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At least 847 records · Page 47Linked to original sources
PubMed1990

A 3'-coterminal nested set of independently transcribed mRNAs is generated during Berne virus replication.

By using poly(A)-selected RNA from Berne virus (BEV)-infected embryonic mule skin cells as a template, cDNA was prepared and cloned in plasmid pUC9. Recombinants covering a contiguous sequence of about 10 kilobases were identified. Northern (RNA) blot hybridizations with various restriction fragments from these clones showed that the five BEV mRNAs formed a 3'-coterminal nested set. Sequence analysis revealed the presence of four complete open reading frames of 4743, 699, 426, and 480 nucleotides, with initiation codons coinciding with the 5' ends of BEV RNAs 2 through 5, respectively. By using primer extension analysis and oligonucleotide hybridizations, RNA 5 was found to be contiguous on the consensus sequence. The transcription of BEV mRNAs was studied by means of UV mapping. BEV RNAs 1, 2, and 3 were shown to be transcribed independently, which is also likely--although not rigorously proven--for RNAs 4 and 5. Upstream of the AUG codon of each open reading frame a conserved sequence pattern was observed which is postulated to function as a core promoter sequence in subgenomic RNA transcription. In the area surrounding the core promoter region of the two most abundant subgenomic BEV RNAs, a number of homologous sequence motifs were identified.

Animals↗
PubMed1992

Southern blot hybridization analysis of polyoma virus-specific RNA synthesized under the block of virus replication by 5-bromo-2'-deoxyuridine.

Polyoma (Py) virus-specific RNA, synthesized at reduced level in infected cells in the presence of antiviral substance 5-bromo-2'-deoxyuridine was characterized in more detail by Southern blot hybridization analysis. Virus-specific RNA present in 3H-uridine labelled cytoplasmic preparations hybridized to individual viral DNA restriction fragments in a characteristic manner and extent, which was quantitatively assayed. BrdUrd (6.34 micrograms/ml) lowers the hybridization profiles essentially in a proportional manner, which indicates that no new, atypical transcription products are formed in the presence of BrdUrd.

Animals↗
PubMed1991

Northern blot hybridization analysis of polyoma virus-specific RNA synthesized under the block of virus replication by 5-bromo-2'-deoxyuridine.

Polyoma (Py) virus-specific RNA, synthesized at reduced level in infected cells in the presence of antiviral compound 5-bromo-2'-deoxyuridine (BrdUrd) was characterized in more detail by Northern blot hybridization analysis. The results obtained with total, cytoplasmic and poly(A)RNA, isolated from mouse embryo cell cultures 42 hrs p.i. indicate that BrdUrd (6.34 micrograms/ml) lowers the level of typical classes of major virus DNA transcripts to a similar extent and that no new, atypical transcription products are formed.

Blotting, Northern↗
PubMedDate not supplied

[Effect of the physiological state of ticks on the intensity of tick-borne encephalitis virus replication in them].

The features of tick-borne encephalitis (TBE) virus reproduction in Ixodes ricinus and Dermacentor anderson ticks were studied in relation to the age of ticks, duration of fasting, feeding, and conditions of keeping the ticks before and after the experiment. Parenteral infection of ticks at various intervals after molting showed that the most favourable conditions for reproduction, survival, transphase and transovarial transmission of the virus occurred when the virus was given to the ticks in the first days after the period of postlarval development followed by feeding the vectors on vertebrates. In this case, owing to increased metabolic activity and intensive organogenesis and histogenesis, the virus persisted in ticks in high titers till the moment of oviposition and penetrated into the egg batch. When the virus was inoculated into old exhausted females (8-18 months) virus titers in ticks were low and penetration of the virus into the egg batch was irregular.

Animals↗
PubMed1999

Generation of bovine respiratory syncytial virus (BRSV) from cDNA: BRSV NS2 is not essential for virus replication in tissue culture, and the human RSV leader region acts as a functional BRSV genome promoter.

