Inducible expression of IκBα represser mutants interferes with NF-κB activity and HIV-1 replication in Jurkat T cells

Abstract

Human immunodeficiency virus (HIV-1) utilizes the NF-κB/Rel proteins to regulate transcription through NF-κB binding sites in the HIV-1 long terminal repeat (LTR). Normally, NF-κB is retained in the cytoplasm by inhibitory IκB proteins; after stimulation by multiple activators including viruses, IκBα is phosphorylated and degraded, resulting in NF-κB release. In the present study, we examined the effect of tetracycline-inducible expression of transdominant repressors of IκBα (TDIκBα) on HIV-1 multiplication using stably selected Jurkat T cells. TD-IκBα was inducibly expressed as early as 3 h after doxycycline addition and dramatically reduced both NF-κB DNA binding activity and LTR-directed gene activity. Interestingly, induced TD-IκBα expression also decreased endogenous IκBα expression to undetectable levels by 24 h after induction, demonstrating that TD-IκBα repressed endogenous NF-κB-dependent gene transcription. TD- IκBα expression also sensitized Jurkat cells to tumor necrosis factor- induced apoptosis. De nero HIV-1 infection of Jurkat cells was dramatically altered by TD-IκBα induction, resulting in inhibition of HIV-1 multiplication, as measured by p24 antigen, reverse transcriptase, and viral RNA. Given the multiple functions of the NF-κB/IκB pathway, TD-IκBα expression may interfere with HIV-1 multiplication at several levels: LTR- mediated transcription, Rev-mediated export of viral RNA, inhibition of HIV- l-induced pro-inflammatory cytokines, and increased sensitivity of HIV1- infected cells to apoptosis.