Intracellular double-stranded RNA (dsRNA) is a chief sign of replication for many viruses. Host mechanisms detect the dsRNA and initiate antiviral responses. In this report, we identify retinoic acid inducible gene I (RIG-I), which encodes a DExD/H box RNA helicase that contains a caspase recruitment domain, as an essential regulator for dsRNA-induced signaling, as assessed by functional screening and assays. A helicase domain with intact ATPase activity was responsible for the dsRNA-mediated signaling. The caspase recruitment domain transmitted 'downstream' signals, resulting in the activation of transcription factors NF-kappaB and IRF-3. Subsequent gene activation by these factors induced antiviral functions, including type I interferon production. Thus, RIG-I is key in the detection and subsequent eradication of the replicating viral genomes.

The cellular protein retinoic acid-inducible gene I (RIG-I) senses intracellular viral infection and triggers a signal for innate antiviral responses including the production of type I IFN. RIG-I contains a domain that belongs to a DExD/H-box helicase family and exhibits an N-terminal caspase recruitment domain (CARD) homology. There are three genes encoding RIG-I-related proteins in human and mouse genomes. Melanoma differentiation associated gene 5 (MDA5), which consists of CARD and a helicase domain, functions as a positive regulator, similarly to RIG-I. Both proteins sense viral RNA with a helicase domain and transmit a signal downstream by CARD; thus, these proteins share overlapping functions. Another protein, LGP2, lacks the CARD homology and functions as a negative regulator by interfering with the recognition of viral RNA by RIG-I and MDA5. The nonstructural protein 3/4A protein of hepatitis C virus blocks the signaling by RIG-I and MDA5; however, the V protein of the Sendai virus selectively abrogates the MDA5 function. These results highlight ingenious mechanisms for initiating antiviral innate immune responses and the action of virus-encoded inhibitors.

Epstein-Barr virus (EBV)-encoded small RNAs (EBERs) are nonpolyadenylated, untranslated RNAs, exist most abundantly in latently EBV-infected cells, and are expected to show secondary structures with many short stemloops. Retinoic acid-inducible gene I (RIG-I) is a cytosolic protein that detects viral double-stranded RNA (dsRNA) inside the cell and initiates signaling pathways leading to the induction of protective cellular genes, including type I interferons (IFNs). We investigated whether EBERs were recognized by RIG-I as dsRNA. Transfection of RIG-I plasmid induced IFNs and IFN-stimulated genes (ISGs) in EBV-positive Burkitt's lymphoma (BL) cells, but not in their EBV-negative counterparts or EBER-knockout EBVinfected BL cells. Transfection of EBER plasmid or in vitro-synthesized EBERs induced expression of type I IFNs and ISGs in RIG-I-expressing, EBV-negative BL cells, but not in RIG-I-minus counterparts. EBERs activated RIG-I's substrates, NF-jB and IFN regulatory factor 3, which were necessary for type I IFN activation. It was also shown that EBERs co-precipitated with RIG-I. These results indicate that EBERs are recognized by RIG-I and activate signaling to induce type I IFN in EBV-infected cells.

RIG-I senses intracellular virus-specific nucleic acid structures and initiates an antiviral response that induces interferon (IFN) production, which, in turn, activates the transcription of RIG-I to increase RIG-I protein levels. Upon intracellular poly(I:C) stimulation, however, the levels of RIG-I protein did not correlate with the expression patterns of RIG-I transcripts. When the ISG15 conjugation system was overexpressed, ISG15 was conjugated to RIG-I and cellular levels of the unconjugated form of RIG-I decreased. The ISGylation of RIG-I reduced levels of both basal and virus-induced IFN promoter activity. Levels of unconjugated RIG-I also decreased when 26S proteasome activity was blocked by treatment with MG132, ALLN, or Lactacystin. In the presence of MG132, ISG15 conjugation to RIG-I increased, and hence, the unconjugated form of RIG-I was reduced. In Ube1L ؊/؊ cells, which lack the ability to conjugate ISG15, basal levels of both RIG-I protein and transcripts were increased compared to those in wild-type cells. As a result, enhanced production of ISGs and enhanced IFN promoter activity in Ube1L؊/؊ cells were observed, and the phenotype was restored to that of wild-type cells by the overexpression of Ube1L. Based on these results, we propose a novel negative feedback loop which adjusts the strength of the RIG-I-mediated antiviral response and IFN production through the regulation of RIG-I protein by IFN-induced ISG15 conjugation.

CX3CL1/Fractalkine, a CX3C chemokine, is a potent agonist for the chemotaxis and adhesion of monocytes and lymphocytes. It was first identified as a membrane protein in endothelial cells activated with IL-1 or TNF-alpha. We have found the enhanced expression of fractalkine in human umbilical vein endothelial cells stimulated with interferon-gamma (IFN-gamma). Pretreatment of the cells with cycloheximide did not inhibit the expression of fractalkine mRNA. The majority of fractalkine protein was found in the cell lysate, and an antibody-blocking experiment disclosed that fractalkine contributes to the adhesion of mononuclear cells to endothelial monolayers stimulated with IFN-gamma. Vascular endothelial cells produce fractalkine in response to IFN-gamma, and this may play an important role in immune responses by eliciting a traffic of mononuclear cells through the vascular wall.

We first cloned galectin-9 (Gal-9)/ecalectin as a T cell-derived eosinophil chemoattractant. Gal-9 plays a role in not only accumulation but also activation of eosinophils in experimental allergic models and human allergic patients, because Gal-9 induces eosinophil chemoattraction in vitro and in vivo and activates eosinophils in many aspects. Gal-9 requires divalent galactoside-binding activity but not the linker peptide of Gal-9 to exhibit its biological functions, and an unidentified matrix metalloproteinase is involved in the release of Gal-9. Our recent studies also showed that Gal-9 has other functions, such as cell differentiation, aggregation, adhesion, and death. Now, we and other groups are on the way of investigating the regulation and function of Gal-9 in a variety of physiological and pathological conditions. In this article, we will show the possible role of Gal-9 in physiological and pathological conditions by using our recent findings.

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