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1.
We have measured the effect of heat shock on three mycoplasmas (Acholeplasma laidlawii K2 and JA1 and Mycoplasma capricolum Kid) and demonstrated the induction of mycoplasma heat shock proteins under these conditions. Increased synthesis of at least 5 heat shock proteins in A. laidlawii K2, 11 heat shock proteins in A. laidlawii JA1, and 7 heat shock proteins in M. capricolum was observed by electrophoretic analysis of proteins from heat-shocked cells in sodium dodecyl sulfate-polyacrylamide gels. In all three strains, major heat shock proteins (66 to 68 and 26 to 29 kilodaltons [kDa]) were found. The 66- to 68-kDa protein cross-reacted with antibody to Escherichia coli DnaK protein, suggesting that this heat shock protein has been conserved in spite of major reductions in genetic complexity during mycoplasma evolution. A. laidlawii also contained a 60-kDa protein that cross-reacted with eubacterial GroEL protein and a 40-kDa protein that cross-reacted with E. coli RecA protein. Unlike with coliphages, the mycoplasma virus L2 progeny yield was not increased when virus was plated on heat-shocked A. laidlawii host cells. However, UV-irradiated L2 virus could be host cell reactivated by both A. laidlawii SOS repair and heat shock systems. 相似文献
2.
Interaction of mycoplasma virus type 2 with cellular components of Acholeplasma laidlawii strain JA1. 总被引:2,自引:1,他引:1
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Mycoplasma virus type 2 was shown to adsorb specifically to intact cells, membranes, and lipoglycan of Acholeplasma laidlawii strain JA1 but not to these components of Acholeplasma oculi. The oligosaccharide chain of the lipoglycan defined the specificity of the receptor site since deacylation not only did not reduce adsorption but increased it threefold. Actual adsorption of virus to lipoglycan was demonstrated by sucrose density gradient separation of the virus-lipoglycan complex. A strain of A. laidlawii, JA1r, resistant to infection with mycoplasma virus type 2, was incapable of adsorbing the virus and was devoid of lipoglycan. 相似文献
3.
Polyethylene glycol-dependent transfection of Acholeplasma laidlawii with mycoplasma virus L2 DNA 总被引:10,自引:5,他引:5
Phenol-extracted DNA from mycoplasma virus L2 was able to transfect Acholeplasma laidlawii in the presence of polyethylene glycol. Transfection was sensitive to DNase and was most efficient with 36% (wt/vol) polyethylene glycol 8000 and cells in logarithmic growth. Virus production by the transfected cells was similar to that of the cells infected by intact virus. L2 DNA transfected A. laidlawii with a single-hit dose-response curve, reaching saturation at high DNA concentrations. Optimum transfection frequencies were about 10(-7) transfectants per L2 DNA molecule and 10(-4) transfectants per CFU. When DNA was present in saturating amounts, the number of transfectants increased linearly with the number of CFU present in the transfection mixture, suggesting that DNA uptake does not occur by a mechanism involving cell fusion. The cleavage of the superhelical mycoplasma virus L2 genome with restriction endonucleases that cleave the DNA molecule once reduced the transfection frequency. Host cell modification and restriction of transfecting L2 DNA were similar to those for infecting L2 virions. 相似文献
4.
Soluble forms of herpes simplex virus glycoprotein D bind to a limited number of cell surface receptors and inhibit virus entry into cells. 总被引:44,自引:37,他引:7
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Herpes simplex virus type 1 (HSV-1) and HSV-2 plaque production was inhibited by treating cells with soluble forms of HSV-1 glycoprotein D (gD-1t) and HSV-2 glycoprotein D (gD-2t). Both glycoproteins inhibited entry of HSV-1 and HSV-2 without affecting virus adsorption. In contrast, a soluble form of HSV-2 glycoprotein B had no effect on virus entry into cells. Specific binding of gD-1t and gD-2t to cells was saturable, and approximately 4 x 10(5) to 5 x 10(5) molecules bound per cell. Binding of gD-1t was markedly reduced by treating cells with certain proteases but was unaffected when cell surface heparan sulfate glycosaminoglycans were enzymatically removed or when the binding was carried out in the presence of heparin. Together, these results suggest that gD binds to a limited set of cell surface receptors which may be proteins and that these interactions are essential for subsequent virus entry into cells. However, binding of gD to its receptors is not required for the initial adsorption of virus to the cell surface, which involves more numerous sites (probably including heparan sulfate) than those which mediate gD binding. 相似文献
5.
