Viral induction of the human beta interferon promoter: modulation of transcription by NF-kappa B/rel proteins and interferon regulatory factors

Abstract

Multiple regulatory domains within the-100 region of the beta interferon (IFN-P) promoter control the inducible response of the IFN gene to virus infection. In this study, we demonstrate that the formation of NF-KB-specific complexes on the positive regulatory domain II (PRDII) precedes the onset of detectable IFN-I transcription in Sendai virus-infected cells. By using NF-KB subunit-specific antibodies, a temporal shift in the composition of NF-KB subunits in association with the PRDII domain is detected as a function of time after virus infection. Furthermore, a virus-induced degradation of IKBBa (MAD3) protein is observed between 2 and 8 h after infection; at later times, de novo synthesis of IKBa restores IKBC(to levels found in uninduced cells and correlates with the down regulation of IFN-0 transcription. In cotransfection experiments using various NF-KB subunit expression plasmids and two copies of PRDIIINF-KB linked to a chloramphenicol acetyltransferase reporter gene, we demonstrate that expression of p65, c-Rel, or p50 or combinations of p5O-p65 and p65-c-Rel differentially stimulated PRDII-dependent transcription. Coexpression of IK.Ba completely abrogated p65-, c-Rel-, or p65-p50-induced gene activity. When the entire IFN-4 promoter (-281 to +19) was used in coexpression studies, synergistic stimulation of IFN-f promoter activity was obtained when NF-KB subunits were coexpressed together with the IFN regulatory factor 1 (IRF-1) transcription factor. Overexpression of either IKB or the IRF-2 repressor was able to abrogate inducibility of the IFN-jI promoter. Thus, multiple regulatory events-including differential activation of DNA-binding NF-cB heterodimers, degradation of IKBa, synergistic interaction between IRF-1 and NF-KB, and decreased repression by IKB and IRF-2-are all required for the transcriptional activation of the IFN-, promoter.

Transcriptional activation of the interleukin-8 gene by respiratory syncytial virus infection in alveolar epithelial cells: nuclear translocation of the RelA transcription factor as a mechanism producing airway mucosal inflammation

“…Although our studies are the first to describe RelA activation by a mucosa-restricted virus, NF-B activation has been reported in myeloid cells as a consequence of chronic lentivirus infection (45,47), acute Sendai virus infection of fibroblastic and renal carcinoma cells (22), and picornavirus infection of alveolar epithelial cells (60). Lentivirus infection of myeloid cells activates NF-B chronically; given the ability of lentivirus to stimulate cytokine production, many of which can themselves stimulate NF-B, it is difficult to determine whether RelA activation is a direct effect of viral replication or a secondary paracrine phenomenon.…”

RelA, p50 and Inhibitor of kappa B alpha are Elevated in Human Metastatic Melanoma Cells and Respond Aberrantly to Ultraviolet Light B

“…The cDNA probes were labeled with [a-32 P]dCTP (NEN) using random nonamer priming (Pharmacia Biotech). The p50 cDNA probe was a 1.5-kb EcoRI insert, p65 was a 0.95-kb EcoRI insert, IkBa was a 1.2-kb EcoRI insert (gifts of John Hiscott, McGill University) (28), and an 18S rRNA probe was used (Ambion, 18S Decatemplate). The filters were hybridized with the [a-32 P]dCTP-labeled cDNA probes at 42°C for 2 hr in Rapid-Hyb Buffer (Amersham) and washed following the manufacturer's recommendation.…”

IκB-Mediated Inhibition of Virus-Induced Beta Interferon Transcription

“…Transfections and CAT reporter gene assays. Subconfluent 293 cells, CMVt-rtTA 293 cells, or CMVt-rtTA-IB-expressing 293 cells were transfected with the IFN-␤-CAT reporter plasmid, by the calcium phosphate coprecipitation method (17). All of the transfections contained equivalent amounts of DNA standardized with the CMV-B1 vector.…”