Interleukin-6 and glucocorticoids synergistically induce human immunodeficiency virus type-1 expression in chronically infected U1 cells by a long terminal repeat independent post-transcriptional mechanism

Background: Glucocorticoids (GC) such as dexamethasone (Dex) can directly u pregulate human immunodeficiency virus type-1 (HIV-1) replication in acutel y infected cells and potentiate HIV expression from chronically infected pr omonocytic UI cells stimulated with tumor necrosis factor-alpha (TNF-alpha) . We have here investigated the potential effect of Dex in UI cells stimula ted with interleukin-6 (IL-6), a cytokine inducing virus expression by acti ng mostly at a post-transcriptional level on the virus life cycle. Materials and Methods: Virus production in culture supernatants was evaluat ed by reverse transcriptase (RT) activity. GC receptor expression was teste d by both binding of [H-3]-Dexamethasone 21-mesylate and Northern blotting. Cell-associated HIV protein expression was analyzed by Western blotting, w hereas both HIV and monocyte chemoattractant protein-1 (MCP-1) RNA accumula tion were evaluated by Northern blotting. HIV transcription was tested by l ong terminal repeat (LTR) chloramphenicol acetyl transferase (CAT) assay af ter transient transfection of UI or U937 cells. Formation of activating pro tein-1 (AP-1) DNA binding complex in nuclear cell extracts was visualized b y electrophoretic mobility shift assay (EMSA), whereas ERK1/2 mitogen-activ ated protein kinase (MAPK) phosporylation was studied by Western blotting. Results: IL-6 and Dex synergistically induced HIV expression in U I cells, and this effect was blocked by RU 486. No substantial HIV RNA accumulation was demonstrated in UI cells co-stimulated with IL-6 and Dex, whereas IL-6 upregulated the expression of MCP-I RNA, and this effect was inhibited by D ex. In contrast, Dex potentiated IL-6 induced activation of AP-1 and ERK1/2 MAPK phosphorylation, as revealed by EMSA. HIV-1 LTR driven transcription was observed in U1 cells stimulated with TNF-alpha and this effect was pote ntiated by Dex. In sharp contrast, no induction of LTR-directed CAT activit y was observed in transfected UI cells (or in their parental uninfected U93 7 cells) stimulated with IL-6 and Dex either alone or in combination. Conclusions: High levels of virion production can be induced in latently in fected cells by stimulation with IL-6 and Dex in the absence of activation of the HIV LTR or viral transcription in spite of activation of both ERK1/2 MAPK and AP-1. These findings suggest the existence of LTR-independent pat hways influenced by cytokine and GC through which HIV can maintain substant ial levels of protein expression and virion production.