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101.
Identification and mapping of Epstein-Barr virus early antigens and demonstration of a viral gene activator that functions in trans. 总被引:31,自引:22,他引:9
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The BamHI M DNA fragment of the Epstein-Barr virus (EBV) genome was inserted in two orientations into a simian virus 40-based expression vector, and the EBV-specific proteins produced in COS-7 monkey cells were examined. In one orientation, termed BamHI-M rightward reading frame 1 (BMRF1), a set of phosphoproteins ranging in size from 47,000 to 54,000 daltons was synthesized. These proteins reacted with monoclonal and polyclonal antisera, defining them as components of the EBV early antigen diffuse set of proteins (EA-D). The BamHI M DNA fragment in the opposite orientation, termed BamHI-M leftward reading frame 1 (BMLF1), directed the synthesis of a nuclear antigen detected by antibodies in serum from a patient with nasopharyngeal carcinoma. The BMLF1 antigen was not detected by monoclonal or polyclonal antibodies directed against the EA-D complex. A series of deletion mutants were constructed in the BamHI M DNA fragment, and the EA-D complex and BMLF1 antigen were mapped to discrete open reading frames in this DNA fragment. A test for several possible functions of these antigens showed that the BMLF1 antigen had the ability to activate or enhance, in trans, the level of expression of a gene under the control of the adenovirus early region 3 promoter or the simian virus 40 early promoter in the absence of its cis-acting enhancer. These experiments demonstrate a new gene function, encoded by EBV, that may be important in the positive regulation of viral or cellular genes. 相似文献
102.
Summary Plant survival rate, root disease index and fresh shoot weight of subterranean clover seedlings inoculated with fungal pathogens
(Fusarium avenaceum, F. oxysporum, Phoma medicaginis, Pythium irregulare andRhizoctonia solani, both singly and in combinations) were generally inter-correlated over a wide range of temperature (10, 15, 20 and 25°C)
and moisture conditions (45, 65% water holding capacity and flooding). There was a negative correlation between root disease
index and shoot weight for all treatments exceptF. avenaceum + P. irregulare. Root disease index and seedling survival rate were negatively correlated except forF. oxygsporum, Phoma medicaginis, P. irregulare andF. oxysporum + F. avenaceum. However, a good positive correlation was found between the survival rate and shoot weight for all treatments with the exception
ofPhoma medicaginis. 相似文献
103.
Simultaneous Measurements of Steady State Chlorophyll a Fluorescence and CO(2) Assimilation in Leaves: The Relationship between Fluorescence and Photosynthesis in C(3) and C(4) Plants 总被引:3,自引:3,他引:0
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Rates of CO2 assimilation and steady state chlorophyll a fluorescence were measured simultaneously at different intercellular partial pressures of CO2 in attached cotton (Gossypium hirsutum L. cv Deltapine 16) leaves at 25°C. Electron transport activity for CO2 assimilation plus photorespiration was calculated for these experiments. Under light saturating (1750 microeinsteins per square meter per second) and light limiting (700 microeinsteins per square meter per second) conditions there was a good correlation between fluorescence and the calculated electron transport activity at 19 and 200 millibars O2, and between fluorescence and rates of CO2 assimilation at 19 millibars but not 200 millibars O2. The values of fluorescence measured at about 220 microbars intercellular CO2 were not greatly affected by increasing O2 from 19 to 800 millibars. Fluorescence increased with light intensity at any one intercellular CO2 partial pressure. But the values obtained for fluorescence, expressed as a ratio of the maximum fluorescence obtained in DCMU-treated tissue, over the same range of CO2 partial pressure at 500 microeinsteins per square meter per second were similar to those obtained at 1000 and 2000 microeinsteins per square meter per second. There were two phases in the observed correlation between fluorescence and calculated electron transport activity: an initial inverse relationship at low CO2 partial pressures which reversed to a positive correlation at higher values of CO2 partial pressures. Similar results were observed in the C3 species Helianthus annuus L., Phaseolus vulgaris L., and Brassica chinensis. In all C4 species (Zea mays L., Sorghum bicolor L., Panicum maximum Jacq., Amaranthus edulis Speg., and Echinochloa frumentacea [Roxb.] Link) examined changes in fluorescence were directly correlated with changes in CO2 assimilation rates. The nature and the extent to which Q (primary quencher) and high-energy state (qE) quenching function in determining the steady state fluorescence obtained during photosynthesis in leaves is discussed. 相似文献
104.
105.
Structural and enzymological characterization of immunoaffinity-purified DNA polymerase alpha.DNA primase complex from KB cells 总被引:16,自引:0,他引:16
S W Wong L R Paborsky P A Fisher T S Wang D Korn 《The Journal of biological chemistry》1986,261(17):7958-7968
We describe the polypeptide structure and some of the catalytic properties of a DNA polymerase alpha.DNA primase complex that can be prepared from KB cells by immunoaffinity purification. The procedure is based on monoclonal antibodies that were raised against a biochemically purified, catalytically active core protomer of the polymerase. In all respects tested, the basic mechanism of substrate recognition and binding by the immunoaffinity-purified polymerase is qualitatively identical to that of the core protomer. The immunoaffinity-purified KB cell polymerase alpha X DNA primase is structurally complex. On the basis of extensive immunochemical analyses with five independent monoclonal antibodies, three of which are potent neutralizers of polymerase alpha activity, peptide mapping studies, and the application of a sensitive immunoassay that permits detection of polymerase alpha antigens in crude cell lysates, we have established that the principal form of catalytically active DNA polymerase alpha in KB cells is a phosphoprotein with a molecular mass of 180 kilodaltons. This protein is stable in vivo, with an estimated half-life of greater than or equal to 15 h. In contrast, the polypeptide is extremely fragile in vitro and generates partial degradation products of p165, p140, and p125 that explain the "microheterogeneity" typically exhibited by polymerase alpha peptides in denaturing polyacrylamide gels. In addition to the catalytically active polymerase alpha polypeptide(s), the immunopurified enzyme fraction typically contains three other proteins, p77, p55, and p49, the functions of which have not yet been established. These proteins do not display polymerase alpha epitopes and have been shown by peptide mapping to be independent species that are unrelated either to the large polymerase peptides or to one another. The polypeptide p77 is also a phosphoprotein, and in both p180 and p77 the phosphorylated amino acids are exclusively serine and threonine. 相似文献
106.
