Innate Immune Response In Drosophila Research Articles

A Feedback Regulatory Loop Involving dTrbd/dTak1 in Controlling IMD Signaling in Drosophila Melanogaster.

Negative regulators of the inflammatory responses are essential for the maintenance of immune homeostasis and organismal fitness. In Drosophila, the deubiquitinase (Dub) dTrbd selectively restricts the K63-linked ubiquitination modification of dTak1, a pivotal kinase of the IMD signaling pathway, to regulate the IMD innate immune response. However, which domain and how it functions to enable dTrbd’s activity remain unexplored. Here, we provide compelling evidence showing that the NZF domain of dTrbd is essential for its association with dTak1. Meanwhile, the Linker region of dTrbd is involved in modulating its condensation, a functional state representing the Dub enzymatical activity of dTrbd. Of interest, the activated IMD signals following bacterial stimuli enhance the dTrbd/dTak1 interaction, as well as the condensate assembly and Dub enzymatical activity of dTrbd. Collectively, our studies shed light on the dual mechanisms by which the IMD signaling-mediated feedback loop of dTrbd/dTak1 precisely regulates the innate immune response in Drosophila.

RNA-binding protein Roquin negatively regulates STING-dependent innate immune response in Drosophila.

Drosophila melanogaster utilizes innate immune response to defend against exogenous pathogens. The molecular regulation mechanism of the process is evolutionarily conserved. Research of the regulatory mechanisms of Drosophila innate immunity is greatly significant for understanding the modulation of the human innate immunity and the pathogenesis of related diseases. To explore novel regulators in the STING-dependent innate immune response in Drosophila, we utilized the double-stranded RNA-mediated gene expression silencing technique and the dual-luciferase reporter system in knockdown experiments on 9 genes encoding the ubiquitin ligase such as echinus (CG2904), usp16 (CG4165), smurf (CG4943), pellino (CG5212), usp47 (CG5486), diap2 (CG8293), dtraf2 (CG10961), roquin (CG16807) and usp10 (CG32479) in the S2 cells in vitro. The results suggested a negative correlation between CG16807 (roquin) and the STING signaling pathway. Further studies showed that over-expression of roquin in S2 cells significantly inhibited STING innate immune signaling. Meanwhile, Listeria infection experiments showed that knocking down of roquin markedly elevated the expression levels of anti-microbial peptides and inhibited the proliferation of Listeria, thus increasing the survival rates post pathogenic infection. Taken together, our results suggested that the RNA-binding protein Roquin negatively regulates the STING-dependent innate immune response in Drosophila. In view of the high correlation between Drosophila genes and human genes, this study provides a theoretical basis for further development of treatments for STING-related innate immune diseases in humans.

Reduced Dpp expression accelerates inflammation-mediated neurodegeneration through activated glial cells during altered innate immune response in Drosophila

The progression of neurodegenerative disease is very complex biological process and the molecular crosstalk of inflammatory cytokines during neurodegeneration is associated with multiple cascade signalling. Few evidences suggest that environmental toxin, Paraquat (PQ) administration activates the microglia and intensify the release of proinflamatory cytokines during progression of Parkinson’'s disease (PD) but the proper aetiology remained unknown. However, the fundamental role of anti-inflammatory molecule Decapentaplegic (Dpp), homologue of the secreted mammalian Transforming growth factor-β (TGF-β) signalling molecule during neurodegeneration of invertebrate fly model is yet to establish. To elucidate the molecular processes during early stage of Parkinson's disease, we observed neuro-toxin plays a determining role in the increased vulnerability to a particular PQ exposure that is attended by decreased lifespan, severe locomotor deficits, and more loss of dopaminergic (DA) neuron in PQ-treated Dpp deficient fly than wild type (WT). Simultaneously, activated microglia induced the inflammatory response with the release of pro-inflammatory and anti-inflammatory cytokine in Drosophila during neurodegeneration. Moreover, neuro-toxin exposure altered the expression of innate immune genes in both WT and mutant fly compared to the respective PQ-treated flies. Interestingly, PQ exposure reduced the expression of innate immune genes in mutant fly compared to WT. It may indicate that PQ exposure had broken down the immune defence response in mutant fly than WT whereas, without PQ exposure the innate immune tolerance level was higher in fly with reduced Dpp expression than WT. Thus, we observed the conserve anti-inflammatory factor TGF-β may exhibit a crucial defensive role during inflammation mediated neurodegeneration in invertebrate Drosophila melanogaster.

