During progression of the Acquired Immune Deficiency Syndrome (AIDS), the human immunodeficiency virus type 1 (HIV‐1) is harbored in CD4+ T cells, which act as the primary reservoir for the virus. In vitro, HIV‐1 requires activated T cells for a productive infection; however, in vivo, the number of circulating T cells in the activated state that are potential targets for HIV‐1 infection is low. We have investigated the ability of HIV‐1 to infect resting T cells, and the consequences of such an infection. T cell activation was not required for HIV‐1 infection; however, viral DNA was unable to integrate in resting T cells and was maintained extrachromosomally. Subsequent T cell activation allowed integration of extrachromosomal forms and led to a productive viral life cycle. Extrachromosomal forms of viral DNA were found to persist for several weeks after infection of resting T cells and, following T cell activation, these forms maintained their ability to integrate and act as a template for infectious virus. Several lines of evidence, including temporal analysis of HIV‐1 replication and analysis of an HIV‐1 integrase deletion mutant, indicated that extra‐chromosomal HIV‐1 DNA genomes were transcriptionally active. These results are compatible with a model whereby HIV‐1 can persist in a non‐productive extra‐chromosomal state in resting T cells until subsequent antigen‐induced or mitogen‐induced T cell activation, virus integration and release. Thus agents that induce T cell activation may control the rate of HIV‐1 replication and spread during AIDS progression.

After cell infection by the human immuno deficiency virus type 1 (HIV-1), nascent viral DNA in the form of a high molecular weight nucleoprotein preintegration comin plex must be transported to the nucleus of the host cell prior to integration of viral DNA with the host genome. The mechm used by retroviruses for nuclear targeting of preintegration complexes and dependence on cell division has not been established. Our studies show that, after infection, the preintegration complex of HIV-1 was rapidly transported to the nucleus of the host cell by a process that required ATP but was independent of cell division. Functional HIV-1 integrase, an essential component of the preintegration complex, was not required for nuclear import of these complexes. The ability of iHV-1 to use host cell active transport processes for nuclear import of the viral preintegration complex obviates the requireenlt for host cell division in establishment of the integrated provires. These findgs are pertinent to our underding of early events in the life cycle of HIV-1 and to the mode of HIVE1 replication in terminally differentiated nondividing cells such as macrophages (monocytes, tissue macrophages, follicular dendritic cells, and microglial cells).Integration of the retroviral genome with cellular DNA and establishment of the provirus is an essential step in retrovirus replication (1). The integration reaction is catalyzed by a virus-encoded integrase, which is derived from the virus particle and which, after reverse transcription of genomic viral RNA, remains associated with the viral cDNA in a high molecular weight nucleoprotein preintegration complex (2). Targeting of the viral preintegration complex to host cell DNA is therefore dependent on transport of this complex to the nucleus of the host cell. The process that directs nuclear localization of retroviral preintegration complexes after infection and the dependence ofthis process on cell division are unknown. Oncogenic HIV-1-infected cells were lysed in hypotonic medium (6) using multiple strokes of a Dounce homogenizer, and nuclear integrity during cell lysis was monitored by phase-contrast microscopy. Nuclei were extracted with a hypertonic buffer (6) and both nuclear and cytoplasmic extracts were fractionated on nonionic density gradients as described (2). Integration activity in each gradient fraction was analyzed in vitro by a modification of a previous protocol (7). Briefly, 100 ul of each gradient fraction was mixed with 1 pug of ir AN7 target DNA (8) in a reaction volume of 150 Al and was incubated 60 min at 220C. Samples were treated with DNA polymerase I and deproteinated; in vitro integration products were identified by two rounds of PCR with nested HIV-1 U5 and R long terminal repeat (LTR) primers. PCR products were visualized by Southern blot hybridization with 32P-end-labeled oligonucleotide probes as described elsewhere (4).PCR Analysis of HIV-1 DNA in Nuclear and Cytoplasmic Cell Extracts. Cells were washed once in ice-cold phosphatebuffered saline (pH 7....

To better understand the basis for human immunodeficiency virus type 1 (HIV-1) persistence and latency, the form in which viral DNA exists in the peripheral T lymphocyte reservoir of infected individuals was investigated. In asymptomatic individuals, HIV-1 was harbored predominantly as full-length, unintegrated complementary DNA. These extrachromosomal DNA forms retained the ability to integrate upon T cell activation in vitro. In patients with acquired immunodeficiency syndrome (AIDS), there was an increase in integrated relative to extrachromosomal DNA forms. By analysis of DNA from patient lymphocyte subpopulations depleted of human lymphocyte antigen-Dr receptor-positive cells, quiescent T cells were identified as the source of extrachromosomal HIV-1 DNA. Thus quiescent T lymphocytes may be a major and inducible HIV-1 reservoir in infected individuals.

Acyclovir and human interferon displayed an additive to synergistic effect in reducing the number of herpes plex viral plaque-forming units in Vero cells, suggesting a therapeutic potential for such combination therapy.The promising new antiherpes compound, 9-(2-hydroxyethoxymethyl)guanine, referred to as acyclovir, has been found to have marked antiviral activity in animal models (5,8,9,10,12) and is presently being evaluated in a series of clinical tials. Human interferon (Hu IFN) is also currently being studied for the prevention or treatment of local and systemic herpes simplex infections (3,13 Both HSV-1 and HSV-2 were less susceptible to the antiviral action of IFN than was VSV (Fig. 1). Also, as previously reported (11,14), inhibition of plaques by IFN was more pronounced with HSV-1 than with HSV-2. The observed effects of IFN in combination with acyclovir were compared with the expected values. The latter were calculated by multiplying the fraction of viral control plaques (FP)

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