TRANSCRIPTIONAL EFFECTS OF SUPERINFECTION IN HIV CHRONICALLY INFECTEDT-CELLS - STUDIES IN DUALLY INFECTED CLONES

We had previously shown that chronically infected ACH-2 cells (HIVLAI) could be superinfected with HIVRF, that the frequency of superinfecti on increased with time, and that the transcription of the superinfecti ng virus exceeded that of the host HIVLAI provirus. In contrast, ACH-2 cells superinfected with a nef-substituted neomycin-resistant (proNEO ) provirus were not detectable by DNA polymerase chain reaction (PCR) until geneticin (G418) was added, suggesting that the ability to propa gate progressively in culture may be HIV strain specific. Clonal popul ations of ACH-2 superinfected with proNEO did not demonstrate preferen tial transcription of the superinfecting virus. However, clones of ACH -2 superinfected with HIVRF (ACH2/RF) showed a preponderance of HIVRF transcripts similar to that seen in bulk populations. Induction of the superinfecting virus by phorbol ester (PMA) occurred more rapidly tha n the host provirus and did not equalize transcriptional activity. PCR -derived long terminal repeat (LTR) fragments and Tat cDNAs from A3.01 cells acutely infected with HIVRF or from ACH-2 cells were sequenced and tested for transactivation. The HIVLAI LTR was two to three times more Tat-responsive than the HIVRF LTR. Tat(RF) was two to three times more transcriptionally active on either LTR than Tat(LAI). Demethylat ion with 5-azacytidine did not significantly affect HIV expression fro m the HIVLAI host provirus of superinfected ACH2/RF cell clones. These data suggest that the mechanism of preferential transcription in HIVR F superinfected ACH2/RF may be attributed to the Tat/TAR axis and the effect of the specific locus of host proviral integration.