Beyond Host Defense: Deregulation of Drosophila Immunity and Age-Dependent Neurodegeneration.

Age-dependent neurodegenerative disorders are a set of diseases that affect millions of individuals worldwide. Apart from a small subset that are the result of well-defined inherited autosomal dominant gene mutations (e.g., those encoding the β-amyloid precursor protein and presenilins), our understanding of the genetic network that underscores their pathology, remains scarce. Genome-wide association studies (GWAS) especially in Alzheimer's disease patients and research in Parkinson's disease have implicated inflammation and the innate immune response as risk factors. However, even if GWAS etiology points toward innate immunity, untangling cause, and consequence is a challenging task. Specifically, it is not clear whether predisposition to de-regulated immunity causes an inadequate response to protein aggregation (such as amyloid or α-synuclein) or is the direct cause of this aggregation. Given the evolutionary conservation of the innate immune response in Drosophila and humans, unraveling whether hyperactive immune response in glia have a protective or pathological role in the brain could be a potential strategy in combating age-related neurological diseases.

Identification and Analysis of a Gene with Potential to Repress the IMD Pathway in Drosophila melanogaster.

The IMD pathway is one of the signal transduction pathways that regulates innate immune responses in Drosophila. To understand the regulation mechanism of the IMD pathway, we performed a mis-expression screening and identified a gene, Dyro, which potentially repress the expression of the downstream target genes of the IMD pathway. We confirmed that Dyro was expressed in the fat body where the IMD pathway is functional and that the overexpression of Dyro increases susceptibility. However, we detected neither an increased expression of target genes nor reduced susceptibility in mutants. During the analysis, we observed that the Dyro mutant exhibits a female sterile phenotype, and observed oogenesis defects. The results suggest that Dyro have potential to suppress immune response but its main role is regulation of oogenesis.

A high-sugar diet affects cellular and humoral immune responses in Drosophila

A high-sugar diet (HSD) induces Type 2 diabetes (T2D) and obesity, which severely threaten human health. The Drosophila T2D model has been constructed to study the mechanisms of insulin resistance, diet-induced cardiovascular diseases and other conditions. Innate immunity is the first line of defense against invading pathogens and parasites. However, few studies have focused on the relationship between a HSD and the innate immune response in Drosophila. In this study, we fed flies a high-sucrose diet and observed defects in the phagocytosis of latex beads and B. bassiana spores. The actin cytoskeleton was also disrupted in hemocytes from HSD-fed larvae. Furthermore, HSD induced the differentiation of lamellocytes in the lymph gland and circulating hemolymph, which rarely occurs in healthy bodies, via JNK signaling. In addition, the Toll and JNK pathways were excessively activated in the fat bodies of HSD-fed larvae, and a large number of dead cells were observed. Finally, HSD induced the aberrant activation of the innate immune system, including inflammation. Our results have established a connection between T2D and the innate immune response.

The distinct function of Tep2 and Tep6 in the immune defense of Drosophila melanogaster against the pathogen Photorhabdus

ABSTRACT Previous and recent investigations on the innate immune response of Drosophila have identified certain mechanisms that promote pathogen elimination. However, the function of Thioester-containing proteins (TEPs) in the fly still remains elusive. Recently we have shown the contribution of TEP4 in the antibacterial immune defense of Drosophila against non-pathogenic E. coli, and the pathogens Photorhabdus luminescens and P. asymbiotica. Here we studied the function of Tep genes in both humoral and cellular immunity upon E. coli and Photorhabdus infection. We found that while Tep2 is induced after Photorhabdus and E. coli infection; Tep6 is induced by P. asymbiotica only. Moreover, functional ablation of hemocytes results in significantly low transcript levels of Tep2 and Tep6 in response to Photorhabdus. We show that Tep2 and Tep6 loss-of-function mutants have prolonged survival against P. asymbiotica, Tep6 mutants survive better the infection of P. luminescens, and both tep mutants are resistant to E. coli and Photorhabdus. We also find a distinct pattern of immune signaling pathway induction in E. coli or Photorhabdus infected Tep2 and Tep6 mutants. We further show that Tep2 and Tep6 participate in the activation of hemocytes in Drosophila responding to Photorhabdus. Finally, inactivation of Tep2 or Tep6 affects phagocytosis and melanization in flies infected with Photorhabdus. Our results indicate that distinct Tep genes might be involved in different yet crucial functions in the Drosophila antibacterial immune response.

Bap180/Baf180 is required to maintain homeostasis of intestinal innate immune response in Drosophila and mice.

