Search PubMed⌕ Search

Biomedical subjects

A Brandenburger

Publications and source records attributed to A Brandenburger.

8 recordsLinked to original sources
PubMed1992

Transformation-dependent expression of interleukin genes delivered by a recombinant parvovirus.

The prototype strain of minute virus of mice [MVM(p)] is an autonomous parvovirus with a tropism for cells expressing a neoplastically transformed phenotype. To generate gene transfer vectors for tumor-specific gene expression, human interleukin-2 (IL-2) and murine interleukin-4 (IL-4) genes were cloned under the control of the p38 late promoter of MVM(p). Upon transfection into permissive cells, the recombinant MVMIL2 or MVMIL4 DNA was excised, amplified, and, in the presence of a helper plasmid, packaged into recombinant viral particles. The recombinant viruses were able to transfer fully functional IL-2 and IL-4 genes to permissive target cells and retained the oncotropic host range properties of the parental virus. Following infection with MVMIL2, nontransformed fibroblasts of rodent (FR3T3) or human (MRC-5) origin produced minimal IL-2 compared with the high levels of IL-2 production observed in their transformed derivatives (FREJ4 and MRC-5V1).

Cell Line, Transformed↗
PubMed1990

Programmed killing of human cells by means of an inducible clone of parvoviral genes encoding non-structural proteins.

Although its dependence on the target cell type is well established, the cytopathogenicity of parvoviruses has remained elusive to date as far as its mechanism is concerned. However, indirect evidence suggested that parvoviral non-structural (NS) proteins may be the cytotoxic effectors. In order to test this hypothesis, a molecular clone of parvovirus MVMp was modified, by replacing the P4 promoter of the NS transcription unit by the glucocorticoid-inducible promoter of the mouse mammary tumour virus. Clones of neoplastic human cells that had incorporated this construct and that were induced to produce NS proteins by dexamethasone, showed a cytopathic effect and eventually died. Our data strongly suggest that the intracellular accumulation of parvoviral NS products jeopardizes the survival of the cells, which cannot be detected unless a threshold protein concentration is reached. Interestingly, a cell variant could be isolated which resisted dexamethasone-induced killing, although it was fully inducible for the production of NS proteins. This variant was also unusually resistant to infection with MVMp virions, thus confirming the essential role played by the NS proteins in the parvoviral cytotoxicity and indicating that the cytocidal activity of the parvoviral NS products is modulated by cellular factors that may vary from one cell to another.

Capsid↗
PubMed1990

NS-1 and NS-2 proteins may act synergistically in the cytopathogenicity of parvovirus MVMp.

The interaction of parvovirus minute virus of mice (prototype strain, MVMp) with simian virus 40 (SV40)-transformed human cells (NB-E) was investigated by means of transfection with MVMp molecular clones derived from the infectious recombinant plasmid (pMM984). pMM984 inhibits stable transformation of NB-E cells to geneticin resistance (G418R) upon cotransfection with the selectable pSV2neo plasmid. We show here that this inhibition is not merely caused by a repression of marker gene expression from the SV40 early region promoter in pSV2neo and rather is likely to reflect the cytotoxic action of the parvovirus. Starting from plasmid pMM984, defined mutations were introduced into the genome of MVMp and more particularly into sequences coding for the NS-1 and/or NS-2 nonstructural proteins. In this way we could show that the NS-1 protein is necessary for the inhibition of transformation to G418R and that the NS-2 protein acts synergistically to enhance this effect. Moreover, results obtained with different viral mutants indicate that the inhibitory action of NS-1 on stable transformation can be dissociated from the ability of this protein both to transactivate the parvoviral p39 promoter of the capsid protein-encoding region and to drive parvoviral DNA amplification. Altogether these data point to a probable direct toxicity of MVMp nonstructural proteins for permissive host cells.

Capsid↗
PubMed1984

Indirect SOS induction is promoted by ultraviolet light-damaged miniF and requires the miniF lynA locus.

Indirect prophage induction is produced by transfer to recipients of u.v.-damaged F plasmid (95 kb). We tested whether the SOS signal can be produced by miniF, a 9.3 kb restriction fragment, coding for the replication and segregation functions of plasmid F. We used lambda miniF, a hybrid phage-plasmid. u.v.-irradiated lambda miniF induced prophages phi 80 or lambda and sfiA, a chromosomal SOS gene, in more than 50% of the infected cells. The maximal inducing dose produced about 0.5 pyrimidine dimers per kb and left 1% of lambda miniF survivors. Thus, the SOS signal produced by u.v.-damaged lambda miniF was almost as potent as that resulting from direct u.v.-irradiation of the lysogens. The u.v.-damaged vector lambda, devoid of miniF, failed to promote SOS induction. In contrast, efficient induction was observed when u.v.-damaged lambda miniF infected a lambda immune host, in which replication and expression of the phage genome were repressed. When replication and expression of the miniF genome was repressed by Hfr incompatibility, SOS induction was largely prevented. All these facts indicate that, in the hybrid lambda-miniF, it is the u.v.-damaged miniF that generates an SOS signal. To locate on the miniF genome the loci that are involved in the production of the SOS signal, we isolated deletions spanning all the miniF restriction fragments. We characterized six mutant phenotypes (Par+, Rep-, Fid-, Par-2, Par-1 and SOS-) related to four functions; partition, copy number, replication and SOS induction. A locus, we call lynA, 800 bp long, located by deletion mapping between the two origins of replication oriP and oriS is required for the production of an inducing signal. We postulate that indirect SOS induction by u.v.-damaged miniF results from the disturbance of the lynA function that may be involved in the co-segregation of F plasmid with the host chromosome.

Bacteriophage lambda↗
PubMed1984

Gratuitous induction.

We describe a novel mode of SOS induction, called gratuitous indirect induction, which is elicited when the maintenance of an intact lambda miniF introduced into a recipient was inhibited by a resident plasmid or by mutations in miniF that impaired partition or replication. Gratuitous induction required the presence of the lynA locus on miniF and was dependent on the host recA and lexA alleles. To account for gratuitous induction, we postulate that impairment of the normal co-regulation between partition and replication of miniF affects lynA functions whose disturbance leads to the production of an SOS signal.

Bacteriophage lambda↗
PubMed1978

Requirement of protein and RNA synthesis for lambda repressor inactivation by tif-1: effects of chloramphenicol, neomycin and rifampicin.

The inactivation of lambda repressor was followed by the specific DNA binding assay during the course of lysogenic induction provoked by incubation at 42 degrees C of an E. coli tif-1 lysogenic strain. The presence of up to 400 microgram/ml chloramphenicol during the inducing treatment did not impair the loss of repressor binding activity, whilst concentrations of 200 microgram/ml neomycin and 100 microgram/ml rifampicin effectively inhibited the inactivation of lambda repressor. Residual protein synthesis in the presence of chloramphenicol, neomycin and rifampicin was 5%, 5% and 27% respectively of that observed in the drug-free control. This residual synthesis did not appear to involve amplification of the X-protein. These results suggest that tif-mediated inactivation of the lambda repressor requires the activation of some specific gene(s), the translation of which appears to be resistant to chloramphenicol.

Autoradiography↗