The Drosophila Baramicin polypeptide gene protects against fungal infection.

The fruit fly Drosophila melanogaster combats microbial infection by producing a battery of effector peptides that are secreted into the haemolymph. Technical difficulties prevented the investigation of these short effector genes until the recent advent of the CRISPR/CAS era. As a consequence, many putative immune effectors remain to be formally described, and exactly how each of these effectors contribute to survival is not well characterized. Here we describe a novel Drosophila antifungal peptide gene that we name Baramicin A. We show that BaraA encodes a precursor protein cleaved into multiple peptides via furin cleavage sites. BaraA is strongly immune-induced in the fat body downstream of the Toll pathway, but also exhibits expression in other tissues. Importantly, we show that flies lacking BaraA are viable but susceptible to the entomopathogenic fungus Beauveria bassiana. Consistent with BaraA being directly antimicrobial, overexpression of BaraA promotes resistance to fungi and the IM10-like peptides produced by BaraA synergistically inhibit growth of fungi in vitro when combined with a membrane-disrupting antifungal. Surprisingly, BaraA mutant males but not females display an erect wing phenotype upon infection. Here, we characterize a new antifungal immune effector downstream of Toll signalling, and show it is a key contributor to the Drosophila antimicrobial response.

New insights on Drosophila antimicrobial peptide function in host defense and beyond.

Since the first animal antimicrobial peptides (AMPs) were discovered in insects, Drosophila melanogaster has emerged as a powerful model for their characterization. Drosophila AMPs have been used extensively to monitor the activity of the Toll and Imd NF-κB pathways, but little was known of their precise functions. In this review, we summarize recent findings on the function of Drosophila AMPs not only for antimicrobial defense, but also in the gut, tumor control, and neurology. The integration of these new studies allows a new framework to be drawn that explains how AMPs can contribute simultaneously to microbe killing whilst also regulating important host cellular functions. These functions require that AMPs target not only negatively charged microbes but also aberrant host cells.

More Than Black or White: Melanization and Toll Share Regulatory Serine Proteases in Drosophila.

The melanization response is an important defense mechanism in arthropods. This reaction is mediated by phenoloxidases (POs), which are activated by complex extracellular serine protease (SP) cascades. Here, we investigate the role of SPs in the melanization response using compound mutants in D. melanogaster and discover phenotypes previously concealed in single-mutant analyses. We find that two SPs, Hayan and Sp7, activate the melanization response in different manners: Hayan is required for blackening wound sites, whereas Sp7 regulates an alternate melanization reaction responsible for the clearance of Staphylococcus aureus. We present evidence that Sp7 is regulated by SPs activating the Toll NF-κB pathway, namely ModSP and Grass. Additionally, we reveal a role for the combined action of Hayan and Psh in propagating Toll signaling downstream of pattern recognition receptors activating either Toll signaling or the melanization response.

Synergy and remarkable specificity of antimicrobial peptides in vivo using a systematic knockout approach.

Antimicrobial peptides (AMPs) are host-encoded antibiotics that combat invading microorganisms. These short, cationic peptides have been implicated in many biological processes, primarily involving innate immunity. In vitro studies have shown AMPs kill bacteria and fungi at physiological concentrations, but little validation has been done in vivo. We utilized CRISPR gene editing to delete all known immune-inducible AMPs of Drosophila, namely: 4 Attacins, 4 Cecropins, 2 Diptericins, Drosocin, Drosomycin, Metchnikowin and Defensin. Using individual and multiple knockouts, including flies lacking all 14 AMP genes, we characterize the in vivo function of individual and groups of AMPs against diverse bacterial and fungal pathogens. We found that Drosophila AMPs act primarily against Gram-negative bacteria and fungi, contributing either additively or synergistically. We also describe remarkable specificity wherein certain AMPs contribute the bulk of microbicidal activity against specific pathogens, providing functional demonstrations of highly specific AMP-pathogen interactions in an in vivo setting.