Immune homeostasis is a prerequisite to protective immunity against gastrointestinal infections. In Drosophila, immune deficiency (IMD) signalling (tumour necrosis factor receptor/interleukin-1 receptor, TNFR/IL-1R in mammals) is indispensable for intestinal immunity against invading bacteria. However, how this local antimicrobial immune response contributes to inflammatory regulation remains poorly defined. Here, we show that flies lacking intestinal Bap180 (a subunit of the chromatin-remodelling switch/sucrose non-fermentable (SWI/SNF) complex) are susceptible to infection as a result of hyper-inflammation rather than bacterial overload. Detailed analysis shows that Bap180 is induced by the IMD-Relish response to both enteropathogenic and commensal bacteria. Upregulated Bap180 can feed back to restrain overreactive IMD signalling, as well as to repress the expression of the pro-inflammatory gene eiger (TNF), a critical step to prevent excessive tissue damage and elongate the lifespan of flies, under pathological and physiological conditions, respectively. Furthermore, intestinal targeting of Baf180 renders mice susceptible to a more aggressive infectious colitis caused by Citrobacter rodentium. Together, Bap180 and Baf180 serve as a conserved transcriptional repressor that is critical for the maintenance of innate immune homeostasis in the intestines.

Genome-wide miRNA screening reveals miR-310 family members negatively regulate the immune response in Drosophila melanogaster via co-targeting Drosomycin

Although innate immunity mediated by Toll signaling has been extensively studied in Drosophila melanogaster, the role of miRNAs in regulating the Toll-mediated immune response remains largely unknown. In this study, following Gram-positive bacterial challenge, we identified 93 differentially expressed miRNAs via genome-wide miRNA screening. These miRNAs were regarded as immune response related (IRR). Eight miRNAs were confirmed to be involved in the Toll-mediated immune response upon Gram-positive bacterial infection through genetic screening of 41 UAS-miRNA lines covering 60 miRNAs of the 93 IRR miRNAs. Interestingly, four out of these eight miRNAs, miR-310, miR-311, miR-312 and miR-313, are clustered miRNAs and belong to the miR-310 family. These miR-310 family members were shown to target and regulate the expression of Drosomycin, an antimicrobial peptide produced by Toll signaling. Taken together, our study implies important regulatory roles of miRNAs in the Toll-mediated innate immune response of Drosophila upon Gram-positive bacterial infection.

Characterization of the effect of Cr(VI) on humoral innate immunity using Drosophila melanogaster.

With the advancement of human race, different anthropogenic activities have heaped the environment with chemicals that can cause alteration in the immune system of exposed organism. As a first line of barrier, the evolutionary conserved innate immunity is crucial for the health of an organism. However, there is paucity of information regarding in vivo assessment of the effect of environmental chemicals on innate immunity. Therefore, we examined the effect of a widely used environmental chemical, Cr(VI), on humoral innate immune response using Drosophila melanogaster. The adverse effect of Cr(VI) on host humoral response was characterized by decreased gene expression of antimicrobial peptides (AMPs) in the exposed organism. Concurrently, a significantly decreased transcription of humoral pathway receptors (Toll and PGRP) and triglyceride level along with inhibition of antioxidant enzyme activities were observed in exposed organism. This in turn weakened the immune response of exposed organism that was manifested by their reduced resistance against bacterial infection. In addition, overexpression of the components of humoral immunity particularly Diptericin benefits Drosophila from Cr(VI)-induced humoral immune-suppressive effect. To our knowledge, this is the first report regarding negative impact of an environmental chemical on humoral innate immune response of Drosophila along with subsequent protection by AMPs, which may provide novel insight into host-chemical interactions. Also, our data validate the utility and sensitivity of Drosophila as a model that could be used for screening the possible risk of environmental chemicals on innate immunity with minimum ethical concern that can be further extrapolated to higher organisms.

Fat facets induces polyubiquitination of Imd and inhibits the innate immune response in Drosophila

The IMD pathway is one of the major regulators of the innate immune response in Drosophila. Although extensive analysis of the IMD pathway has been carried out, precise mechanisms for how each target gene of the pathway is down-regulated remain to be clarified. Here, we carried out genetic screening and found that fat facets (faf), which encodes a deubiquitinating enzyme, inhibited the expression of the target genes of the IMD pathway. Overexpression of faf suppressed the infection-induced expression of Diptericin and increased susceptibility to bacterial infection in flies, whereas faf loss-of-function mutants decreased susceptibility. Time course analysis revealed that specific subsets of the target genes of the IMD pathway were affected by faf. Biochemical analysis showed that Faf made a complex with Imd, and both Faf and Imd were polyubiquitinated when they were co-overexpressed. Given that faf-dependent Imd polyubiquitination did not seem to cause protein degradation of Imd, Faf might inhibit the IMD pathway by modulating the state of Imd ubiquitination and/or stability.