Interaction of Mycoplasma with Viruses I. Primary Adsorption of Virus Is Ionic in Mechanism 总被引:7,自引:3,他引:4
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We have studied the adsorption of the Gourlay Acholeplasma virus MVL-1 to the host cell Acholeplasma laidlawii JA-1. Successful adsorption depends primarily on some unknown action of the serum factor in the medium, but evaluation of various physical parameters indicates clearly that given this factor the kinetics is pseudo first order (K = 3 x 10(-9) cm(3)/min) and the mechanism ionic. Chiefly important are the ionic strength of the cation and pH (optimal Na(+) = 0.08 M, pH = 6). The system seems indifferent to whether the cation is Na, K, NH(4), Ca, or Mg. There is little effect of temperature over the range 0 to 42 C. The diffusion constant of the virus, calculated from its geometry or its maximum adsorption rate, is consistent with its reported size and shape. 相似文献
6.
Glucose transport in Acholeplasma laidlawii B: dependence on the fluidity and physical state of membrane lipids. 总被引:3,自引:2,他引:1
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The uptake of D-glucose by Acholeplasma laidlawii B occurs via a mediated transport process, as shown by the following observations: (i) glucose permeates A. laidlawii B cells at a rate at least 100 times greater than would be expected if its entry occurred only by simple passive diffusion; (ii) the apparent activation energy for glucose uptake in A. laidlawii is significantly lower than that expected and observed for the passive permeation of this sugar; (iii) glucose uptake appears to be a saturable process; (iv) glucose uptake can be completely inhibited by low concentrations of phloretin and phlorizin; and (v) glucose uptake is markedly inhibited at temperatures above 45 C, whereas the passive entry of erythritol continues to increase logarithmically until at least 60 C. The metabolism of D-glucose by this organism is rapid and, at low glucose concentrations, the intracellular radioactivity derived from D-[14-C]glucose is at any given time a reflection of the net effect of glucose transport, glucose metabolism, and loss from the cell of radioactive metabolic products. Care must thus be taken when attempting to determine the rate of glucose transport by measuring the accumulation by the cells of the total radioactivity derived from D-[14-C]glucose. The rate of uptake of D-glucose by A. laidlawii B cells is markedly dependent on the fatty acid composition and cholesterol content of the plasma membrane and exhibits a direct dependence on the fluidity of the membrane lipids as measured by their reversible, thermotropic gel to liquie-crystalline phase transition temperatures. In contrast to the transport rates, the apparent activation energy for glucose uptake above the phase transition temperature is not dependent on membrane lipid composition. At the temperature range within the membrane lipid phase transition region, the apparent activation energy of glucose uptake is different from the activation energy observed at temperatures above the phase transition. This may reflect the superimposed operation within the phase transition region of more than one temperature-dependent process. 相似文献
7.
