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 upregulate human immunodeficiency virus type-1 (HIV-1) replication in acutely infected cells and potentiate HIV expression from chronically infected promonocytic U1 cells stimulated with tumor necrosis factor-alpha (TNF-alpha). We have here investigated the potential effect of Dex in U1 cells stimulated with interleukin-6 (IL-6), a cytokine inducing virus expression by acting mostly at a post-transcriptional level on the virus life cycle.


METHODS
Virus production in culture supernatants was evaluated by reverse transcriptase (RT) activity. GC receptor expression was tested by both binding of [3H]-Dexamethasone 21-mesylate and Northern blotting. Cell-associated HIV protein expression was analyzed by Western blotting, whereas both HIV and monocyte chemoattractant protein-1 (MCP-1) RNA accumulation were evaluated by Northern blotting. HIV transcription was tested by long terminal repeat (LTR) chloramphenicol acetyl transferase (CAT) assay after transient transfection of U1 or U937 cells. Formation of activating protein-1 (AP-1) DNA binding complex in nuclear cell extracts was visualized by electrophoretic mobility shift assay (EMSA), whereas ERK1/2 mitogen-activated protein kinase (MAPK) phosphorylation was studied by Western blotting.


RESULTS
IL-6 and Dex synergistically induced HIV expression in U1 cells, and this effect was blocked by RU 486. No substantial HIV RNA accumulation was demonstrated in U1 cells co-stimulated with IL-6 and Dex, whereas IL-6 upregulated the expression of MCP-1 RNA, and this effect was inhibited by Dex. 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 potentiated by Dex. In sharp contrast, no induction of LTR-directed CAT activity was observed in transfected U1 cells (or in their parental uninfected U937 cells) stimulated with IL-6 and Dex either alone or in combination.


CONCLUSIONS
High levels of virion production can be induced in latently infected 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 pathways influenced by cytokine and GC through which HIV can maintain substantial levels of protein expression and virion production.
Kinter
A L
AL
Laboratory of Immunoregulation, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA.
Biswas
P
P
Alfano
M
M
Justement
J S
JS
Mantelli
B
B
Rizzi
C
C
Gatti
A R
AR
Vicenzi
E
E
Bressler
P
P
Poli
G
G
D001324:Autoantigens / Q000235:genetics / Q000378:metabolism; D015152:Blotting, Northern; D015153:Blotting, Western; D018932:Chemokine CCL2; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D003907:Dexamethasone / Q000031:analogs & derivatives* / Q000494:pharmacology*; D004357:Drug Synergism; D024202:Electrophoretic Mobility Shift Assay; D005938:Glucocorticoids / Q000494:pharmacology*; D016325:HIV Long Terminal Repeat / Q000187:drug effects / Q000502:physiology; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D015850:Interleukin-6 / Q000494:pharmacology*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000502:physiology; D009000:Monocytes / Q000187:drug effects* / Q000821:virology; D012367:RNA, Viral / Q000096:biosynthesis; D011965:Receptors, Glucocorticoid / Q000378:metabolism; D015398:Signal Transduction; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014775:Virus Activation / Q000187:drug effects; D014779:Virus Replication / Q000187:drug effects
D001324:Autoantigens; C111835:CCL2 protein, human; D018932:Chemokine CCL2; D005938:Glucocorticoids; D015850:Interleukin-6; D012367:RNA, Viral; D011965:Receptors, Glucocorticoid; D018808:Transcription Factor AP-1; D003907:Dexamethasone; D015500:Chloramphenicol O-Acetyltransferase; D020929:Mitogen-Activated Protein Kinase Kinases; C031710:dexamethasone 21-methanesulfonate