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At least 847 records · Page 47Linked to original sources
PubMed1997

Transfection of Actinomyces spp. by genomic DNA of bacteriophages from human dental plaque.

Bacteriophages that produced turbid or clear zones of lysis in strains of Actinomyces were isolated from 22 of 124 samples of fresh human dental plaque. All human and nonhuman strains of Actinomyces viscosus or Actinomyces naeslundii tested in this study were sensitive to infection by one or more of these phages. In contrast, none of the Actinomyces odontolyticus, Actinomyces israelii, or Actinomyces bovis strains tested were susceptible. Results of restriction endonuclease analyses indicated that the genomes of these phages consisted of double-stranded DNA molecules ranging in size between 16 and 60 kbp. Sequence homology under hybridization conditions of high stringency was observed among a few of the isolated phages. A lysogenized isolate of A. viscosus MG-1 was obtained following infection with a temperate phage, designated phi 225. Results of Southern blot analyses indicated that phi 225 replicated as a plasmid in the lysogenized strain. Genomic DNA from several lytic phages was used to establish conditions for transfection by electroporation of strains of Actinomyces spp. Efficiencies of DNA transfer ranged from 10(2) to 10(5) plaque-forming units per microgram of DNA were obtained under optimal transfection conditions. The results of these studies demonstrate that transfer of genetic information in Actinomyces spp. can be achieved by transfection.

Actinomyces↗
PubMed1988

Viral infection of neurons can depress neurotransmitter mRNA levels without histologic injury.

Neonatal mice inoculated with lymphocytic choriomeningitis virus (LCMV) have non-lytic persistent neuronal infection and disturbed behavior. We now show that LCMV replicates in neurons containing the neurotransmitter somatostatin without morphologic evidence of injury and that persistent neuronal LCMV infection in mice is attended by a decrease in brain levels of somatostatin mRNA. Brain levels of mRNA for another neurotransmitter peptide, cholecystokinin, are not decreased. These data are the first to localize a virus to a specific neurotransmitter-containing cell during in vivo infection and suggest that persistent viral infections could cause neurologic or psychiatric diseases through selective effects on brain levels of neurotransmitter mRNAs.

Animals↗
PubMed1997

Evidence for regulation of transcription and replication of the human neurotropic virus JCV genome by the human S(mu)bp-2 protein in glial cells.

Glial factor 1 (GF-1) is a partial cDNA isolated from a human brain cDNA library which encodes a truncated protein with binding ability to the B-regulatory domain of the human neurotropic virus, JCV. GF-1 exhibits sequence homology to the central region of the newly identified human DNA-binding protein S(mu)bp-2. GF-1 appears to be a partial cDNA for human S(mu)bp-2 based on its sequence homology to S(mu)bp-2 and their chromosomal co-localization. In this report, we have employed transfection assay and have compared the ability of GF-1 and its full-length form, S(mu)bp-2, on regulating the activity of JCV promoters in glial cells. Our results demonstrate that, unlike GF-1 which stimulates JCV early promoter in glial cells, overexpression of S(mu)bp-2 exhibits no drastic effect on the transcription of the viral early promoter. The activity of the viral late promoter was noticeably increased by both GF-1 and S(mu)bp-2, although the level of induction by GF-1 was consistently higher than that detected by S(mu)bp-2. Use of deletion constructs in co-transfection assay revealed that the B-domain of the JCV promoter is required for transcriptional activation by GF-1 and S(mu)bp-2. Expression of GF-1 and S(mu)bp-2 in glial cells increased the induced level of JCV late gene transcription by the viral early protein, T-antigen. Examination of the viral DNA replication by DpnI assay indicated that, unlike GF-1, S(mu)bp-2 has the ability to decrease the level of JCV DNA replication in glial cells. These observations suggest that the N-terminal portion of S(mu)bp-2 which encompasses several helicase motifs and/or its C-terminus, both of which are missing in GF-1, may confer differential effects on viral gene transcription and replication. The biological importance of our findings in regulation of the JCV lytic cycle in glial cells is discussed.

Antigens, Viral, Tumor↗
PubMed1990

Epstein-Barr virus BZLF1 trans activator induces the promoter of a cellular cognate gene, c-fos.

The Epstein-Barr virus BZLF1 gene product ZEBRA is a DNA-binding protein that is partially homologous to c-Fos, binds specifically to AP-1 sites, and can induce the lytic cycle in latently infected B lymphocytes. Induction of the viral lytic cycle can also be achieved by treatment with the phorbol ester 12-O-tetrade-canoylphorbol-13-acetate, a reagent which activates gene expression in part through AP-1 (Jun/Fos). In this article the interrelationship between ZEBRA and AP-1 is extended by the demonstration that ZEBRA can induce c-Fos expression through AP-1 and "AP-1-like" sites present in the c-fos promoter. Induction of c-Fos may be necessary for the expression of other viral lytic genes and perhaps cellular genes whose products are required for viral replication.