In order to generate recombinant bovine respiratory syncytial virus (BRSV), the genome of BRSV strain A51908, variant ATue51908, was cloned as cDNA. We provide here the sequence of the BRSV genome ends and of the entire L gene. This completes the sequence of the BRSV genome, which comprises a total of 15,140 nucleotides. To establish a vaccinia virus-free recovery system, a BHK-derived cell line stably expressing T7 RNA polymerase was generated (BSR T7/5). Recombinant BRSV was reproducibly recovered from cDNA constructs after T7 RNA polymerase-driven expression of antigenome sense RNA and of BRSV N, P, M2, and L proteins from transfected plasmids. Chimeric viruses in which the BRSV leader region was replaced by the human respiratory syncytial virus (HRSV) leader region replicated in cell culture as efficiently as their nonchimeric counterparts, demonstrating that all cis-acting sequences of the HRSV promoter are faithfully recognized by the BRSV polymerase complex. In addition, we report the successful recovery of a BRSV mutant lacking the complete NS2 gene, which encodes a nonstructural protein of unknown function. The NS2-deficient BRSV replicated autonomously and could be passaged, demonstrating that NS2 is not essential for virus replication in cell culture. However, growth of the mutant was considerably slower than and final infectious titers were reduced by a factor of at least 10 compared to wild-type BRSV, indicating that NS2 provides a supporting factor required for full replication capacity.

5' Untranslated Regions↗
PubMed1987

Pathogenesis of visna/maedi and caprine arthritis-encephalitis: new leads on the mechanism of restricted virus replication and persistent inflammation.

Lentiviruses are unique retroviruses which cause diseases with long incubation periods and prolonged clinical courses. The prototype lentiviruses, visna/maedi of sheep and arthritis-encephalitis virus of goats (CAEV), infect cells of the monocyte-macrophage system and replicate at a restricted level in these cells. The virus life cycle is closely associated with maturational factors in the cells; monocytes support the early stages of the replication cycle which goes to completion only when the cells mature to macrophages. Virus replication in the monocyte-macrophage results in lesions characterized by mononuclear cell infiltration of the central nervous system (CNS), lungs, synovium and mammary gland and their draining lymph nodes. Co-cultivation of sheep or goat lymphocytes with macrophages infected with visna or CAE viruses results in production of a unique interferon (LV-IFN). LV-IFN is a non-glycosylated protein of 54,000 to 64,000 daltons and has biological properties which have several implications for pathogenesis. Firstly, it retards the rate of maturation of monocytes and thus indirectly slows the rate of virus replication. Second, it restricts the rate of virus replication in mature macrophages by preventing virus maturation. Third, it induces expression of class II (Ia) antigens of the major histocompatibility complex on cells of macrophage lineage. Thus, by curtailing virus replication and enhancing expression of MHC class II antigens, LV-IFN may contribute to the induction and augmentation of the host's lymphoproliferative response to the virus.

Animals↗
PubMed1998

Temporal analyses of virus replication, immune responses, and efficacy in rhesus macaques immunized with a live, attenuated simian immunodeficiency virus vaccine.

Despite evidence that live, attenuated simian immunodeficiency virus (SIV) vaccines can elicit potent protection against pathogenic SIV infection, detailed information on the replication kinetics of attenuated SIV in vivo is lacking. In this study, we measured SIV RNA in the plasma of 16 adult rhesus macaques immunized with a live, attenuated strain of SIV (SIVmac239Deltanef). To evaluate the relationship between replication of the vaccine virus and the onset of protection, four animals per group were challenged with pathogenic SIVmac251 at either 5, 10, 15, or 25 weeks after immunization. SIVmac239Deltanef replicated efficiently in the immunized macaques in the first few weeks after inoculation. SIV RNA was detected in the plasma of all animals by day 7 after inoculation, and peak levels of viremia (10(5) to 10(7) RNA copies/ml) occurred by 7 to 12 days. Following challenge, SIVmac251 was detected in all of the four animals challenged at 5 weeks, in two of four challenged at 10 weeks, in none of four challenged at 15 weeks, and one of four challenged at 25 weeks. One animal immunized with SIVmac239Deltanef and challenged at 10 weeks had evidence of disease progression in the absence of detectable SIVmac251. Although complete protection was not achieved at 5 weeks, a transient reduction in viremia (approximately 100-fold) occurred in the immunized macaques early after challenge compared to the nonimmunized controls. Two weeks after challenge, SIV RNA was also reduced in the lymph nodes of all immunized macaques compared with control animals. Taken together, these results indicate that host responses capable of reducing the viral load in plasma and lymph nodes were induced as early as 5 weeks after immunization with SIVmac239Deltanef, while more potent protection developed between 10 and 15 weeks. In further experiments, we found that resistance to SIVmac251 infection did not correlate with the presence of antibodies to SIV gp130 and p27 antigens and was achieved in the absence of significant neutralizing activity against the primary SIVmac251 challenge stock.

