The human cytomegalovirus (HCMV) tegument phosphoprotein pp71 activates viral immediate early (IE) transcription and thus has a role in initiating lytic infection. Protein pp71 stimulates expression from a range of promoters in a sequence-independent manner, and in this respect behaves similarly to the herpes simplex virus type 1 (HSV-1) IE protein ICP0. The intracellular localization of pp71 was investigated after its expression from transfected plasmids or from HSV-1 mutants constructed to produce pp71 transiently. The protein colocalized with the cell promyelocytic leukaemia (PML) protein at nuclear domain 10 (ND10) structures but, unlike ICP0, pp71 did not induce disruption of ND10. The activity of pp71 in mouse sensory neurons in vivo was investigated after co-inoculation of animals with pairs of HSV-1 mutants, one expressing pp71 and the second containing the E. coli lacZ gene controlled by various promoters. In this system, pp71 stimulated β-galactosidase expression from a range of viral IE promoters when mice were analysed at 4 days postinoculation. At later times, expression of pp71 resulted in a reduction in numbers of neurons containing β-galactosidase, indicating cytotoxicity or promoter shutoff. The HSV-1 latency-active promoter was not responsive to pp71, demonstrating specificity in the activity of the protein. Pp71 was as active in mice lacking both copies of the PML gene (PMLN/N) as in control animals, and in PMLN/N fibroblasts pp71 stimulated gene expression as effectively as in other cell types. Therefore, neither the PML protein nor the normal ND10 structure is necessary for pp71 to stimulate gene expression.

These HHLs retain primary hepatocyte phenotype and should be useful for investigating mechanisms of entry and replication of hepatotropic viruses, and should also be valuable in the study of hepatocyte biology and pathology.

Herpes simplex virus type 1 (HSV-1) transcription can be arrested at the immediate early (IE) stage by continuous treatment of cells with inhibitors of protein synthesis, usually cycloheximide, from the time of infection. We have analysed the effect of cycloheximide on IE gene expression with HSV-1 mutants deficient in the production of functional levels of the three major transactivators, the virion protein (VP16) and two IE proteins (ICP0 and ICP4). Expression from the HSV-1 IE promoters that control synthesis of ICP0 and ICP27 was, unexpectedly, stimulated by inhibition of protein synthesis. The effect was observed for the ICP0 promoter in its normal genome location and also when cloned upstream of the Escherichia coli lacZ coding sequences and inserted into the viral thymidine kinase locus. Expression from the human cytomegalovirus

Recent reports suggest an association between Chlamydia pneumoniae and Helicobacter pylori bacteria and atherosclerosis. We studied 51 patients (mean age, 68.3 years) who underwent abdominal aortic aneurysm surgery. For each patient we performed a microimmunofluorescence test for immunoglobulin G (IgG), IgA, and IgM antibodies to C. pneumoniae specific antigen (TW-183). Anti-H. pylori antibodies were determined by means of an EIA-G test. Each aortic aneurysm surgical specimen was sampled into multiple sections of 0.3 cm 2 each and frozen at ؊20؇C. Two samples of each aneurysm were used for a nested PCR with two sets of C. pneumoniae and two sets of H. pylori specific primers. Specimens were treated with a solution containing 20 mM Tris-HCl, Tween 20-Nonidet P-40 (0.5% [vol/vol] each), and 100 g of proteinase K per ml and incubated at 60؇C for 1 h and at 98؇C for 10 min. DNA was extracted twice with phenol-chloroform-isoamylic alcohol and precipitated with sodium acetate-ethanol by standard methods. Forty-one patients were seropositive for C. pneumoniae with past-infection patterns in 32 patients (16 < IgG < 512; 32 < IgA < 256) and high antibody titers in 9 patients (IgG > 512). In 26 of 51 patients, C. pneumoniae DNA was detected in aortic aneurysm plaque specimens. Of these patients, 23 had a serologic past-infection pattern, 2 had an acute reinfection pattern, and 1 was seronegative. Forty-seven of 51 patients were seropositive for H. pylori. In all cases PCR showed no evidence of H. pylori presence in plaque specimens. This study provides data on a possible C. pneumoniae involvement in the pathogenesis of aortic aneurysm and additional evidence for an association between this agent and atherosclerosis. Conversely, notwithstanding a high H. pylori seroprevalence observed, our results tend to rule out the possibility of a direct involvement of H. pylori in atherosclerosis.

The role of viral immediate-early (IE) gene expression in herpes simplex virus type 1 (HSV-1) latency was investigated. The HSV-1 multiple mutant in1312, defective for the expression of the virion transactivator VP16 and the IE proteins ICP0 and ICP4, was used as the parent for these studies. The coding sequences of the Escherichia coli lacZ gene, preceded by the encephalomyocarditis virus internal ribosome entry site, were inserted into the region of in1312 that encodes the latency-associated transcripts (LATs) such that transcription of the transgene was controlled by the LAT promoter. This insert has previously been shown to direct long-term latent-phase expression of ␤-galactosidase in a wild-type HSV-1 genome (R. H. Lachmann and S. Efstathiou, J. Virol. 71, 3197-3207, 1997). The resulting recombinant, in1388, was apathogenic after inoculation into mice via the footpad and did not detectably replicate in dorsal root ganglia (DRG) or footpads. Infection with herpes simplex virus type 1 (HSV-1) normally results in productive replication of virus and death of the host cell. Neurons, however, are able to survive infection and retain the HSV-1 genome in a latent state for the lifetime of the host. Reactivation of latent virus and, in some instances, reappearance of disease occur in response to stimuli that cause stress to the neuron or to the host organism (reviewed in references 43,54,60).Transcription of the HSV-1 genome is largely controlled by the immediate-early (IE) proteins ICP4 (Vmw175) and ICP0 (Vmw110) and by the virion protein VP16 (Vmw65 or ␣-TIF). ICP4 is a transcription activator that is absolutely required for productive infection. Early and late gene transcription does not occur after infection with virus mutants lacking functional ICP4 (10,38,62). ICP0 alters the intranuclear environment such that entry of HSV-1 into the lytic cycle is facilitated (17,18). Infection at low multiplicity of infection (MOI) with viruses possessing mutations that inactivate ICP0 results in only a small proportion of infected cells supporting replication and most viral genomes being retained in a quiescent state (16,40,44,45,56,57). The absence of ICP0 can, however, be overcome by carrying out infection at a high MOI (16,44,57). Transcription of the IE genes is stimulated by VP16, a component of the incoming virus particle which interacts with the cell factors Oct-1 and HCF to form a multiprotein complex at the TAATGARAT (R is a purine nucleotide) sequences found in all IE promoters (reviewed in reference 36). The virus mutant in1814, which expresses nonfunctional VP16, exhibits a phenotype similar to that of ICP0 mutants, with most cells infected at low MOI failing to initiate early or late gene expression and retaining the HSV-1 genome in a quiescent state (1, 23). The absence of functional ICP4, ICP0, or VP16 therefore arrests the HSV-1 lytic cycle at early stages.During latency in humans or animals, gene expression characteristic of productive replication cannot be detected; instead only one portion of the genome,...