Base Sequence↗
PubMed1993

Immediate-early transcription over covalently joined genome ends of bovine herpesvirus 1: the circ gene.

Herpesvirus genomes are linear molecules in virions. Prior to replication in host cells, they form circular templates by unknown mechanisms. Examining lytic infection with bovine herpesvirus 1, we observed immediate-early transcription over joined genome ends, which suggested that circles are present at the initial stage of infection. Among the transcripts was a spliced immediate-early RNA (1.5 kb) sharing exon 1 with previously described major immediate-early transcripts from the right genome end and exon 2 with a late transcript located near the left genome end. Exon 2 encodes a putative circ-encoded protein with homology to the varicella-zoster virus open reading frame 2 and equine herpesvirus 1 open reading frame 3 products. The novel features reported here for bovine herpesvirus 1 may constitute a more general property of herpesviruses.

Amino Acid Sequence↗
PubMed1999

Role of the epstein-barr virus RTA protein in activation of distinct classes of viral lytic cycle genes.

Initiation of the Epstein-Barr virus (EBV) lytic cycle is controlled by two immediate-early genes, BZLF1 and BRLF1. In certain epithelial and B-cell lines, their protein products, ZEBRA and Rta, stimulate their own expression, reciprocally stimulate each other's expression, and activate downstream viral targets. It has been difficult to examine the individual roles of these two transactivators in EBV-infected lymphocytes, as they are expressed simultaneously upon induction of the lytic cycle. Here we show that the Burkitt lymphoma cell line Raji represents an experimental system that allows the study of Rta's role in the lytic cycle of EBV in the absence and presence of ZEBRA. When expressed in Raji cells, exogenous Rta does not activate endogenous BZLF1 expression, yet Rta remains competent to transactivate certain downstream viral targets. Some genes, such as BaRF1, BMLF1, and a late gene, BLRF2, are maximally activated by Rta itself in the absence of detectable ZEBRA. The use of the Z(S186A) mutant form of ZEBRA, whose transactivation function is manifest only by coexpression of Rta, allows identification of a second class of lytic cycle genes, such as BMRF1 and BHRF1, that are activated in synergy by Rta and ZEBRA. It has already been documented that of the two activators, only ZEBRA stimulates the BRLF1 gene in Raji cells. Thus, there is a third class of viral genes activated by ZEBRA but not Rta. Moreover, ZEBRA exhibits an inhibitory effect on Rta's capacity to stimulate the late gene, BLRF2. Consequently ZEBRA may function to repress Rta's potential to activate some late genes. Raji cells thus allow delineation of the combinatorial roles of Rta and ZEBRA in control of several distinct classes of lytic cycle genes.

DNA Replication↗
PubMed1991

Prevention and treatment of hepatitis delta virus infection.

The prevention of HDV infection can readily be achieved by immunisation with the envelope proteins (HBsAg/pre-S) of the helper virus (HBV) on which HDV is dependent. In patients with established chronic HBV infection, superinfection with HDV may theoretically be controlled, but not prevented, by immunisation to the internal components of HDV which may be the target of a cytotoxic T-cell response to HDV infected hepatocytes. Such a response, by analogy to the effects of immunisation to HBcAg, may result in rapid lysis of infected hepatocytes thereby limiting the spread of HDV through the liver. In our preliminary experiments using recombinant HDAg (amino-acids 13-76) we have shown that a humoral immune response to HDV prior to HDV superinfection, does not control HDV infection but may facilitate it. We do not know whether this immunisation protocol successfully induced a cytotoxic T-cell response or whether the region of HDAg included is recognised by cytotoxic T-cells and involved in the immune destruction of HDV infected cells. Further studies of the mechanisms of lysis of HDV infected hepatocytes to determine the role, if any, of cytotoxic T-cells or humoral lytic mechanisms, is needed. Although short courses of alpha interferon inhibit HDV replication, relapses are common and amelioration of the inflammatory liver disease is not always seen. Longer periods of therapy may be effective if they can be continued until clearance of the helper virus (HBV), as well as HDV, has occurred. In three patients treated for greater than one year, clearance of HBsAg has occurred. If these patients remain in this state after stopping therapy, further studies of long term alpha interferon therapy may be fruitful.

Animals↗
PubMed1986

Enhanced lysis of herpes simplex virus type 1-infected mouse cell lines by NC and NK effectors.

