AEGiS-11IAC: CD8+ T cell culture fluids from HIV-infected individuals suppress HIV-1 long terminal repeat (LTR) driven transcription.

11th International AIDS Conference

Vancouver, British Columbia — July 7-12, 1996

CD8+ T cell culture fluids from HIV-infected individuals suppress HIV-1 long terminal repeat (LTR) driven transcription.

Int Conf AIDS 1996 Jul 7-12; 11:211 (abstract no. Th.A.105)
Sato A, Mackewicz CE, Gaynor RH, Levy JA; Cancer Research Institute, University of California-San Francisco, San Francisco, CA, USA.

OBJECTIVE: CD8+ cells from HIV-infected individuals suppress HIV replication in cultured CD4+ cells by a noncytolytic mechanism that involves a secreted CD8+ cell antiviral factor (CAF). To elucidate the mechanism of the anti-HIV effect of CAF, we examined the inhibitory effect of CAF on transcriptional activity of HIV-1 LTR directing the chloramphenicol acetyltransferase (CAT) gene.

METHODS: Culture fluids containing CAF were obtained from stimulated purified CD8+ T cells recovered from the blood of HIV-infected Long-term survivors (LTS). CD8+ cell culture fluids lacking CAF activity came from HIV-infected Progressors. The cells were cultured in serum-free medium and fluids were collected every 2 days. CAF content was determined by the inhibitory effect of the fluid on HIV-1 replication in acutely infected CD4+ cells or 1G5 cells. The 1G5 cell line is a derivative of Jurkat T cells, containing a stable LTR-luciferase construct. Jurkat cells were transfected with HIV-1 LTR-CAT and SV40-tat plasmids and subsequently grown in a 50% of the culture fluids containing CAF. After 48 hours, the extent of CAT production was measured by ELISA. LTR mutants were also used to evaluate further the mechanism of HIV inhibition.

RESULTS: Six CAF-containing fluids have been evaluated. Five fluids suppressed CAT activity up to 40 % and one fluid showed no inhibitory effect. In the 1G5 cell line, these CAF-containing fluids suppressed induction of luciferase by at least 50%. Culture fluids obtained from CD8+ cells of Progressors lacked CAF activity and enhanced CAT production; they had no effect on luciferase expression. Preliminary studies using LTR-CAT with NFkB element or SP-1 binding site mutations have suggested that CAF has differential effects on regulatory elements within the LTR. Further studies with these LTR mutants are in progress to define HIV-1 regulatory elements that may be targeted for CAF-mediated decrease in gene expression.

CONCLUSIONS: The noncytolytic CD8+ cell antiviral activity mediated by CAF exerts its effect, at least in part, by specifically interrupting HIV transcription via genetic elements within the HIV LTR.

Keywords: AEGIS, HIV Long Terminal Repeat, Antigens, CD8, HIV, T-Lymphocytes, Transcription, Genetic, HIV-1, Virus Replication, Jurkat Cells, HIV Infections, Cell Line, HIV-1 Reverse Transcriptase, Culture, Reverse Transcriptase Inhibitors, Human, Cats, Animal, ethnology, genetics, virology, ICA11

960707
ThA105