Cell-by-cell flow cytometric analysis of chromosomes

The authors discuss various aspects of a recently developed method permitting a detailed flow cytometric analysis of the individual cell karyotypes such as instrumentation, histochemistry, data proceeding algorithms. Possible drawbacks of the method and the ways of their overcoming are considered. Results of analysis of the Chinese hamster cells are presented that illustrate the possibilities of the method, including the metaphase chromosome distribution according to their fluorescence intensity, the analysed cell distribution according to their chromosomes number, the table in which the individual cell karyotypes are distributed according to their fluorescence. The results obtained show that the developed method may be successfully used for investigating chromosomal iNstability and heterogeneity of the mammalian cells.     

The dose dependence of the development of compensatory-restorative body reactions to irradiation. The EPR method]

The study deals with the mechanism of organism's adaptive responses to the effect of radiation in widely ranging dose. Post-irradiation metabolic changes were evaluated in canine blood as well as in murine blood, spleen, bone marrow and liver using the EPR spectroscopy. It was shown that the dynamics of changes in transferrin and ceruloplasmin pools and ribonucleotide reductase activity were phase-dependent with the maxima at the 2nd, 6th and 10-12th days after irradiation. Such dynamics was observed at various irradiation doses applied. The data allow us to suggest that the nonspecific compensatory--adaptive reactions of organisms develop as the response to irradiation. The dose-response function of the reaction intensity was found to be linear. The shape of the dose-response curve indicates that the minimum response of organism depends on the dose linearly up to 3.2 Gy (for dogs) as well as the maximum one. However, in the case of low-dose irradiation (0.25 or 0.5 Gy) there were deviations of maximum responses from the linearity, i.e. the amplification of the amplitude of compensatory adaptive reactions. These effect were shown to be dependent upon initial individual characteristics of animal blood and to be related to the "depressed" or "activated" state of organism prior to irradiation. The ribonucleotide reductase activity was measured in bone marrow and spleen of animals by the EPR method. The nature of non-repairable DNA damage is discussed in view of the inactivation of ribonucleotide reductase.     

Sequence variation in the outer-surface-protein genes of Borrelia burgdorferi

Borrelia burgdorferi is a spirochete pathogen transmitted among warm- blooded hosts by ixodid ticks. Frequency-dependent selection for variant outer-surface proteins might be expected to arise in this species, since rare variants are more likely to avoid immune surveillance in previously infected hosts. We sequenced the OspA and OspB genes of nine North American strains and compared them with nine strains previously described. For each gene, the mean number of synonymous substitutions per synonymous site and the mean number of nonsynonymous substitutions per nonsynonymous site show only a twofold excess of silent mutations. Synonymous rates vary widely along the OspB protein. Some regions show a significant excess of silent substitutions, while divergence in other regions is constrained by biased base composition or selection. The presence, in antigenically important regions of the protein, of significant variation among strains, as well as evidence for recombination among strains, should be considered in attempts to develop vaccines against this disease.      

Characterisation of a 34 kD protein from potato cyst nematodes, using monoclonal antibodies with potential for species diagnosis

Two monoclonal antibodies, which differentially recognise the two species of potato cyst nematodes (PCN), Globodera pallida and G. rostochiensis, are described. They have been shown to have potential for quantification of these two species, recognising proteins of the same molecular weight (34 kD) in both species. Further investigation showed these proteins to have isoelectric points at pH values of 5.7 in G. pallida and 5.9 in G. rostochiensis, in common with the proteins used by Fleming & Marks (1983) to differentiate the species of PCN. They are likely to be structurally very similar, with the same physiological function (and therefore similar concentrations) in the two species. In cross-reactivity tests with a wide range of soil nematode species, the antibodies reacted strongly only with species of the genus Globodera, and thereby confirmed their potential as the basis of a quantitative immunoassay likely to be useful in management of PCN populations.     