Efficient RNA virus control in Drosophila requires the RNA methyltransferase Dnmt2

Drosophila use small-interfering RNA mechanisms to limit the amplification of viral genomes. However, it is unclear how small RNA interference components recognize and separate viral from cellular RNA. Dnmt2 enzymes are highly conserved RNA methyltransferases with substrate specificity towards cellular tRNAs. We report here that Dnmt2 is required for efficient innate immune responses in Drosophila. Dnmt2 mutant flies accumulate increasing levels of Drosophila C virus and show activated innate immune responses. Binding of Dnmt2 to DCV RNA suggests that Dnmt2 contributes to virus control directly, possibly by RNA methylation. These observations demonstrate a role for Dnmt2 in antiviral defence.

Quantitative Measurement of the Immune Response and Sleep in <em>Drosophila</em>

A complex interaction between the immune response and host behavior has been described in a wide range of species. Excess sleep, in particular, is known to occur as a response to infection in mammals (1) and has also recently been described in Drosophila melanogaster(2). It is generally accepted that sleep is beneficial to the host during an infection and that it is important for the maintenance of a robust immune system(3,4). However, experimental evidence that supports this hypothesis is limited(4), and the function of excess sleep during an immune response remains unclear. We have used a multidisciplinary approach to address this complex problem, and have conducted studies in the simple genetic model system, the fruitfly Drosophila melanogaster. We use a standard assay for measuring locomotor behavior and sleep in flies, and demonstrate how this assay is used to measure behavior in flies infected with a pathogenic strain of bacteria. This assay is also useful for monitoring the duration of survival in individual flies during an infection. Additional measures of immune function include the ability of flies to clear an infection and the activation of NFκB, a key transcription factor that is central to the innate immune response in Drosophila. Both survival outcome and bacterial clearance during infection together are indicators of resistance and tolerance to infection. Resistance refers to the ability of flies to clear an infection, while tolerance is defined as the ability of the host to limit damage from an infection and thereby survive despite high levels of pathogen within the system(5). Real-time monitoring of NFκB activity during infection provides insight into a molecular mechanism of survival during infection. The use of Drosophila in these straightforward assays facilitates the genetic and molecular analyses of sleep and the immune response and how these two complex systems are reciprocally influenced.

Phosphoinositide Binding by the Toll Adaptor dMyD88 Controls Antibacterial Responses in Drosophila

The cell biological principles that govern innate immune responses in Drosophila are unknown. Here, we report that Toll signaling in flies was dictated by the subcellular localization of the adaptor protein dMyD88. dMyD88 was located at the plasma membrane by a process dependent on a C-terminal phosphoinositide-binding domain. In vivo analysis revealed that lipid binding by dMyD88 was necessary for its antimicrobial and developmental functions as well as for the recruitment of the downstream cytosolic adaptor Tube to the cell surface. These data are reminiscent of the interactions between the mammalian Toll adaptors MyD88 and TIRAP with one major exception. In the mammalian system, MyD88 is the cytosolic adaptor that depends on the phosphoinositide-binding protein TIRAP for its recruitment to the cell surface. We therefore propose that dMyD88 is the functional homolog of TIRAP and that both proteins function as sorting adaptors to recruit downstream signaling adaptors to activated receptors.