Spontaneously cytotoxic murine lymphocytes lysed certain cell types infected by herpes simplex virus type 1 (HSV-1) better than uninfected cells. The levels of virus-directed lysis varied widely from target to target, and we found that differences in virus-directed lytic efficiency could be attributed both to the characteristics of HSV-1 replication in the different targets and to the subgroup of natural effector cells which mediated lysis. Although HSV-1 adsorbed to the surface of all the target cells, those in which the virus replicated more efficiently were lysed to a greater extent. As targets, we used cell lines that, when uninfected, were spontaneously lysed by NK cells (YAC-1) or by NC cells (WEHI-164). We also used a fibroblastoid cell line (M50) and a monocytic tumor line (PU51R), which were not spontaneously killed. Using complement-mediated elimination of Qa-5-positive or asialo-GM1-positive NK cells to distinguish NK from NC activity, we found that NK cells lysed HSV-1-infected YAC cells better than uninfected cells, and an NC-like activity selectively lysed HSV-1-infected WEHI cells. In addition, we showed that both NK and NC cytotoxicities contributed to the lysis against the HSV-1-infected fibroblastoid line, M50, but the infected PU51R cells were killed by only NK effectors. These findings were consistent with the results of experiments performed to define the role of interferon in induction of virus-augmented cytolysis. Increased lysis of YAC-HSV and PU51R-HSV was entirely due to interferon activation and was completely abolished by performing the 51Cr-release assay in the presence of anti-interferon serum. Because NC activity was not augmented by interferon, virus-enhanced NC lysis of M50-HSV and WEHI-HSV was not due to this nonspecific mechanism. Together, our data show that HSV-1 infection of NK/NC targets induces increased cytotoxicity, but the effector cell responsible for lysis is determined by the uninfected target, or by an interaction between the virus and target cell, rather than by a viral determinant alone.

Animals↗
PubMed1997

How rolling circle plasmids control their copy number.

Rolling circle DNA replication is inherently continuous and unregulated. This 'go-for-broke' strategy works well for lytic phages but is suicidal for plasmids that must coexist with their host. Plasmids have consequently evolved elaborate copy number control systems that operate at the transcriptional, translational and post-translational levels.

DNA Helicases↗
PubMed2006

Identifying cellular genes crucial for the reactivation of Kaposi's sarcoma-associated herpesvirus latency.

Kaposi's sarcoma-associated herpesvirus (KSHV) is the latest addition to the long list of human herpesviruses. Reactivation of latent herpesvirus infections is still a mystery. It was demonstrated recently that the phorbol ester TPA was efficient in inducing a reactivation of KSHV infection in the S phase of the cell cycle. In the present study, flow cytometry-sorted, TPA-induced, KSHV-infected haematopoietic cells (BCBL-1) were used to analyse the expression profiles of cancer-related cellular genes in the S phase of the cell cycle compared with the G0/1 phase by using microarrays. Overall, the S phase of the cell cycle seems to provide KSHV with an apt environment for a productive lytic cycle of infection. The apt conditions include cellular signalling that promotes survivability, DNA replication and lipid metabolism, while blocking cell-cycle progression to M phase. Some of the important genes that were overexpressed during the S phase of the cell cycle compared with the G0/1 phase of TPA-induced BCBL-1 cells are v-myb myeloblastosis (MYBL2), protein kinase-membrane associated tyrosine/threonine 1 (PKMYT1), ribonucleotide reductase M1 polypeptide (RRM1) and peroxisome proliferator-activated receptors delta (PPARD). Inhibition of PKMYT1 expression by the use of specific short interfering RNAs significantly lowered the TPA-induced KSHV lytic cycle of infection. The significance of these and other genes in the reactivation of KSHV is discussed in the following report. Taken together, a flow cytometry-microarray-based method to study the cellular conditions critical for the reactivation of KSHV infection is reported here for the first time.

Cell Line, Tumor↗
PubMed1978

Genetic evidence for a temperature-sensitive lesion in the adenovirus 7 region of the PARA genome.