Gap junction structures: Analysis of the x-ray diffraction data

Models for the spatial distribution of protein, lipid and water in gap junction structures have been constructed from the results of the analysis of X-ray diffraction data described here and the electron microscope and chemical data presented in the preceding paper (Caspar, D. L. D., D. A. Goodenough, L. Makowski, and W.C. Phillips. 1977. 74:605-628). The continuous intensity distribution on the meridian of the X-ray diffraction pattern was measured, and corrected for the effects of the partially ordered stacking and partial orientation of the junctions in the X-ray specimens. The electron density distribution in the direction perpendicular to the plane of the junction was calculated from the meridional intensity data. Determination of the interference function for the stacking of the junctions improved the accuracy of the electron density profile. The pair-correlation function, which provides information about the packing of junctions in the specimen, was calculated from the interference function. The intensities of the hexagonal lattice reflections on the equator of the X-ray pattern were used in coordination with the electron microscope data to calculate to the two-dimensional electron density projection onto the plane of the membrane. Differences in the structure of the connexons as seen in the meridional profile and equatorial projections were shown to be correlated to changes in lattice constant. The parts of the junction structure which are variable have been distinguished from the invariant parts by comparison of the X-ray data from different specimens. The combination of these results with electron microscope and chemical data provides low resolution three- dimensional representations of the structures of gap junctions.     

Effect of 57Co-bleomycetin on the immune response induced in mice by ram erythrocytes

The effect of 57Co-bleomycin on the immune response induced by sheep red cells was studied on mice. It was found that the immune response was not suppressed when the labeled antibiotic was used in a single dose of 5-20 mg/kg. By the 5th day the level of 57Co-bleomycin in the skin, thymus and blood was 3-10 times higher than that of 57CoCl2 and in the spleen it was 1.3-1.5 times higher.     

Bead rings at the endoplasmic reticulum-golgi complex boundary: morphological changes accompanying inihibition of intracellular transport of secretory proteins in arthropod fat body tissue

Golgi complex beads are 10-nm particles arranged in rings on the smooth surface of rough endoplasmic reticulum (ER) makind the forming face of the Golgi complex (GC). In arthropod cells they stain specifically with bismuth. Their morphology has been studied after treatment with reagents known to interfere with GC function. Inhibitors of oxidative phosphorylation (antimycin A, cyanide, and anoxia), but not an inhibitor of glycolysis (iodoacetate), both cause the bead rings to collapse and the GC saccules to round up, and inhibit transition vesicle (TV) formation. Cycloheximide blocks protein synthesis on ribosomes but does not stop TV formation or disrupt bead rings, even after prolonged treatment (6 h) to allow emptying of the rough ER cisternae. Thus the collapse of bead rings is not attributable to inhibition of protein synthesis, and the ring structure of beads does not require continued protein synthesis and secretion for its maintenance. Valinomycin has effects on the GC similar to those of antimycin A, but {"type":"entrez-nucleotide","attrs":{"text":"A23187","term_id":"833253","term_text":"A23187"}}A23187, monensin, and lasalocid do not affect bead ring structure or TV formation. These results are consistent with valinomycin’s secondarily uncoupling mitochondria, which collapses bead rings and prevents TV formation. Thus inhibitors of oxidative phosphorylation do not influence the beads through cation movement. Because mononsin and lasalocid block secretion at the level of the condensing vacuoles, bead rings are not influenced by blocks in secretion distal to them or by the backup of secretory material. These experiments are consistent with inhibitors of oxidative phosphorylation collapsing bead rings by decreasing intracellular ATP. The concomitant block to TV formation and the collapse of bead rings suggests that integrity of the bead rings is essential for the transport of secretory material from the rough ER to the GC.     

Study of the conformational mobility of globular proteins in aqueous solutions according to their proton relaxation in a rotating system of coordinates

Measurements of the nuclear magnetization decay in the rotating frame for protons of SA and RNAase proteins in aqueous solutions indicate the dispersion of the relaxation rate for SA protons within the region of correlation frequencies of 10(5)--10(6) s-1. These frequencies are much lower than frequencies of rotational diffusion movements of the SA macromolecules in aqueous solutions. These must be ascribed to internal movement within protein globules due to which the distances between interacting magnetic moments are changed. This conclusion gives direct evidence in favour of existence of conformational mobility in most of the protein globule volume.