Animals↗
PubMed1993

A mutation of cauliflower mosaic virus gene I interferes with virus movement but not virus replication.

A 300-bp deletion was made in gene I of a multimeric copy of cauliflower mosaic virus (CaMV). The deleted and wild-type constructs were mobilized into Agrobacterium tumefaciens and used for the inoculation of plants and leaf discs of the host plant, turnip, by "agroinfection." Infection induced by the wild-type construct gave characteristic viral symptoms on the inoculated and systemically infected leaves of whole plants whereas no infection was detected using the deletion mutant. The possibility that the deletion in gene I affected a cis-related function (e.g., encapsidation) was tested by complementation of mutant gene I with wild-type CaMV. Double infection experiments showed that replication and spread of both viruses occurred without detectable recombination of the mutant. In contrast to the situation in plants, agroinfection (Grimsley et al., 1986, Proc. Natl. Acad. Sci. USA 83, 3282-3286) of leaf discs, detected as the accumulation of encapsidated forms of virion DNA, was recorded for both the wild-type and the deleted constructs. The reduced level of virus accumulation from the mutant was consistent with virus replication occurring only in cells involved in the primary interaction with Agrobacterium, and accordingly progeny viral DNA was only detected in cells around the edge of the tissue disc. These data provide definitive evidence that CaMV gene I encodes a movement protein and show that this function can act in trans in a virus infection.

Brassica↗
PubMed1987

Herpes simplex virus replication and protein synthesis in a human blood-derived cell line.

Herpes simplex virus (HSV) types 1 and 2 were shown to replicate in a newly described human cell line (Meg) derived from the peripheral blood of a healthy volunteer. The cell line has both megakaryocyte-like and B cell-like properties. Upon infection with HSV-1 or -2, at a m.o.i. between 0.5 and 5, unlike B and T cells, the Meg cells were growth-arrested and this was accompanied by cytopathic effects and virus replication. The HSV proteins and glycoproteins B and D (gB and gD) made in the blood-derived Meg cells were compared to the corresponding proteins made in the non-blood-derived cell lines, Vero (African green monkey kidney cell) and HEp-2 (human epidermoid carcinoma cell). The maximum level of HSV protein synthesis occurred earlier in the Meg cells than in the Vero and HEp-2 cells. The electrophoretic pattern of HSV-1 and -2 proteins made in the Meg cell line was similar to the corresponding proteins made in the Vero and HEp-2 cell lines; however, some qualitative and quantitative differences were evident. There were no apparent differences detected in the migration pattern of gB made in all three cell lines while significant differences were observed with the gD species. However, upon hydrolysis with Staphylococcus aureus V-8 protease of the monoclonal antibody-purified gB and gD, distinct differences were observed in the electrophoretic pattern of the generated peptide fragments of both gB and gD made in the three cell lines. The results demonstrate that a human blood cell can support HSV replication and that species-specific post-translational modification of gB and gD occurs in HSV-infected Vero cells as compared to HSV-infected human cells.

Animals↗
PubMed2000

1,3-Dihydroxyacridone derivatives as inhibitors of herpes virus replication.