It has previously been shown that the helper adenovirus (Ad) present in the defective Ad 7-SV40 hybrid population is temperature-sensitive (ts) for replication in human cells. This study has shown that the replication of the defective hybrid virus (PARA) in green monkey kidney cells is also restricted at 40.5 percent. Complementation tests between the parental or transcapsidant PARA populations and SV40 or various Ad serotypes revealed that the ts lesion is located in the Ad region of the hybrid genome. Wild-type (wt) Ad type 7 and Ad type 21 were able to complement the replication of PARA while Ad type 31 was unable to do so. Complementation was more efficient after the PARA genome was transcapsidated to the wt isolates of helper Ad than during multiple infections with the parental hybrid population. The SV40 function which complements the replication of human adenoviruses in simian cells is not expressed by PARA at the nonpermissive temperature. However, the ts lesion does not affect the expression of SV40 functions coded for by PARA under other conditions, i.e., complementation of the replication of human adenoviruses in transformed monkey cells, and synthesis of T antigen in lytically infected or transformed cells. Although the exact nature of the mutated Ad 7 gene product is unknown, heat inactivation data suggest that it may be a structural protein.

Adenoviridae↗
PubMed1994

Defective infection of rabbit peripheral blood monocyte cultures with human immunodeficiency virus type 1.

Human immunodeficiency virus type 1 (HIV-1) produces abortive infections of primary cultures of rabbit peripheral blood mononuclear cells (PMBCs). Mitogen activation of rabbit PBMCs or the addition of exogenous cytokines to the cultures does not change the level of release of the early HIV core protein, p24, into the culture medium. The amount of p24 increases steadily and reaches peak levels about 7 days after infection. HIV-1-specific DNA env sequences are also detected in infected PBMCs. However, reverse transcription of RNA of samples from activated infected cultures to cDNA, followed by amplification by the polymerase chain reaction, revealed transcription of gag, but not env, regions, suggesting that HIV-1 infection of rabbit PBMC does not lead to the replication and maturation of complete HIV-1 virions. In addition, neither CPE nor lytic infection was observed in HIV-1-infected rabbit cells and infectivity could not be transferred from rabbit cells infected with HIV for up to 2 weeks to MT-2 or H9 indicator cell lines. In addition, no CPE was seen in long-term cultures of the HTLV-I-transformed rabbit cell line PLT-1441, after inoculation with HIV. It is concluded that primary rabbit PBMCs may be infectable by HIV-1 but are not permissive for production of infectious virus. This conclusion is consistent with the apparent long-term latent infection seen after inoculation of rabbits with HIV-1 or HIV-1-infected cells.

Animals↗
PubMed1983

Construction and characterization of an adenovirus type 5/adenovirus type 12 recombinant virus.

We have constructed an adenovirus type 5 (Ad5) recombinant virus in which the early region 1b (E1b) of the nononcogenic Ad5 is replaced by the E1b region of the highly oncogenic Ad12. Analysis of cells lytically infected with the recombinant virus showed that both the Ad5 E1a genes and the Ad12 E1b genes are faithfully expressed. The recombinant virus replicates efficiently in human embryonic kidney cells and in HeLa cells, indicating that the Ad12 E1b region can fully replace the Ad5 E1b region in lytic infection. Inoculation of the Ad5/Ad12 hybrid virus into newborn hamsters did not result in development of tumors. This shows that the E1b region of Ad12, previously shown to be responsible for the high oncogenic potential of Ad12-transformed cells in nude mice is not capable of converting the nononcogenic Ad5 into an oncogenic virus.

Adenoviruses, Human↗
PubMed1988

Constitutive expression in human cells of herpes simplex virus type 1 glycoprotein B gene cloned in an episomal eukaryotic vector.

Expression of herpes simplex virus type 1 (HSV-1) glycoprotein B (gB-1) was obtained in human cells from the gB-1 gene cloned in the episomal replicating vector pBK-1, which contains the origin of replication and early region of the human papovavirus BK. Selective systems for the TK+ phenotype in TK-143B cells and for resistance to G418 in adenovirus 5-transformed 293 cells were used to obtain stable transformants that produced gB-1. While gB-1 expression in 143B cells required induction by HSV-1 early proteins, constitutive gB1 production was observed in 293 cells, where endogenous trans-acting factors probably replace the need for early viral products in the activation of the cloned gB-1 gene. The amount of recombinant gB-1 was comparable to that produced during HSV-1 lytic infection in human cells, due to amplification of the inserted gene in the replicating episomal vector. Expression of gB-1 was induced by cadmium and zinc when the promoter of the mouse metallothionein-I gene was placed upstream of gB1 structural sequences. The inducible system where the gB-1 gene is under the control of its own promoter could be employed to clarify the role of early viral products in induction of gB-1 synthesis. Constitutive expression of gB-1 in human cells could provide useful material for diagnostic purposes and for the preparation of a subunit vaccine against HSV infections.

Blotting, Southern↗
PubMed1999

Phosphoantigen-reactive Vgamma9Vdelta2 T lymphocytes suppress in vitro human immunodeficiency virus type 1 replication by cell-released antiviral factors including CC chemokines.

