont été étudiés à 25°C. Les oeufs sont pondus sous les quatre stades larvaires (les deuxième et troizième sont préférés) mais pas sous les nymphes. Les oeufs n'éclosent que sous les larves du quatrième stade ou les nymphes. Les temps de développement médiaux sont à 25°, les suivants: stade I: 2,5j; stade II: 4j; stade III: 4j et nymphe 8j. Pendant la ponte, la femelle est à 90° sur son hôte, les ailes dressées, et insère sa tarière sous la larve d'aleurode. L'oeuf est déposé près du point d'insertion de la trompe dans la feuille. Après l'émission, la femelle marque apparemment son hôte pendant qu'elle tambourine avec ses pattes postérieures. Elle distingue les hôtes parasités ou non, et limite sa ponte dans les premiers. La sélection est effectuée seulement après tambourinage antennaire.

     

Factors Affecting Competence for Transformation in Staphylococcus aureus

A chemically defined medium has been developed for isolation of amino acid-requiring mutants of Staphylococcus aureus strain 8325, and for use as a selective medium in transformation assays. Variables affecting transformation of both plasmid and chromosomal markers have been studied. The optimal pH and temperature for transformation are 6.75 to 7.0 and 30 C, respectively. Ca ions are required for transformation, and only cells lysogenic for the phage phi11 can be transformed. Superinfection of competent cells with phi11 does not increase the transformation frequency. Maximal number of transformants is obtained after 20 min of contact between cells and deoxyribonucleic acid. The transformation frequencies for the plasmid marker erythromycin resistance (ero) and the chromosomal markers trp, thy, and cyt are of the same order of magnitude, whereas the frequency for the chromosomal marker tyr is approximately one order of magnitude lower.     

Patterns of meiotic double-strand breakage on native and artificial yeast chromosomes

The preferred positions for meiotic double-strand breakage were mapped on Saccharomyces cerevisiae chromosomes I and VI, and on a number of yeast artificial chromosomes carrying human DNA inserts. Each chromosome had strong and weak double-strand break (DSB) sites. On average one DSB-prone region was detected by pulsed-field gel electrophoresis per 25 kb of DNA, but each chromosome had a unique distribution of DSB sites. There were no preferred meiotic DSB sites near the telomeres. DSB-prone regions were associated with all of the known ”hot spots” for meiotic recombination on chromosomes I, III and VI. Received: 19 March 1996; in revised form: 26 July 1996 / Accepted: 18 August 1996