The fruit fly, Drosophila melanogaster, has helped us to understand how innate immunity is activated. In addition to the Toll receptor and the Toll signaling pathway, the Drosophila immune response is regulated by another evolutionarily conserved signaling cascade, the immune deficiency (Imd) pathway, which activates NF-κB. In fact, the Imd pathway controls the expression of most of the antimicrobial peptides in Drosophila; thus, it is indispensable for normal immunity in flies. In this article, we review the current literature on the Drosophila Imd pathway, with special emphasis on its role in the (patho)physiology of different organs. We discuss the systemic response, as well as local responses, in the epithelial and mucosal surfaces and the nervous system.

NF-κB transcription factors are involved in evolutionarily conserved signaling pathways controlling multiple cellular processes including apoptosis and immune and inflammatory responses. Immune response of the fruit fly Drosophila melanogaster to Gram-negative bacteria is primarily mediated via the Imd (immune deficiency) pathway, which closely resembles the mammalian TNFR signaling pathway. Instead of cytokines, the main outcome of Imd signaling is the production of antimicrobial peptides. The pathway activity is delicately regulated. Although many of the Imd pathway components are known, the mechanisms of negative regulation are more elusive. In this study we report that a previously uncharacterized gene, pirk, is highly induced upon Gram-negative bacterial infection in Drosophila in vitro and in vivo. pirk encodes a cytoplasmic protein that coimmunoprecipitates with Imd and the cytoplasmic tail of peptidoglycan recognition protein LC (PGRP-LC). RNA interference-mediated down-regulation of Pirk caused Imd pathway hyperactivation upon infection with Gram-negative bacteria, while overexpression of pirk reduced the Imd pathway response both in vitro and in vivo. Furthermore, pirk-overexpressing flies were more susceptible to Gram-negative bacterial infection than wild-type flies. We conclude that Pirk is a negative regulator of the Imd pathway.

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Because NF-κB signaling pathways are highly conserved in evolution, the fruit fly Drosophila melanogaster provides a good model to study these cascades. We carried out an RNA interference (RNAi)-based genome-wide in vitro reporter assay screen in Drosophila for components of NF-κB pathways. We analyzed 16,025 dsRNA-treatments and identified 10 novel NF-κB regulators. Of these, nine dsRNA-treatments affect primarily the Toll pathway. G protein-coupled receptor kinase (Gprk)2, CG15737/Toll pathway activation mediating protein, and u-shaped were required for normal Drosomycin response in vivo. Interaction studies revealed that Gprk2 interacts with the Drosophila IκB homolog Cactus, but is not required in Cactus degradation, indicating a novel mechanism for NF-κB regulation. Morpholino silencing of the zebrafish ortholog of Gprk2 in fish embryos caused impaired cytokine expression after Escherichia coli infection, indicating a conserved role in NF-κB signaling. Moreover, small interfering RNA silencing of the human ortholog GRK5 in HeLa cells impaired NF-κB reporter activity. Gprk2 RNAi flies are susceptible to infection with Enterococcus faecalis and Gprk2 RNAi rescues Toll^10b-induced blood cell activation in Drosophila larvae in vivo. We conclude that Gprk2/GRK5 has an evolutionarily conserved role in regulating NF-κB signaling.

JAK/STAT signaling pathway is evolutionarily conserved and tightly regulated. We carried out a reporter‐based genome‐wide RNAi in vitro screen to identify genes that regulate Drosophila JAK/STAT pathway and found 5 novel regulators. Of these, CG14225 is a negative regulator structurally related to the Drosophila JAK/STAT pathway receptor Domeless, especially in the extracellular domain, and to the mammalian IL‐6 receptor and the signal transducer gp130. CG14225 coimmunoprecipitates with Domeless and its associated kinase hopscotch in S2 cells. CG14225 RNAi caused hyperphosphorylation of the transcription factor Stat92E in S2 cells on stimulation with the Drosophila JAK/STAT pathway ligand unpaired. CG14225 RNAi in vivo hyperactivated JAK/STAT target genes on septic injury and enhanced unpaired‐induced eye overgrowth, and was thus named the eye transformer (ET) . In the gastrointestinal infection model, where JAK/STAT signaling is important for stem cell renewal, CG14225/ET RNAi was protective in vivo . In conclusion, we have identified ET as a novel negative regulator of the Drosophila JAK/STAT pathway both in vitro and in vivo , and it functions in regulating Stat92E phosphorylation.—Kallio, J., Myllymäki, H., Grönholm, J., Armstrong, M., Vanha‐aho, L.‐M., Mäkinen, L., Silvennoinen, O., Valanne, S., Rämet, M. Eye transformer is a negative regulator of Drosophila JAK/STAT signaling. FASEBJ . 24, 4467–4479 (2010). www.fasebj.org