Prizes in an Age of Collaborative Research

BioTechniquesVol. 52, No. 1 From the EditorOpen AccessPrizes in an age of collaborative researchNathan S. BlowNathan S. BlowBioTechniquesSearch for more papers by this authorPublished Online:3 Apr 2018https://doi.org/10.2144/000113792AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinkedInRedditEmail When the 2011 Nobel Prize in Physiology or Medicine was announced in October for immunologists Ralph Steinman, Bruce Beutler, and Jules Hoffmann, I was delighted. Having worked on the innate immune response in Drosophila a little during my postdoctoral fellowship, I was pleased to see the discovery of Toll and Toll-like receptors as regulators of immunity being awarded arguably science's highest honor. However, as I reflect on this award, and the controversy surrounding the recipients three months later, I can now see how this award has again shed light on what is becoming a growing issue when bestowing science prizes in general—celebrating individual accomplishment in an era of collaborative research.The controversy surrounding this year's physiology or medicine prize started shortly after the announcement when a group of 26 immunologists from around the globe co-signed a letter that appeared in the pages of journal Nature critiquing the Nobel committee for their omission in acknowledging the work of Charles Janeway Jr. and Ruslan Medzhitov, two pioneers in the field of immunology and innate immunity. This letter was then followed in early December by greater criticism from Bruno Lemaitre—a researcher who worked in Hoffmann's group—for the conferring of the award to Hoffmann. Lemaitre, who initiated the Toll project and is the first author of the article cited by the Nobel committee in their award announcement for Hoffmann, has gone so far as to set up a website devoted to explaining his role in the history of Toll discovery and the minor role he says Hoffmann played as a supervisor.The question of who uncovered the role of Toll in the immune response and which deserving scientists the committee missed will continue to be debated, I suspect. But the larger issue for the Nobel committee that is now coming to light is what to do in cases where collaborative contributions lead to seminal insights in biology? Today, a growing number of researchers are working together, and with this trend, issues of credit will most certainly continue to arise, especially if only three or less scientists can be honored for a discovery. What has taken place with the 2011 prize in physiology or medicine might just be the tip of the iceberg; other worthy efforts that involve multiple researchers working on a problem using a variety of techniques and skills could prove even more problematic to award, and potentially stir even more controversy within the scientific community.For example, the Human Genome Project is one of the crowning accomplishments in modern biology. And yet, there has been no Nobel Prize awarded for this technical feat that has fueled both novel biological discovery as well as our current march toward the field of personalized medicine. While some will no doubt argue that deciphering the human genome is not deserving of such an award, I have to wonder if another reason is that awarding a Nobel Prize in this case would truly highlight the challenges faced when trying to honor large-scale collaborative research efforts. Conjecture aside, one point has become clear during this scientific awards season: as science changes and grows so too should the manner in which we acknowledge scientific contributions. If we stay on the current track, more controversy will certainly arise in the coming years and diminish the impact of such prestigious honors.As science changes and grows, so too should the manner in which we acknowledge scientific contributionsPlease share your thoughts with us by posting at our Molecular Biology Forums under “To the Editor” (http://molecularbiology.forums.biotechniques.com) or sending an email directly to the editors (bioeditor@biotechniques.com).FiguresReferencesRelatedDetailsCited ByIs the Nobel Prize good for science?5 September 2013 | The FASEB Journal, Vol. 27, No. 12 Vol. 52, No. 1 Follow us on social media for the latest updates Metrics History Published online 3 April 2018 Published in print January 2012 Information© 2012 Author(s)PDF download

Diversity of Innate Immune Recognition Mechanism for Bacterial Polymeric meso-Diaminopimelic Acid-type Peptidoglycan in Insects

In Drosophila, the synthesis of antimicrobial peptides in response to microbial infections is under the control of the Toll and immune deficiency (Imd) signaling pathway. The Toll signaling pathway responds mainly to the lysine-type peptidoglycan of Gram-positive bacteria and fungal β-1,3-glucan, whereas the Imd pathway responds to the meso-diaminopimelic acid (DAP)-type peptidoglycan of Gram-negative bacteria and certain Gram-positive bacilli. Recently we determined the activation mechanism of a Toll signaling pathway biochemically using a large beetle, Tenebrio molitor. However, DAP-type peptidoglycan recognition mechanism and its signaling pathway are still unclear in the fly and beetle. Here, we show that polymeric DAP-type peptidoglycan, but not its monomeric form, formed a complex with Tenebrio peptidoglycan recognition protein-SA, and this complex activated the three-step proteolytic cascade to produce processed Spätzle, a Toll receptor ligand, and induced Drosophila defensin-like antimicrobial peptide in Tenebrio larvae similarly to polymeric lysine-type peptidoglycan. Monomeric DAP-type peptidoglycan induced Drosophila diptericin-like antimicrobial peptide in Tenebrio hemocytes. In addition, both polymeric and monomeric DAP-type peptidoglycans induced expression of Tenebrio peptidoglycan recognition protein-SC2, which is DAP-type peptidoglycan-selective N-acetylmuramyl-l-alanine amidase that functions as a DAP-type peptidoglycan scavenger, appearing to function as a negative regulator of the DAP-type peptidoglycan signaling by cleaving DAP-type peptidoglycan in Tenebrio larvae. Taken together, these results demonstrate that molecular recognition mechanism for polymeric DAP-type peptidoglycan is different between Tenebrio larvae and Drosophila adults, providing biochemical evidences of biological diversity of innate immune responses in insects.

