Transcriptional activation of the IFN-β gene by virus infection requires the cooperative assembly of an enhanceosome. We report that the stochastic and monoallelic expression of the IFN-β gene depends on interchromosomal associations with three identified distinct genetic loci that could mediate binding of the limiting transcription factor NF-κB to the IFN-β enhancer, thus triggering enhanceosome assembly and activation of transcription from this allele. The probability of a cell to express IFN-β is dramatically increased when the cell is transfected with any of these loci. The secreted IFN-β protein induces high-level expression of the enhanceosome factor IRF-7, which in turn promotes enhanceosome assembly and IFN-β transcription from the remaining alleles and in other initially nonexpressing cells. Thus, the IFN-β enhancer functions in a nonlinear fashion by working as a signal amplifier.

The control of innate immune responses through activation of the nuclear transcription factor NF-{kappa}B is essential for the elimination of invading microbial pathogens. We showed that the bacterial N-(3-oxo-dodecanoyl) homoserine lactone (C12) selectively impairs the regulation of NF-{kappa}B functions in activated mammalian cells. The consequence is specific repression of stimulus-mediated induction of NF-{kappa}B–responsive genes encoding inflammatory cytokines and other immune regulators. These findings uncover a strategy by which C12-producing opportunistic pathogens, such as Pseudomonas aeruginosa, attenuate the innate immune system to establish and maintain local persistent infection in humans, for example, in cystic fibrosis patients.

Key to the pathogenicity of several viruses is activation of the canonical nuclear factor-κB (NF-κB) transcriptional pathway. Subversion of this tightly regulated mechanism is achieved through the production of host mimetic viral proteins that deregulate the transcription process. One such protein is ks-vFLIP (produced by the Kaposi's sarcoma herpes virus [KSHV]), which associates with IKKγ, an essential component of the IKK complex or signalosome. This interaction renders the canonical NF-κB pathway constitutively active and has been linked to Kaposi's sarcoma and other malignancies. In order to elucidate the molecular basis underpinning ks-vFLIP-induced activation of the IKK signalosome, we have determined the crystal structure of a complex involving a fragment of IKKγ bound to ks-vFLIP at 3.2 Å. In addition to identifying and subsequently probing the ks-vFLIP-IKKγ interface, we have also investigated the effects of a mutation implicated in the genetic disorder anhydrotic ectodermal dysplasia with immunodeficiency (EDA-ID).

Symbiotic mutualism with gut microbes occurs in all metazoans, and it is well established that commensal bacteria influence multiple aspects of host gut physiology such as innate immunity and development. However, our understanding of these coevolved interactions between prokaryotes and eukaryotes remains unclear. One mechanism by which commensal bacteria modulate host intracellular signaling pathways in order to avoid excess inflammation has now been determined. In this process, bacterial-induced reactive oxygen species in gut epithelial cells act as key messengers that inhibit the cullin-1–dependent protein degradation machinery, which in turn results in the stabilization of a master negative regulator of inflammation, inhibitor of nuclear factor-{kappa}B (I{kappa}B). Furthermore, this bacterial-mediated system also appears to be involved in the stabilization of a key developmental regulator, β-catenin. These findings provide new insights into the molecular mechanisms by which commensal microbes shape host cellular physiology.

Chlamydia trachomatis is an obligate intracellular bacterial pathogen that causes various human diseases, including blindness caused by ocular infection and sexually-transmitted diseases resulting from urogenital infection. After infecting host cells, Chlamydiae avoid alarming the host's immune system. Among the immune evasion mechanisms, Chlamydiae can inhibit NF-kappaB activation, a crucial pathway for host inflammatory responses. In this study, we show that ChlaDub1, a deubiquitinating and deneddylating protease from C. trachomatis, is expressed in infected cells. In transfection experiments, ChlaDub1 suppresses NF-kappaB activation induced by several pro-inflammatory stimuli and binds the NF-kappaB inhibitory subunit IkappaBalpha, impairing its ubiquitination and degradation. Thus, we provide further insight into the mechanism by which C. trachomatis may evade the host inflammatory response by demonstrating that ChlaDub1, a protease produced by this microorganism, is capable of inhibiting IkappaBalpha degradation and blocking NF-kappaB activation.