Vgamma9Vdelta2 T lymphocytes are broadly reactive against various intracellular pathogens and display both lytic and proliferative responses to human immunodeficiency virus (HIV)-infected cells. HIV infection of peripheral blood mononuclear cell cultures led to absolute increases in Vgamma9Vdelta2 T cells accompanied by decreased p24 levels. Strong gammadelta T cell activation with nonpeptidic mycobacterial phosphoantigens (TUBAg1 extract or synthetic isopentenyl pyrophosphate) resulted in potent inhibition of HIV replication through soluble released factors. Subsequent analyses showed that phosphoantigen-activated gammadelta T cells produced substantial amounts of beta-chemokines (macrophage inflammatory protein [MIP]-1alpha, MIP-1beta, and regulated-on-activation, normal T-cell-expressed and -secreted beta-chemokine [RANTES]), which represent the natural ligand for the CCR5 HIV coreceptor. Accordingly, anti-beta-chemokine antibodies neutralized the inhibition of monocytotropic HIV strains by gammadelta T cell-released factors. Moreover, a T-tropic HIV strain using the CXCR4 coreceptor for virus entry was potently inhibited. Together, these data reveal that phosphoantigen-activated gammadelta T cells are an important source of CC chemokines and may suppress HIV replication through cell-released antiviral factors.

Antigens, Bacterial↗
PubMed1973

Relationship of replication and transcription of Simian Virus 40 DNA.

RNA produced by the Simian Virus 40 (SV40) mutant tsA30 during lytic infection of kidney cells of African green monkeys was examined by RNA-DNA competition-hybridization. This mutant is temperature-sensitive in a function (gene A) that regulates synthesis of viral DNA. No detectable difference between mutant RNA synthesized at the permissive temperature (33 degrees ) and wild-type viral RNA was found. During continuous infection with the mutant at the restrictive temperature (41 degrees ) only early viral RNA was produced. When mutant DNA and late RNA synthesis were initiated at the permissive temperature, a shift to the restrictive temperature rapidly terminated synthesis of viral DNA but not that of late viral RNA. The data indicate that the function of gene A is required before synthesis of late viral RNA and that after initiation, the production of late RNA continues without further expression of gene A or concomittant viral DNA synthesis.

Animals↗
PubMed2001

Biophysical analysis of natural variants of the multimerization region of Epstein-Barr virus lytic-switch protein BZLF1.

BZLF1 plays a key role in the induction of Epstein-Barr virus (EBV) replication. On the basis of limited sequence homology and mutagenesis experiments, BZLF1 has been described as a member of the bZip family of transcription factors, but this prospect has not been rigorously tested to date. Here, we present biophysical analysis of the multimerization domain of BZLF1, from three natural variants of EBV, and demonstrate for the first time that the region between amino acids 196 and 227 is sufficient to direct folding as a coiled-coil dimer in vitro.

Amino Acid Sequence↗
PubMed1992

Molecular characterization of a second abortive phage resistance gene present in Lactococcus lactis subsp. lactis ME2.

The fifth phage resistance factor from the prototype phage-insensitive strain Lactococcus lactis subsp. lactis ME2 has been characterized and sequenced. The genetic determinant for Prf (phage resistance five) was subcloned from the conjugative plasmid pTN20, which also encodes a restriction and modification system. Typical of other abortive resistance mechanisms, Prf reduces the efficiency of plaquing to 10(-2) to 10(-3) and decreases the plaque size and burst size of the small isometric-headed phage p2 in L. lactis subsp. lactis LM0230. However, normal-size plaques occurred at a frequency of 10(-4) and contained mutant phages that were resistant to Prf, even after repeated propagation through a sensitive host. Prf does not prevent phage adsorption or promote restriction and modification activities, but 90% of Prf+ cells infected with phage p2 die. Thus, phage infections in Prf+ cells are aborted. Prf is effective in both L. lactis subsp. lactis and L. lactis subsp. cremoris strains against several small isometric-headed phages but not against prolate-headed phages. The Prf determinant was localized by Tn5 mutagenesis and subcloning. DNA sequencing identified a 1,056-nucleotide structural gene designated abiC. Prf+ expression was obtained when abiC was subcloned into the lactococcal expression vector pMG36e. abiC is distinct from two other lactococcal abortive phage resistance genes, abiA (Hsp+, from L. lactis subsp. lactis ME2) and abi416 (Abi+, from L. lactis subsp. lactis IL416). Unlike abiA, the action of abiC does not appear to affect DNA replication. Thus, abiC represents a second abortive system found in ME2 that acts at a different point of the phage lytic cycle.

Amino Acid Sequence↗