The nuclear enzyme DNA topoisomerase II is a candidate pharmacological target for treating herpes virus infections and the novel catalytic inhibitors, 7-chloro-1,3-dihydroxyacridone (compound 1), and 1,3,7-trihydroxyacridone (2) are potential lead compounds [Bastow, K.F., Itoigawa, M., Furukawa, H., Kashiwada, Y., Bori, I.D., Ballas, L.M., Lee, K.-H., 1994. Antiproliferative actions of 7-substituted 1,3-dihydroxyacridones; possible involvement of DNA topoisomerase II and protein kinase C as biochemical targets. Bioorg. Med. Chem. 2, 1403-1411; Vance, J.R., Bastow, K.F., 1999. Inhibition of DNA topoisomerase II catalytic activity by the antiviral agents 7-chloro,1,3-dihydroxyacridone and 1,3,7-trihydroxyacridone. Biochem. Pharmacol. 58, 703-708]. In this report, four new 1,3-dihydroxyacridone analogs with functional groups at either the 5-, 6- or 8-positions (compounds 3-6) were synthesized. Target compounds, three other analogs including compounds 1 and 2 and three anticancer drugs that inhibit DNA topoisomerase II (etoposide, amsacrine and aclarubicin) were then evaluated as selective inhibitors of herpes simplex virus (HSV) replication in cell culture and as enzyme inhibitors in vitro. Etoposide and amsacrine inhibited HSV but acted non-selectively. In general, the activities of 1,3-dihydroxyacridone derivatives as selective anti-HSV agents and as enzyme inhibitors varied inversely suggesting that DNA topoisomerase II probably is not the critical antiviral target. The 5-Cl congener (compound 3) was the most selective agent (about 26-fold under a stringent assay condition) but was not an enzyme inhibitor. Results of exploratory mechanistic studies with compounds 1 and 3 show that HSV replication was blocked at a stage after DNA and late protein synthesis. The acridone derivatives were also tested against human cytomegalovirus (HCMV) replication but none of them were active.

Acridines↗
PubMed1994

Characterization of rubella virus replication complexes using antibodies to double-stranded RNA.

A feature of the rubella virus (RV) replication cycle is the formation of cytoplasmic vesicle-containing structures known as replication complexes. Following detergent treatment of RV-infected cells, pre-embedding immunogold labeling electron microscopy using antiserum to double-stranded (ds) RNA was employed to characterize the replication complexes. Concentrations of gold particles were found associated with amorphous material located within the RV replication complex. Unlabeled long fine strands, 3-5 nm in width, were also frequently seen associated with this gold-labeled material. On some occasions gold-labeled vesicles within the replication complexes were also detected. The gold-labeled amorphous material was first detected in RV replication complexes at 12 hr postinfection, soon after the reported latent period of 8 hr. Concentrations of gold particles were not detected in mock-infected cells. The findings in this study indicate that the amorphous material is released from detergent-disrupted vesicles within the replication complex and that the vesicles contain the dsRNA. When cells were infected with the related Semliki Forest virus (SFV) and examined using the same antibody, similar gold-labeled material associated with unlabeled fine strands was also observed in SFV replication complexes. For both RV and SFV, the vesicles which line the inner membrane of the replication complexes contain the dsRNA which represent the viral replicative forms and replicative intermediates.

Animals↗
PubMed1989

The role of viral and cellular nuclear proteins in herpes simplex virus replication.

Following infection of cells by herpes simplex virus, the cell nucleus is subverted for transcription and replication of the viral genome and assembly of progeny nucleocapsids. The transition from host to viral transcription involves viral proteins that influence the ability of the cellular RNA polymerase II to transcribe a series of viral genes. The regulation of RNA polymerase II activity by viral gene products seems to occur by several different mechanisms: (1) viral proteins complex with cellular proteins and alter their transcription-promoting activity (e.g., alpha TIF), (2) viral proteins bind to specific DNA sequences and alter transcription (e.g., ICP4), and (3) viral proteins affect the posttranslational modification of viral or cellular transcriptional regulatory proteins (e.g., possibly ICP27). Thus, HSV may utilize several different approaches to influence the ability of host-cell RNA polymerase II to transcribe viral genes. Although it is known that viral transcription uses the host-cell polymerase II, it is not known whether viral infection causes a change in the structural elements of the nucleus that promote transcription. In contrast, HSV encodes a new DNA polymerase and accessory proteins that complex with and reorganize cellular proteins to form new structures where viral DNA replication takes place. HSV may encode a large number of DNA replication proteins, including a new polymerase, because it replicates in resting cells where these cellular gene products would never be expressed. However, it imitates the host cell in that it localizes viral DNA replication proteins to discrete compartments of the nucleus where viral DNA synthesis takes place. Furthermore, there is evidence that at least one specific viral gene protein can play a role in organizing the assembly of the DNA replication structures. Further work in this system may determine whether assembly of these structures is essential for efficient viral DNA replication and if so, why assembly of these structures is necessary. Thus, the study of the localization and assembly of HSV DNA replication proteins provides a system to examine the mechanisms involved in morphogenesis of the cell nucleus. Therefore, several critical principles are apparent from these discussions of the metabolism of HSV transcription and DNA replication. First, there are many ways in which the activity of RNA polymerase II can be regulated, and HSV proteins exploit several of these in controlling the transcription of a single DNA molecule. Second, the interplay of these multiple regulatory pathways is likely to control the progress of the lytic cycle and may play a role in determining the lytic versus latent infection decision.(ABSTRACT TRUNCATED AT 400 WORDS)

