The ability to detect toxic compounds in foods is essential for animal survival. However, the minimal subunit composition of gustatory receptors required for sensing aversive chemicals in Drosophila is unknown. Here we report that three gustatory receptors, GR8a, GR66a and GR98b function together in the detection of L-canavanine, a plant-derived insecticide. Ectopic co-expression of Gr8a and Gr98b in Gr66a-expressing, bitter-sensing gustatory receptor neurons (GRNs) confers responsiveness to L-canavanine. Furthermore, misexpression of all three Grs enables salt- or sweet-sensing GRNs to respond to L-canavanine. Introduction of these Grs in sweet-sensing GRNs switches L-canavanine from an aversive to an attractive compound. Co-expression of GR8a, GR66a and GR98b in Drosophila S2 cells induces an L-canavanine-activated nonselective cation conductance. We conclude that three GRs collaborate to produce a functional L-canavanine receptor. Thus, our results clarify the full set of GRs underlying the detection of a toxic tastant that drives avoidance behaviour in an insect.

Animals discriminate nutritious food from toxic substances using their sense of taste. Since taste perception requires taste receptor cells to come into contact with water-soluble chemicals, it is a form of contact chemosensation. Concurrent with that contact, mechanosensitive cells detect the texture of food and also contribute to the regulation of feeding. Little is known, however, about the extent to which chemosensitive and mechanosensitive circuits interact. Here, we show Drosophila prefers soft food at the expense of sweetness and that this preference requires labellar mechanosensory neurons (MNs) and the mechanosensory channel Nanchung. Activation of these labellar MNs causes GABAergic inhibition of sweet-sensing gustatory receptor neurons, reducing the perceived intensity of a sweet stimulus. These findings expand our understanding of the ways different sensory modalities cooperate to shape animal behaviour.

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. The immune response is designed to target foreign infectious elements, not self, but it can become destructive when it fails to discriminate self from nonself. Therefore, it is important to restrain the magnitude and duration of the immune response by several mechanisms including receptor down-regulation and inhibitor synthesis. Here, focusing on the immune system of Drosophila, we present a mechanism of control that relies on the transcription factors AP-1 and STAT to prevent the excessive activation of the NF-κB–mediated immune response. Thus, AP-1 and STAT, renowned for their role in activating the NF-κB–mediated immune response, appear also to participate in its attenuation. In their role as negative regulators, AP-1 and STAT form a complex with HMG protein and HDAC. This complex is then recruited to the promoter regions of NF-κB target genes, causing the chromatin structure near the NF-κB target genes to contract and the expression of NF-κB target genes to shut down. Mis-regulation of this negative-feedback process, we found, increased the lethality of bacterial infection in Drosophila. A similar scenario has been noted in mammals with over-activated NF-κB–mediated immune responses, which has been implicated in autoimmune disease. Thus, feedback inhibition of NF-κB appears to be evolutionarily conserved to maintain properly balanced immune responses.

Insect survival depends on contact chemosensation to sense and avoid consuming plant-derived insecticides, such asl-canavanine. Members of a family of ∼60 gustatory receptors (GRs) comprise the main peripheral receptors responsible for taste sensation inDrosophila. However, the roles of mostDrosophilaGRs are unknown. In addition to GRs, a G protein-coupled receptor, DmXR, has been reported to be required for detectingl-canavanine. Here, we showed that GRs are essential for responding tol-canavanine and that flies missing DmXR displayed normall-canavanine avoidance andl-canavanine-evoked action potentials. Mutations disrupting eitherGr8aorGr66aresulted in an inability to detectl-canavanine. We found thatl-canavanine stimulated action potentials in S-type sensilla, which were whereGr8aandGr66awere both expressed, but not inGr66a-expressing sensilla that did not expressGr8a.l-canavanine-induced action potentials were also abolished in theGr8aandGr66amutant animals.Gr8awas narrowly required for responding tol-canavanine, in contrast toGr66a, which was broadly required for responding to other noxious tastants. Our data suggest that GR8a and GR66a are subunits of anl-canavanine receptor and that GR8a contributes to the specificity forl-canavanine.

Phagocytosis of invading microbes requires dynamic rearrangement of the plasma membrane and its associated cytoskeletal actin network. The polarization of Cdc42 and Rac1 Rho GTPases to the site of plasma membrane protrusion is responsible for the remodeling of actin structures. However, the mechanism of Rho GTPase recruitment to these sites and the identities of accessory molecules involved in this process are not well understood. In this study, we uncovered several new components involved in innate immunity in Drosophila melanogaster. Our data demonstrate that Rab35 is a regulator of vesicle transport required specifically for phagocytosis. Moreover, recruitment of Cdc42 and Rac1 to the sites of filopodium and lamellipodium formation is Rab35 dependent and occurs by way of microtubule tracks. These results implicate Rab35 as the immune cell-specific regulator of vesicle transport within the actin-remodeling complex.