The binding of iodinated concanavalin A (Con A) and Ricinus communis agglutinin (RCA) to intact cells and isolated membranes of Acholeplasma laidlawii, Mycoplasma hominis and Mycoplasma capricolum decreased with the progression of the culture from the mid- to the late-logarithmic phase of growth. The binding of the lectins to Acholeplasma laidlawii membranes had no significant effect on membrane fluidity, as assessed by electron-paramagnetic resonance spectroscopy of spin-labelled fatty acids, and had no effect on several membrane-associated enzymic activities. Temperature affected the binding of Con A and RCA in an opposite manner: the binding of Con A increased, whereas that of RCA decreased, on raising the temperature from 4 degrees C to 37 degrees C. No significant difference in lectin binding was found between oleate- and elaidate-enriched membranes at low temperatures where the former was in the liquid-crystalline state and the latter in the gel state, suggesting that membranes fluidity does not influence the binding of Con A and RCA to Acholeplasma laidlawii membranes. 相似文献
8.
Alteration of colonial morphology of Acholeplasma laidlawii and Acholeplasma modicum by infection with Mycoplasmatales viruses.
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Morphologically aberrant colonies resulted from the infection of Acholeplasma laidlawii with two of its three known viruses and from Acholeplasma modicum cells naturally carrying virus. The patterns of colonial alteration differed between cells infected with the two A. laidlawii viruses. Colonies derived from single cells infected with the bullet-shaped virus MV-L1 (Mycoplasmatales virus-laidlawii-1) had a radial sectoring pattern of intracolonial swellings ("blebs"), whereas cells infected with the tailed icosahedral virus MV-L3 contained bubble-like blebs. Colonies from cellsinfected with the enveloped virus MV-L2 appeared identical to those obrained from uninfected cells. Aberrant colonies contained 10(6) colony-forming units of organisms and 10(6) plaque-forming units of virus serologically identical to the infecting type, indicating that both the virus and host organism were capable of simultaneous replication. Enumeration of virus by means of counting aberrant colonies was 30-fold more sensitive than infectious center assay for MV-L1 and 1.2- to 2-fold higher for MV-L3. Furthermore, blebbed colonies plaquing with a new virus specific to A. modicum. Thus, blebbing in colonies provides a valuable marker for detection of the Mycoplasmatales viruses. 相似文献
9.
Host cell and ultraviolet reactivation of ultraviolet-irradiated Mycoplasmaviruses. 总被引:6,自引:1,他引:5
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The mycoplasma Acholeplasma laidlawii was shown to have mechanisms for both host cell and ultraviolet (UV) reactivation of UV-irradiated mycoplasmaviruses. Host cell reactivation was examined by comparing the survival abilities of UV-irradiated double-stranded deoxyribonucleic acid mycoplasmavirus plated on both untreated and on acriflavine-treated cells. Acriflavine treatment inhibited cell exision repair. Decreased survival on the acriflavine-treated cells demonstrated host cell reactivation. UV reactivation was studied by comparing the survival of UV-irradiated virus plated on untreated cells with its survival on cells that received a small UV dose before plating. The UV-irradiated cells gave increased virus survival, showing UV reactivation. Similar experiments with a single-stranded deoxyribonucleic acid mycoplasmavirus showed that this virus could be UV reactivated, but not host cell reactivated. 相似文献
10.
M Furukawa S Arai T Munakata K Kuwano H Inoue K Tomita 《Nihon saikingaku zasshi. Japanese journal of bacteriology》1989,44(6):817-821
Several species of mycoplasmas including M. pneumoniae, the causative agent of human respiratory infection, were investigated for tumor necrosis factor-alpha (TNF-alpha) induction. The cytotoxic activity to Meth A cells of peritoneal macrophages purified from BALB/c mice was enhanced markedly when cultured with either viable or nonviable mycoplasmas. The supernatant of macrophage culture mixed with mycoplasmas, M. pneumoniae or A. laidlawii, showed a potent cytotoxic activity to TNF-alpha-sensitive but not to TNA-alpha-insensitive L cells. Addition of anti-TNA-alpha antiserum inhibited completely the cytotoxic activity of the supernatant, indicating that the cytotoxic activity is due mostly to TNF-alpha. These results strongly suggest that mycoplasmas possess an activity to induce TNF-alpha, which enhances the cytotoxic activity of macrophages and prevent infection with mycoplasmas in vivo. 相似文献
11.