Activated N-ras gene induces neuronal differentiation of PC12 rat pheochromocytoma cells 总被引:19,自引:0,他引:19
Activated mouse N-ras gene transfected into PC12 rat pheochromocytoma cells suppressed proliferation and promoted neuronal differentiation. Normal mouse N-ras in a LTR-containing vector caused differentiation with a reduced efficiency, but normal N-ras in a vector lacking LTR sequences failed to alter the PC12 phenotype. Cultures of NGF-resistant PC12 variant subline U7 also showed outgrowth of neurites and cessation of cell division following transfection with the mutated ras gene. The present findings suggest that ras genes can, in certain cells, play a role in promoting differentiation and suppressing proliferation, in contrast to their established oncogenic neoplasia-promoting activity in other cells. 相似文献
107.
Cytolytic T lymphocytes (CTL) specific for trinitrophenyl (TNP)-altered self antigens can be generated in vivo through the simultaneous injection of TNP-modified syngeneic spleen cells and H-2-compatible, minor histocompatibility locus (Mls)-disparate auxiliary spleen cells into the footpads of mice. The latter stimulates host helper cells to produce differentiative and proliferative signals required for the generation of CTL. Advent of this protocol allowed investigation of the initiation of two different cell-mediated immune responses, delayed-type hypersensitivity (DTH) and the generation of CTL, in the same experimental animal. Mice presensitized for CTL were able to develop DTH as well as normal controls. However, when mice were first sensitized for DTH, they were thereafter incapable of generating CTL. This effect was hapten specific, relatively long lasting, and preventable by treating mice with cyclophosphamide before sensitizing for DTH. Adoptive transfer of lymphoid cells from DTH-immune mice conferred DTH reactivity upon naive recipients but not a suppressed CTL response. Therefore, cells mediating DTH were not responsible for suppression of CTL. The mechanism for suppression has been discussed from the viewpoint of the suppressor-T-cell circuits that are known to be generated when animals are sensitized for DTH and which are susceptible to treatment with cyclophosphamide. 相似文献
108.
L. M. S. Palni S. A. B. Tay S. K. Nandi D. J. Pianca G. J. M. de Klerk O. C. Wong D. S. Letham J. K. MacLeod 《Biologia Plantarum》1985,27(2-3):195-203
A range of endogenous cytokinins have been identified inDatura crown-gall tissue by GC-MS. Incorporation of [3H]adenine into zeatin riboside, zeatin and its nucleotide(s) is also shown. Metabolism studies usingcis- andtrans-isomers of zeatin riboside indicate that interconversion of the two isomers does not occur in this tissue. Data on the identity
of major endogenous cytokinins in a genetic tumour line of tobacco is also provided. 相似文献
109.
A method to quantitate Coomassie blue-stained proteins in cylindrical polyacrylamide gels 总被引:1,自引:0,他引:1
A method for the quantitation of Coomassie blue-stained proteins in cylindrical polyacrylamide gels is described. It involves an elution of the dye with an 80% methanol solution in a sealed Pyrex tube at 100 degrees C for 3 h and a measurement of its concentration at 585 nm. Using a 6.5% polyacrylamide gel and bovine serum albumin as a protein standard, the curve of absorbance of the dye solution as a function of the amount of protein was observed to be linear up to 30-40 micrograms of protein and as little as 0.8-1.0 micrograms of protein could be measured. The validity of the method was indicated by the values obtained for the relative proportions of the human erythrocyte membrane proteins. Using this method, the color yields of several proteins varying widely with respect to their size, amino acid composition, and carbohydrate content were determined in a 6.5% polyacrylamide gel. The results showed that they were generally the same except for proteins having a high carbohydrate content which were significantly lower. 相似文献
110.
N-Acetyl-, N-propionyl-, N-butyryl- and N-valerylglucosamines were synthesized as topographical probes to localize further the interaction site of alpha-lactalbumin on galactosyltransferase. All these compounds were found to be substrates for galactosyltransferase with Km values in the millimolar range. In the presence of alpha-lactalbumin, the Michaelis-Menten constants were diminished. However, the effect on the initial rates of these reactions varied. Thus, at low N-acylglucosamine concentrations, alpha-lactalbumin activated the enzyme activity, but at high concentrations, alpha-lactalbumin became inhibitory. This mixed-type inhibition kinetics indicated that a quaternary complex between galactosyltransferase, alpha-lactalbumin, Mn2+-UDPgalactose and N-acylglucosamine existed during the catalytic process. The ability of these N-acylglucosamine substrates to bind to lactose synthase complex was further substantiated by the physical association of galactosyltransferase onto the solid-bound alpha-lactalbumin in the presence of any one of these compounds. The data revealed that the presence of the N-acyl group up to five carbons in length did not interfere with the interaction between alpha-lactalbumin and galactosyltransferase, suggesting that alpha-lactalbumin was not bound in the vicinity of the C-2 region of the monosaccharide site. The inhibitory effect of alpha-lactalbumin on N-acyllactosamine formation is probably a consequence of conformational changes of galactosyltransferase. 相似文献