Pierre Génin

Transcriptional regulators of the NF-kB/IkB family promote the expression of well over 100 target genes, the majority of which participate in the host immune response (1). These proteins include a multitude of cytokines and chemokines, receptors required for immune recognition, proteins involved in antigen presentation, and adhesion receptors involved in transmigration across blood vessels walls. Because of this extensive role in immune action, NF-kB has been termed the central mediator of the immune response. Gene knockout and other studies establish roles for NF-kB in the ontogeny of the immune system but also demonstrate that NF-kB participates at multiple steps during oncogenesis (2) and the regulation of programmed cell death (3). For several reasons, the NF-kB pathway provides an attractive target to viral pathogens. Activation of NF-kB is a rapid, immediate early (IE) event that occurs within minutes after exposure to a relevant inducer, does not require de novo protein synthesis, and results in a strong transcriptional stimulation of several early viral as well as cellular genes. In this review, we will describe strategies that viruses have evolved to modulate the NF-kB pathway, to enhance viral replication, host cell survival, and evasion of immune responses. Activation of NF-kB constitutes an obvious target because many of its target genes — growth factors, cytokines and their receptors, and proto-oncogenes — profoundly influence the host cell cycle. In addition, some viruses exploit the antiapoptotic properties of NF-kB to evade the host defense mechanisms that limit replication by killing infected cells, or conversely to trigger apoptosis as a mechanism to increase virus spread. Perhaps not surprisingly, the persistent activation of the NF-kB pathway maintained by certain viruses contributes to oncogenic transformation (2). In addition to the classic studies with the avian REV-T retrovirus which contains the v-Rel oncoprotein and induces a rapid and fatal B-cell lymphoma in young birds (4), several lines of evidence demonstrate that NF-kB family members contribute to human oncogenesis. Localization of NF-kB–encoding genes at sites of chromosomal translocations and genomic rearrangements in cancer, high levels of NF-kB activity in many breast cancer cells, and constitutive nuclear NF-kB complexes in Hodgkin’s lymphoma cells all support this view (2). Furthermore, as discussed below, viral oncogene products, including human T-cell leukemia virus type 1 (HTLV-1) Tax protein and Epstein-Barr virus latent infection membrane protein 1 (EBV LMP1), each act by unique mechanisms to disrupt NF-kB regulation and initiate viral transformation.

Recent studies implicate the interferon (IFN) regulatory factors (IRF) IRF-3 and IRF-7 as key activators of the alpha/beta IFN (IFN-alpha/beta) genes as well as the RANTES chemokine gene. Using coexpression analysis, the human IFNB, IFNA1, and RANTES promoters were stimulated by IRF-3 coexpression, whereas the IFNA4, IFNA7, and IFNA14 promoters were preferentially induced by IRF-7 only. Chimeric proteins containing combinations of different IRF-7 and IRF-3 domains were also tested, and the results provided evidence of distinct DNA binding properties of IRF-3 and IRF-7, as well as a preferential association of IRF-3 with the CREB binding protein (CBP) coactivator. Interestingly, some of these fusion proteins led to supraphysiological levels of IFN promoter activation. DNA binding site selection studies demonstrated that IRF-3 and IRF-7 bound to the 5'-GAAANNGAAANN-3' consensus motif found in many virus-inducible genes; however, a single nucleotide substitution in either of the GAAA half-site motifs eliminated IRF-3 binding and transactivation activity but did not affect IRF-7 interaction or transactivation activity. These studies demonstrate that IRF-3 possesses a restricted DNA binding site specificity and interacts with CBP, whereas IRF-7 has a broader DNA binding specificity that contributes to its capacity to stimulate delayed-type IFN gene expression. These results provide an explanation for the differential regulation of IFN-alpha/beta gene expression by IRF-3 and IRF-7 and suggest that these factors have complementary rather than redundant roles in the activation of the IFN-alpha/beta genes.

Localized and systemic cytokine production in virus-infected cells play an important role in the outcome of viral infection and pathogenicity. Activation of the interferon regulatory factors (IRF) in turn is a critical mediator of cytokine gene transcription. Recent studies have focused on the 55-kDa IRF-3 gene product as a direct transcriptional regulator of type 1 interferon (IFN-alpha and IFN-beta) activation in response to virus infection. Virus infection induces phosphorylation of IRF-3 on specific C-terminal serine residues and permits cytoplasmic-to-nuclear translocation of IRF-3, activation of DNA binding and transactivation potential, and association with the CBP/p300 coactivator. We previously generated constitutively active [IRF-3(5D)] and dominant-negative forms of IRF-3 that control IFN-beta and IFN-alpha gene expression. In an effort to characterize the range of immunoregulatory genes controlled by IRF-3, we now demonstrate that endogenous human RANTES gene transcription is directly induced in tetracycline-inducible IRF-3(5D)-expressing cells or paramyxovirus-infected cells. We also show that a dominant-negative IRF-3 mutant inhibits virus-induced expression of the RANTES promoter. Specific mutagenesis of overlapping ISRE-like sites located between nucleotides -123 and -96 in the RANTES promoter reduces virus-induced and IRF-3-dependent activation. These studies broaden the range of IRF-3 immunoregulatory target genes to include at least one member of the chemokine superfamily.