The respiratory activity of the Acholeplasma laidlawii cells was studied in order to elucidate a possible mechanism of coupling of transport with energy. The respiration of the cells is stimulated by ethanol, glucose, NADH, lactate, and pyruvate. The substrates of the Krebs cycle have no effect on the respiration. The respiratory activity, stimulated by ethanol and glucose, is inhibited by the inhibitors of the respiratory chain, SH reagents, and the inhibitors of glycolysis. The results of experiments with inhibitors suggest that the respiratory chain in the A. laidlawii cells is reduced and terminated by flavoprotein. This is confirmed by the results of spectroscopic analysis of cytochromes. Respiration coupled with phosphorylation did not play any important role in the active transport of carbohydrates. Probably, the energy, necessary for the transport of carbohydrates, is supplied by the substrate phosphorylation. This explains the activation of respiration by glucose, which is so sensitive to arsenate. The respiration of the A. laidlawii cells is not stimulated by some carbohydrates (fructose, 3-O-methyl-D-glucose). 相似文献
12.
Mycoplasmal infection of insect cell cultures 总被引:1,自引:0,他引:1
Twenty-five cell cultures of three insect orders from eight laboratories were tested for mycoplasmal infection. Acholeplasma laidlawii was detected in one culture, an incidence of 4.0%. A. laidlawii, Mycoplasma orale, M. arginini, but not M. hyorhinis, could establish infections of drosophila Dm-1 cell cultures at 25 degrees C. In prospective studies, drosophila Dm-1 cultures were intentionally infected with broth-propagated A. laidlawii and M. hyorhinis. M. hyorhinis did not grow and was eliminated from the Dm-1 cultures during consecutive passages. A. laidlawii grew without obvious cytopathic effects during six weekly passages; titers of over 10(7) CFU/ml were recorded at Passages 2 and 5 (p2 and p5). Minimal cell culture infectious doses were also determined during these studies. 0.1 milliliter cell samples were inoculated into Leighton tubes containing either fresh M1A culture medium or 3T6 indicator cells in McCoy's 5a medium. After 4 d of incubation at 25 and 37 degrees C, respectively, the cover slips were stained by DNA fluorochrome Hoechst 33258 (A. laidlawii) or by specific fluorescein-conjugated antiserum (M. hyorhinis). At p2 with both mycoplasma species, the procedure using M1A medium and incubation at 25 degrees C without 3T6 cells was inferior to indicator cells. In five of six experiments at least a two-log higher titer of mycoplasmas was needed to be detected with M1A and 25 degrees C. At p5 no difference could be found. Uridine phosphorylase assays of Dm-1 cultures infected with A. laidlawii, M. hyorhinis, M. orale, and M. arginini gave clearly positive results only with A. laidlawii. The ratio of incorporated uridine to incorporated uracil method yielded false positives with two drosophila cell lines. Suggestions for assay of mycoplasmas in invertebrate cell cultures are given. 相似文献
13.
Influenza viruses A/PR/8/34 (PR8; H1N1), A/Aichi/68 X-31 (HKx31; H3N2), and A/Beijing/89 X-109 (BJx109; H3N2) show marked differences in their ability to infect murine macrophages, including resident alveolar and peritoneal macrophages as well as the macrophage-derived cell line J774. The hierarchy in infectivity of the viruses (PR8 < HKx31 < BJx109) resembles that of their reactivity with mannose-binding lectins of the collectin family. Since the macrophage mannose receptor recognizes the same spectrum of monosaccharides as the collectins do, we investigated the possible involvement of this receptor in infection of macrophages by influenza virus. In competitive binding studies, the binding of (125)I-labeled mannosylated bovine serum albumin to macrophages was inhibited by the purified hemagglutinin and neuraminidase (HANA) glycoproteins of influenza virus but not by HANA that had been treated with periodate to oxidize its oligosaccharide side chains. The inhibitory activity of HANA from the three strains of virus differed markedly and correlated with the infectivity of each virus for macrophages. Infection of macrophages, but not MDCK cells, by influenza virus was inhibited by yeast mannan. A variant line of J774 cells, J774E, which expresses elevated levels of the mannose receptor, was more readily infected than J774, and the sensitivity of J774E cells to infection was greatly reduced by culture in the presence of D-mannose, which down-modulated mannose receptor expression. Together, the data implicate the mannose receptor as a major endocytic receptor in the infectious entry of influenza virus, and perhaps other enveloped viruses, into murine macrophages. 相似文献
14.
