A putative latency promoter/enhancer (PLAT2) region of pseudorabies virus contains a virulence determinant (original) (raw)
Related papers
Microbes and Infection, 2000
Early protein 0 (EP0)-deficient recombinant Aujeszky's disease viruses, Ka-ep0lac and Ba-ep0lac derived from strains Kaplan and Bartha, respectively, were constructed to explore the impact of the mutation on replication, virulence and latency of the virus. Inactivation of the EP0 gene resulted in a mutation of long latency transcript that is located on the complementary DNA strand of EP0 and immediate early protein (IE)175 genes. In infection of immortalized porcine kidney cells, the growth rate and yield of both EP0mutant strains were significantly smaller than that of wild-type virus. Ka-ep0lac was found to be highly virulent, while Ba-ep0lac showed an attenuated phenotype in mice. PCR assay and immunohistochemistry showed that the Ba-ep0lac virus was able to establish latency in the mouse trigeminal ganglia. However, latent virus was not able to reactivate in explant reactivation assays. Accordingly, latent Ba-ep0lac has the potential to be exploited as vectors for the delivery of foreign genes to the nervous system. © 2000 Éditions scientifiques et médicales Elsevier SAS herpesvirus / latency / pseudorabies virus / early protein 0 / LLT
Journal of virology, 1999
The pseudorabies virus (PRV) gE gene encodes a multifunctional membrane protein found in infected cell membranes and in the virion envelope. Deletion of the gE gene results in marked attenuation of the virus in almost every animal species tested that is permissive for PRV. A common inference is that gE mutants are less virulent because they have reduced ability to spread from cell to cell; e.g., gE mutants infect fewer cells and, accordingly, animals live longer. In this report, we demonstrate that this inference does not hold in a rat experimental model for virus invasion of the brain. We find that animals infected with gE mutants live longer despite extensive retrograde, transneuronal spread of virus in the rat brain. In this model of brain infection, virus is injected into the stomach musculature and virions spread to the brain in long axons of brain stem neurons that give rise to the tenth cranial nerve (the vagus). The infection then spreads from neuron to neuron in well-define...
Virus genes, 1998
A recombinant pseudorabies (Aujeszky's disease) virus (PrV) designated as vE16lac was constructed by deleting a 3-kbp DNA segment spanning the junction of long and short components of the viral genome, and by replacing the deleted segment with a lacZ-expression cassette. The aim of constructing this mutant was (a) to determine whether the terminal repeat (Tr) can serve as a template for the regeneration of the internal repeat (Ir), and (b) whether this deletion causes a reduction in the neuroinvasiveness of the virus. To analyze the mechanism of equalization, revertant viruses were selected and structurally characterized from vE16lac infection of PK-15 cells, mice and pigs. Because all revertants acquired Ir sequences identical to that of the wild-type virus, the equalization process occurred using the Tr as a template to reconstitute the Ir. We also found that the recombinant virus vE16lac was virulent in both pigs and mice. The data are discussed in view of studies performed w...
The Journal of Infectious Diseases, 2002
The neurotropic a-herpesviruses are common mammalian pathogens that invade the peripheral and central nervous system of their hosts. Their ability to invade and spread in the nervous system in a directional manner has been exploited to develop them as neuronal circuit tracers. Tracing viruses spread among synaptically connected neurons and, by assaying brain sections for viral antigen or reporter genes expressed from the viruses, chains of synaptically connected neurons can be visualized. Virulent field strains generally are not good tracers, but some attenuated strains perform well. Live attenuated vaccine strains of pseudorabies virus (PRV), such as PRV Bartha, are among the most popular virus circuit tracers. It may be counterintuitive that attenuation results in improved neural tracing that requires extensive replication and spread in the brain. This report summarizes two lines of experiments directed to resolving this apparent paradox and introduces a new paradigm for tracing viruses. a-Herpesviruses, such as herpes simplex virus types 1 and 2, varicella-zoster virus, and pseudorabies virus (PRV), are novel parasites of their host's peripheral nervous system (PNS) (see [1-3]). This tropism for the PNS is remarkable because the viruses are pantropic, capable of engaging multiple receptors and of infecting most cells in the body. In fact, many aspects of aherpesvirus pathogenesis reflect this broad range of cell tropism. However, during infections of a healthy natural host, the primary infection is resolved and viruses invariably invade the PNS where they establish their genomes as quiescent extra chromosomes in neurons in peripheral ganglia (the latent state). Reactivation from latency is associated with viral replication in the primary neuron followed by anterograde transport of newly synthesized virus particles from the neuronal cell body back to the innervated tissue. Although less frequent, virus may also spread from the PNS to the central nervous system (CNS) following primary infection or reactivation with serious, often fatal, consequences for the host. This directional movement of virus to PNS cell bodies during infection and away from the cell bodies during reactivation is noteworthy and suggests that viral genes control direction (reviewed in [4]). Similarly, the relative, but not absolute, inability to cross into the CNS in the natural host, but not in permissive alternative hosts, is of considerable interest. The ability of viruses like PRV to spread efficiently from the Protocols were approved by the Princeton University Animal Welfare Committee and were consistent with regulations of the American Association for Accreditation of Laboratory Animal Care and of the Animal Welfare Act (public law 99-198). Grant support: NIH (RO1 33506).