Toll-like receptors trigger an innate immune response by activating signaling pathways that are dependent on IRAK kinases. According to Kawagoe et al., the least understood IRAK member, IRAK2, is required for the perpetuation of these signals.

To define the roles of endothelial-intrinsic nuclear factor {kappa}B (NF-{kappa}B) activity in host defense and multiple organ injury in response to sepsis, we generated double transgenic (TG) mice (EC-rtTA/I-{kappa}B{alpha}mt) that conditionally overexpress a degradation-resistant form of the NF-{kappa}B inhibitor I-{kappa}B{alpha} (I-{kappa}B{alpha}mt) selectively on vascular endothelium. The EC-rtTA/I-{kappa}B{alpha}mt mice had no basal, but a relatively high level of doxycycline-inducible, I-{kappa}B{alpha}mt expression. I-{kappa}B{alpha}mt expression was detected in endothelial cells, but not in fibroblasts, macrophages, and whole blood cells, confirming that transgene expression was restricted to the endothelium. When subjected to endotoxemia, EC-rtTA/I-{kappa}B{alpha}mt mice showed endothelial-selective blockade of NF-{kappa}B activation, repressed expression of multiple endothelial adhesion molecules, reduced neutrophil infiltration into multiple organs, decreased endothelial permeability, ameliorated multiple organ injury, reduced systemic hypotension, and abrogated intravascular coagulation. When subjected to cecal ligation and puncture–induced sepsis, the TG mice had less severe multiple organ injury and improved survival compared with wild-type (WT) mice. WT and EC-rtTA/I-{kappa}B{alpha}mt mice had comparable capacity to clear three different pathogenic bacteria. Our data demonstrate that endothelial NF-{kappa}B activity is an essential mediator of septic multiple organ inflammation and injury but plays little role in the host defense response to eradicate invading pathogenic bacteria.

Proteins with death effector domains (DED) are key signal transducers involved in cell death and inflammation. In this issue of Cell, Sun et al. (2008) describe TIPE2, a DED protein that negatively regulates both T cell receptor and Toll-like receptor signaling. These findings reveal a new element critical to the maintenance of homeostasis in both the adaptive and innate immune systems.

Immune homeostasis is essential for the normal functioning of the immune system, and its breakdown leads to fatal inflammatory diseases. We report here the identification of a member of the tumor necrosis factor-alpha-induced protein-8 (TNFAIP8) family, designated TIPE2, that is required for maintaining immune homeostasis. TIPE2 is preferentially expressed in lymphoid tissues, and its deletion in mice leads to multiorgan inflammation, splenomegaly, and premature death. TIPE2-deficient animals are hypersensitive to septic shock, and TIPE2-deficient cells are hyper-responsive to Toll-like receptor (TLR) and T cell receptor (TCR) activation. Importantly, TIPE2 binds to caspase-8 and inhibits activating protein-1 and nuclear factor-kappaB activation while promoting Fas-induced apoptosis. Inhibiting caspase-8 significantly blocks the hyper-responsiveness of TIPE2-deficient cells. These results establish that TIPE2 is an essential negative regulator of TLR and TCR function, and its selective expression in the immune system prevents hyperresponsiveness and maintains immune homeostasis.

Members of the IRAK family of kinases mediate Toll-like receptor (TLR) signaling. Here we show that IRAK2 was essential for sustaining TLR-induced expression of genes encoding cytokines and activation of the transcription factor NF-kappaB, despite the fact that IRAK2 was dispensable for activation of the initial signaling cascades. IRAK2 was activated 'downstream' of IRAK4, like IRAK1, and TLR-induced cytokine production was abrogated in the absence of both IRAK1 and IRAK2. Whereas the kinase activity of IRAK1 decreased within 1 h of TLR2 stimulation, coincident with IRAK1 degradation, the kinase activity of IRAK2 was sustained and peaked at 8 h after stimulation. Thus, IRAK2 is critical in late-phase TLR responses, and IRAK1 and IRAK2 are essential for the initial responses to TLR stimulation.