Lee J Im YH Jung HH Kim JH Park JO Kim K Kim WS Ahn JS Jung CW Park YS Kang WK Park K 《Biochemical and biophysical research communications》2005,334(2):313-318
The A549 cells, non-small cell lung cancer cell line from human, were resistant to interferon (IFN)-alpha treatment. The IFN-alpha-treated A549 cells showed increase in protein expression levels of NF-kappaB and COX-2. IFN-alpha induced NF-kappaB binding activity within 30 min and this increased binding activity was markedly suppressed with inclusion of curcumin. Curcumin also inhibited IFN-alpha-induced COX-2 expression in A549 cells. Within 10 min, IFN-alpha rapidly induced the binding activity of a gamma-(32)P-labeled consensus GAS oligonucleotide probe, which was profoundly reversed by curcumin. Taken together, IFN-alpha-induced activations of NF-kappaB and COX-2 were inhibited by the addition of curcumin in A549 cells. 相似文献
15.
In the present study, antibodies were raised against the Mg(2+)-ATPase and the immunological relationships between the enzyme and other ATPase from a variety of biological membranes were determined. The anti Mg(2+)-ATPase antiserum inhibited 85% of the enzyme activity from A. laidlawii membranes. We demonstrate a specific selectivity of Mg(2+)-ATPase antiserum for antigenic determinants of the A. laidlawii membranes. Immunoblot studies of A. laidlawii membrane peptides indicated labeling of five bands, 66KD, 49KD, 34KD, 26KD and 13KD, corresponding to five subunits of the ATPase in A. laidlawii membranes. 相似文献
16.
Binding of rabbit hemorrhagic disease virus to antigens of the ABH histo-blood group family 总被引:6,自引:0,他引:6
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Ruvoën-Clouet N Ganière JP André-Fontaine G Blanchard D Le Pendu J 《Journal of virology》2000,74(24):11950-11954
The ability of rabbit hemorrhagic disease virus to agglutinate human erythrocytes and to attach to rabbit epithelial cells of the upper respiratory and digestive tracts was shown to depend on the presence of ABH blood group antigens. Indeed, agglutination was inhibited by saliva from secretor individuals but not from nonsecretors, the latter being devoid of H antigen. In addition, erythrocytes of the rare Bombay phenotype, which completely lack ABH antigens, were not agglutinated. Native viral particles from extracts of infected rabbit liver as well as virus-like particles from the recombinant virus capsid protein specifically bound to synthetic A and H type 2 blood group oligosaccharides. Both types of particles could attach to adult rabbit epithelial cells of the upper respiratory and digestive tracts. This binding paralleled that of anti-H type 2 blood group reagents and was inhibited by the H type 2-specific lectin UEA-I and polyacrylamide-conjugated H type 2 trisaccharide. Young rabbit tissues were almost devoid of A and H type 2 antigens, and only very weak binding of virus particles could be obtained on these tissues. 相似文献
17.