Virology, 1990
The immediate-early (IE) gene of pseudorabies virus (PRV) has recently been sequenced for two virus strains. To investigate IE gene regulation and to examine the genome segment reported to encode latency-related transcripts in opposite polarity to the IE gene, sequence analysis has been extended by 5 kb from each end of the IE gene. The IE promoter (P1) was found to be more complex than previously recognized: it consisted of nine imperfect repeats, each containing five to six different consensus elements for transcription factor binding. A second promoter (P2) was discovered downstream of the IE gene. It contained numerous octamer consensus sequences (ATGCAAAT) and recognition sites for transcription factor Sp1; specific binding of nuclear proteins to four Sp1 sites was detected. An open reading frame (ORF3) bordering on P2 was identified, oriented antiparallel to the IE gene. Potential enhancer elements (E3 and E4) were isolated by the enhancer trap technique. Linked to P1 and a CAT indicator gene, E3 acted as an enhancer and E4 as a silencer. The PRV IE gene product repressed transcription from its own promoter and activated the SV40 early promoter. The transactivating virion protein Vmw65 of HSV1 had an opposite effect on these promoters.
Brain research. Molecular brain research, 2002
In this study we have modified the neuroinvasiveness of pseudorabies virus strain Bartha, a commonly utilized trans-synaptic tract-tracer. In addition, we sought to facilitate detection of cellular mRNAs in neurons infected with the virus. In order to modify spreading characteristics, we inserted the lacZ or the GFP (green fluorescent protein) genes into the genomic loci containing the putative latency-associated transcript promoter (P(LAT2)), resulting in the disruption of the promoter function. Following rat kidney injection, mutant viruses labeled central autonomic neurons in a slower and much more restricted manner than the parent Bartha strain. Since both reporter genes were controlled by the human cytomegalovirus immediate early (IE) 1 promoter, they exhibited IE expression kinetics. This property proved to be important for the co-detection of reporter proteins with neuronal mRNAs, readily detected at early but not at late stage of infection, as shown in tyrosine-hydroxylase e...
Journal of Virology, 1999
Neurotropic alphaherpesviruses have become popular tools for transynaptic analysis of neural circuitry. It has also been demonstrated that coinfection with two viruses expressing unique reporters can be used to define more complicated circuitry. However, the coinfection studies reported to date have employed nonisogenic strains that differ in their invasive properties. In the present investigation we used two antigenically distinct recombinants of the swine pathogen pseudorabies virus (PRV) in single and double infections of the rat central nervous system. Both viruses are derivatives of PRV-Bartha, a strain with reduced virulence that is widely used for circuit analysis. PRV-BaBlu expresses β-galactosidase, and PRV-D expresses the PRV membrane protein gI, the gene for which is deleted in PRV-BaBlu. Antibodies to β-galactosidase identify neurons infected with PRV-BaBlu, and antibodies monospecific for PRV gI identify neurons infected with PRV-D. The ability of these strains to estab...
Transcriptome Signature of Virulent and Attenuated Pseudorabies Virus-Infected Rodent Brain
Journal of Virology, 2006
Mammalian alphaherpesviruses normally establish latent infections in ganglia of the peripheral nervous system in their natural hosts. Occasionally, however, these viruses spread to the central nervous system (CNS), where they cause damaging, often fatal, infections. Attenuated alphaherpesvirus derivatives have been used extensively as neuronal circuit tracers in a variety of animal models. Their circuit-specific spread provides a unique paradigm to study the local and global CNS response to infection. Thus, we systematically analyzed the host gene expression profile after acute pseudorabies virus (PRV) infection of the CNS using Affymetrix GeneChip technology. Rats were injected intraocularly with one of three selected virulent and attenuated PRV strains. Relative levels of cellular transcripts were quantified from hypothalamic and cerebellar tissues at various times postinfection. The number of cellular genes responding to infection correlated with the extent of virus dissemination...
Veterinary Microbiology, 1998
Pseudorabies virus (PRV) early protein 0 (EP0) functions as a transactivator of the viral gene promoters. In transient expression assays employing chloramphenicol acetyl transferase (CAT) reporter constructs, EP0 and the immediate-early protein IE180 act in an additive manner to activate transcription from the thymidine kinase (TK) and glycoprotein G (gG) gene promoters. EP0 enhanced the synthesis of infectious virus in cotransfection experiments with the EP0-expression plasmid and PRV genomic DNA. EP0 was detected by Western blot analysis in the purified virions. These results may indicate that EP0 in the virions acts as an important transactivator to express the immediate-early gene efficiently in the first stage of infection, and IE180 and EP0 expressed after the infection cooperatively activate the early and late gene expression in the later stage of infection.
Influence of Pseudorabies Virus Proteins on Neuroinvasion and Neurovirulence in Mice
Journal of Virology, 2006
Neurotropism is a distinctive feature of members of the Alphaherpesvirinae. However, its molecular basis remains enigmatic. In the past, research has been focused mainly on the role of viral envelope proteins in modulating herpesvirus neuroinvasion and neurovirulence (T. C. Mettenleiter, Virus Res. 92:192-206, 2003). To further analyze the molecular requirements for neuroinvasion of the alphaherpesvirus pseudorabies virus (PrV), adult mice were infected intranasally with a set of single-or multiple-deletion mutants lacking the UL3, glycoprotein E (gE), gM, UL11/US9, UL11/UL16, UL16/UL21, UL11/UL16/UL21, UL11/gE, UL11/gM, UL43/gK, UL43/gM, or UL43/gK/gM genes. Neurovirulence was evaluated by measuring mean survival times compared to that after wild-type virus infection. Furthermore, by immunohistochemical detection of infected neurons, the kinetics of viral spread in the murine central nervous system was investigated.