aphidicolin and HIV-Infections

We have identified four small molecules that boost transduction of cells by human immunodeficiency virus (HIV) and investigated their mechanism of action. These molecules include etoposide and camptothecin, which induce DNA damage by inhibiting religation of cleaved topoisomerase-DNA complexes, taxol, which interferes with the function of microtubules, and aphidicolin, which inhibits DNA polymerases. All four compounds arrest the cell cycle at G2/M, though in addition high concentrations of aphidicolin arrest in G1. We find that early events of HIV replication, including synthesis of late reverse transcription products, two-long terminal repeat circles, and integrated proviruses, were increased after treatment of cells with concentrations of each compound that arrested in G2/M. Stimulation was seen for both transformed cell lines (293T and HeLa cells) and primary cells (IMR90 lung fibroblasts). Arrest in G1 with high concentrations of aphidicolin boosted transduction, though not much as with lower concentrations that arrested in G2/M. Arrest of IMR90 cells in G1 by serum starvation and contact inhibition reduced transduction. Previously, the proteasome inhibitor MG132 was reported to increase HIV infection-here we investigated the effects of combinations of the cell cycle inhibitors with MG132 and obtained data suggesting that MG132 may also boost transduction by causing G2/M cell cycle arrest. These data document that cell cycle arrest in G2/M boosts the early steps of HIV infection and suggests methods for increasing transduction with HIV-based vectors.

Topics: Aphidicolin; Camptothecin; Cell Cycle; Cell Division; Cell Line; Dose-Response Relationship, Drug; Enzyme Inhibitors; Etoposide; G2 Phase; HIV; HIV Infections; Humans; Nucleic Acid Synthesis Inhibitors; Paclitaxel; Time Factors; Transduction, Genetic; Virus Replication

Human immunodeficiency virus type 1 (HIV-1) infection of the CD4+ SupT and CEM cell lines, blocked in cell replication by the polymerase alpha inhibitor aphidicolin (APC), was studied. The APC-treated cells showed a lack of viral production, but the presence of single cell killing. High levels of unintegrated viral DNA forms were found in the infected APC-treated cells as compared with untreated cells. Moreover, an increased rate of viral replication occurred in the remaining viable cells following removal of APC. The results indicate that HIV-1 entry and reverse transcription can take place in cells blocked in the S phase of the cell cycle. Replication of infectious progeny virions appears to require de novo cell division. Finally, accumulation of viral DNA in cells during APC treatment can result in cytopathological effects and subsequent enhancement of virus production.

Topics: Aphidicolin; Cell Death; Cell Division; Cell Line; DNA, Viral; HIV Infections; HIV-1; Humans; S Phase; T-Lymphocytes; Virion; Virus Replication