DNA Replication↗
PubMed2000

Analysis of classical swine fever virus replication kinetics allows differentiation of highly virulent from avirulent strains.

To study the replication of classical swine fever virus (CSFV) in cell culture, kinetics of viral plus-strand RNA synthesis, of viral structural and non-structural protein expression as well as of secreted and cell-associated infectious virus were determined. Highly virulent, moderately virulent and avirulent strains that were tested in standardized animal experiments to confirm their virulence were used to search for in vitro parameters allowing the differentiation of strains according to their virulence. No significant qualitative or quantitative differences were found between the strains studied when either RNA replication or protein synthesis were investigated. However, the ratio of cell-associated virus versus secreted virus proved to be considerably lower for the highly virulent strains when compared to avirulent or moderately virulent strains. These data suggest that highly virulent strains of CSFV can be distinguished in cell culture from strains with reduced virulence.

Animals↗
PubMed1984

Inhibition of herpes simplex virus replication by tobacco extracts.

Herpes simplex virus type 1 (HSV-1) has been associated with the genesis of leukoplakias, epithelial atypia, and oral cancer. Tobacco habits, such as snuff dipping, are also definitely correlated with this type of lesion. The normal cytolytic HSV-1 infection can, after in vitro inactivation, transform cells. Extracts of snuff were prepared and assayed for their ability to inhibit HSV-1 replication. Plaque formation assays of HSV-1 in the presence of snuff extract showed that a reduced number of plaques was formed. Different batches of one brand of snuff were tested for inhibition of herpes simplex virus (HSV) production. More than 99% inhibition of 24-hr HSV production was obtained with undiluted batches. The 1:5 dilutions of snuff had an inhibitory effect of 85% and 1:25 dilutions, 39%. In agreement, the attachment of the virus to the host cell and penetration of the virus to the cell nuclei were found to be inhibited as was the synthesis of viral DNA. Nicotine had an inhibitory effect, while aromatic additions to snuff were found to have no major inhibitory effect on HSV replication. Snuff extracts were prepared from different brands of snuff reported to contain high and low quantities of tobacco-specific N-nitrosamines. Brands with reported high levels of tobacco-specific N-nitrosamines had significantly greater ability to inhibit HSV replication. In conclusion, this study has shown that extracts of snuff have inhibitory effects on the production of cytolytic HSV-1 infections. A chronic snuff dipper keeps tobacco in the mouth for the major part of the day. Thus, virus shed in the oral cavity in connection with a reactivated latent HSV-1 infection has great possibilities of being affected by snuff or derivatives of snuff. It is suggested that an interaction between tobacco products and HSV-1 might be involved in the development of dysplastic lesions in the oral cavity.

Animals↗
PubMed2006

Pharmacodynamics of non-replicating viruses, bacteriocins and lysins.

The pharmacodynamics of antibiotics and many other chemotherapeutic agents is often governed by a 'multi-hit' kinetics, which requires the binding of several molecules of the therapeutic agent for the killing of their targets. In contrast, the pharmacodynamics of novel alternative therapeutic agents, such as phages and bacteriocins against bacterial infections or viruses engineered to target tumour cells, is governed by a 'single-hit' kinetics according to which the agent will kill once it is bound to its target. In addition to requiring only a single molecule for killing, these agents bind irreversibly to their targets. Here, we explore the pharmacodynamics of such 'irreversible, single-hit inhibitors' using mathematical models. We focus on agents that do not replicate, i.e. in the case of phage therapy, we deal only with non-lytic phages and in the case of cancer treatment, we restrict our analysis to replication of incompetent viruses. We study the impact of adsorption on dead cells, heterogeneity in adsorption rates and spatial compartmentalization.

Anti-Bacterial Agents↗