The emergence of influenza A viruses resistant to the two existing classes of antiviral drugs highlights the need for additional antiviral drugs, particularly considering the potential threat of a pandemic of H5N1 influenza A viruses. Here, we determine whether influenza A virus replication can be selectively inhibited by blocking the ability of its NS1A protein to inhibit the 3'-end processing of cellular pre-mRNAs, including beta interferon (IFN-beta) pre-mRNA. Pre-mRNA processing is inhibited via the binding of the NS1A protein to the cellular CPSF30 protein, and mutational inactivation of this NS1A binding site causes severe attenuation of the virus. We demonstrate that binding of CPSF30 is mediated by two of its zinc fingers, F2F3, and that the CPSF30/F2F3 binding site on the NS1A protein extends from amino acid 144 to amino acid 186. We generated MDCK cells that constitutively express epitope-tagged F2F3 in the nucleus, although at only approximately one-eighth the level of the NS1A protein produced during virus infection. Influenza A virus replication was inhibited in this cell line, whereas no inhibition was observed with influenza B virus, whose NS1B protein lacks a binding site for CPSF30. Influenza A virus, but not influenza B virus, induced increased production of IFN-beta mRNA in the F2F3-expressing cells. These results, which indicate that F2F3 inhibits influenza A virus replication by blocking the binding of endogenous CPSF30 to the NS1A protein, point to this NS1A binding site as a potential target for the development of antivirals directed against influenza A virus. 相似文献
18.
Membrane vesicles obtained from Acholeplasma laidlawii accumulate glucose as well as maltose and fructose against their concentration gradient in the absence of exogenous energy sources. Glucose uptake by membrane vesicles is inhibited by anaerobiosis and by electron transfer inhibitors, such as rotenone and amytal, but not by 2-heptyl-4-hydroxyquinoline N-oxide, antimycin A, cyanide and azide. Rotenone, cyanide and amytal also produce a rapid efflux of glucose from the membrane vesicles. Arsenate, oligomycin and N,N'-dicyclohexylcarbodimide do not inhibit glucose transport. Transport of glucose is markedly inhibited by proton conductors such as CCCP and pentachlorophenol. It is concluded that glucose transport can be driven by a high-energy state of the membrane or by the membrane potential. 相似文献
19.
Double-stranded DNA from mycoplasma virus L2 can transfect Acholeplasma laidlawii cells in the presence of polyethylene glycol (T. L. Sladek and J. Maniloff, J. Bacteriol. 155:734-741, 1983). We report here that both single-stranded DNA and double-stranded replicative form DNA, from the single-stranded DNA mycoplasma virus L51, are also infectious in this system. For both DNAs transfection frequencies were in the range of 10(-8) transfectants per DNA molecule and 10(-3) transfectants per CFU. An unexpected finding was that both DNAs could transfect A. laidlawii strain REP-, a variant which is a nonpermissive host for single-stranded DNA mycoplasma viruses due to a block in viral DNA replication (Nowak et al., J. Bacteriol. 127:832-836, 1976). The number of viruses produced by transfected REP- cells was comparable to the number produced by both transfected and infected wild-type cells. Therefore, transfected L51 DNAs are able to bypass the replication block in REP- cells that occurs when these cells are infected by L51 virions. 相似文献
20.
Two large virion envelope glycoproteins mediate Epstein-Barr virus binding to receptor-positive cells. 总被引:11,自引:9,他引:2
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The four major Epstein-Barr virion envelope components were separated by column chromatography and reconstituted into artificial liposomes. These liposomes were tested for their ability to bind selectively to Epstein-Barr virus receptor-positive cells. Only when the two high-molecular-weight glycoproteins, VE1 and VE2, were present together was a stable binding complex formed. The addition of the other virion envelope components did not increase the levels of binding. This binding was inhibited by unlabeled viable virions and by neutralizing antisera, which recognized the two components. Adsorption of viable virus was also eliminated by the antisera. The enzyme susceptibility pattern of the cell-liposome interaction is similar to that of the virus-cell interaction, thus confirming the specificity of the binding site. A model for Epstein-Barr virus binding in which VE1 and VE2 coordinately recognize the same binding site is presented. 相似文献