IbinA and IbinB regulate the Toll pathway-mediated immune response in Drosophila melanogaster

Abstract

To combat infection, an immune system needs to be promptly activated but tightly controlled to avoid destructive effects on host tissues. IbinA and IbinB are related short peptides with robust expression upon microbial challenge in Drosophila melanogaster. Here, we show that Ibin genes are ubiquitously present in flies of the Drosophila subgenus Sophophora, where they replace a different but probably related gene, Mibin, which is found across a much wider range of cyclorrhaphan flies. Using synthetic peptides, we did not observe any direct bactericidal or bacteriostatic activity for either IbinA or IbinB in vitro. Using mutant Drosophila lines lacking the IbinA gene, IbinB gene, or both, we examined their roles in development and during microbial infections. IbinA is expressed in early pupae, and a lack of IbinA and IbinB leads to temperature-dependent formation of melanized tissue during metamorphosis, frequently around the trachea. IbinA and IbinB have distinct effects on susceptibility to microbial infection. For example, IbinB mutant flies, as well as flies lacking both IbinA and IbinB, had improved survival when challenged with Listeria monocytogenes, an intracellular pathogen, whereas a lack of IbinA alone had no effect. RNA sequencing of wildtype and mutant flies infected with L. monocytogenes showed enhanced Toll target gene expression in flies lacking IbinB, suggesting that IbinB acts as a negative regulator of the Toll pathway. In contrast, IbinA mutants had decreased Toll target gene expression in this context. Correspondingly, IbinB mutant flies had improved and IbinA compromised survival in septic fungal infection, where the Toll pathway has a major role. Our study provides insight into the roles of IbinA and IbinB in regulation of the immune response in Drosophila. Author summaryWhile the immune systems of animals must be able to be rapidly activated, they have the potential to cause severe tissue damage when overactive. Drosophila melanogaster has proven to be a highly effective model for studying core immune system pathways, and for developing concepts in host-pathogen interactions. The main signaling pathways involved in activation of the Drosophila immune system are well described and have contributed to the discovery of homologous pathways in the mammalian immune system. Despite this, the molecular mechanisms of action of the genes expressed downstream of these pathways are generally not well characterized. Regulation of the immune response also requires further investigation. Here, we use Drosophila mutants to show roles for two short peptides, both highly expressed during infection. IbinA and IbinB regulate the humoral immune response and the melanization reaction (an insect-specific immune reaction against parasites). Flies lacking one or both of these genes mount a stronger, more effective response to some pathogenic bacteria compared to wild type flies. During development, IbinA and IbinB have tissue protective roles, with mutants showing tissue damage due to aberrant immune activation. Our results indicate that IbinA and IbinB are important regulators of the immune response with tissue protective properties.