A Large-Scale RNAi Screen Identifies Deaf1 as a Regulator of Innate Immune Responses in Drosophila

Innate immune signalling pathways are evolutionarily conserved between invertebrates and vertebrates. The analysis of NF-ĸB signalling in Drosophila has contributed important insights into how organisms respond to infection. Nevertheless, significant gaps remain in our understanding of how the activation of intracellular signalling elicits specific transcriptional programs. Here we report a genome-wide RNA interference survey for transcription factors that are required for Toll-dependent immune responses. In addition to the NF-ĸB homologs Dif, Dorsal and factors of the general transcription machinery, we identified Deformed Epidermal Autoregulatory Factor 1 (Deaf1) to be required for the expression of the Toll target gene Drosomycin in cultured cells and in Drosophila in vivo. We show that Deaf1 is required for the survival of flies after fungal, but not E. coli, infection. We determine that Deaf1 acts downstream of the NF-ĸB factors Dorsal and Dif. These results indicate that Deaf1 is an important contributor to innate immune responses in vivo.

Insect immune resistance to parasitoids

AbstractInsect host—parasitoid interactions involve complex physiological, biochemical and genetic interactions. Against endoparasitoids, immune‐competent hosts initiate a blood cell‐mediated response that quickly destroys the intruders and envelops them in a multilayered melanotic capsule. During the past decade, considerable progress has been made in identifying some of the critical components of the host response, mainly because of the use of efficient molecular tools. This review examines some of the components of the innate immune response of Drosophila, an insect that has served as an exceptionally good experimental model for studying non‐self recognition processes and immune cell signaling mechanisms. Topics considered in this review include hematopoiesis, proliferation and adhesion of hemocytes, melanogenesis and associated cytotoxic molecules, and the genetic aspects of the host‐parasitoid interaction.

Down-Regulation of NF-κB Target Genes by the AP-1 and STAT Complex during the Innate Immune Response in Drosophila

The activation of several transcription factors is required for the elimination of infectious pathogens via the innate immune response. The transcription factors NF-κB, AP-1, and STAT play major roles in the synthesis of immune effector molecules during innate immune responses. However, the fact that these immune responses can have cytotoxic effects requires their tight regulation to achieve restricted and transient activation, and mis-regulation of the damping process has pathological consequences. Here we show that AP-1 and STAT are themselves the major inhibitors responsible for damping NF-κB–mediated transcriptional activation during the innate immune response in Drosophila. As the levels of dAP-1 and Stat92E increase due to continuous immune signaling, they play a repressive role by forming a repressosome complex with the Drosophila HMG protein, Dsp1. The dAP-1–, Stat92E-, and Dsp1-containing complexes replace Relish at the promoters of diverse immune effector genes by binding to evolutionarily conserved cis-elements, and they recruit histone deacetylase to inhibit transcription. Reduction by mutation of dAP-1, Stat92E, or Dsp1 results in hyperactivation of Relish target genes and reduces the viability of bacterially infected flies despite more efficient pathogen clearance. These defects are rescued by reducing the Relish copy number, thus confirming that mis-regulation of Relish, not inadequate activation of dAP-1, Stat92E, or Dsp1 target genes, is responsible for the reduced survival of the mutants. We conclude that an inhibitory effect of AP-1 and STAT on NF-κB is required for properly balanced immune responses and appears to be evolutionarily conserved.

Structure of Tracheal Cytotoxin in Complex with a Heterodimeric Pattern-Recognition Receptor

Tracheal cytotoxin (TCT), a naturally occurring fragment of Gram-negative peptidoglycan, is a potent elicitor of innate immune responses in Drosophila. It induces the heterodimerization of its recognition receptors, the peptidoglycan recognition proteins (PGRPs) LCa and LCx, which activates the immune deficiency pathway. The crystal structure at 2.1 angstrom resolution of TCT in complex with the ectodomains of PGRP-LCa and PGRP-LCx shows that TCT is bound to and presented by the LCx ectodomain for recognition by the LCa ectodomain; the latter lacks a canonical peptidoglycan-docking groove conserved in other PGRPs. The interface, revealed in atomic detail, between TCT and the receptor complex highlights the importance of the anhydro-containing disaccharide in bridging the two ectodomains together and the critical role of diaminopimelic acid as the specificity determinant for PGRP interaction.