Cecropins contribute toDrosophilahost defence against fungal and Gram-negative bacterial infection

Abstract

Cecropins are small helical secreted peptides with antimicrobial activity that are widely distributed among insects. Genes encoding cecropins are strongly induced upon infection, pointing to their role in host-defense. In Drosophila, four cecropin genes clustered in the genome (CecA1, CecA2, CecB and CecC) are expressed upon infection downstream of the Toll and Imd pathways. In this study, we generated a short deletion ΔCecA-C removing the whole cecropin locus. Using the ΔCecA-C deficiency alone or in combination with other antimicrobial peptide (AMP) mutations, we addressed the function of cecropins in the systemic immune response. ΔCecA-C flies were viable and resisted challenge with various microbes as wild-type. However, removing ΔCecA-C in flies already lacking ten other AMP genes revealed a role for cecropins in defense against Gram-negative bacteria and fungi. Measurements of pathogen loads confirm that cecropins contribute to the control of certain Gram-negative bacteria, notably Enterobacter cloacae and Providencia heimbachae. Collectively, our work provides the first genetic demonstration of a role for cecropins in insect host defense, and confirms their in vivo activity primarily against Gram-negative bacteria and fungi. Generation of a fly line (ΔAMP14) that lacks fourteen immune inducible AMPs provides a powerful tool to address the function of these immune effectors in host-pathogen interactions and beyond.

Drosophilaimmunity: theDrosocingene encodes two host defence peptides with pathogen-specific roles

“…Previous efforts were unable to link this 2307 Da peptide to a gene in the Drosophila reference genome. However, during our studies, we noticed that IM7 was absent in flies lacking 14 AMP genes, indicating that it is probably produced by one of these genes [6,23]. We repeated these MALDI-TOF proteomic experiments with haemolymph samples from flies carrying systematic combinations of AMP mutations, ultimately honing in on the gene Drosocin (Dro) .…”

“…We focused our screen on a panel of Gram-negative bacteria of interest: En. cloacae β12 bacteria that Dro gene mutants are specifically susceptible to [6,23], a recently-isolated Acetobacter sp . that can kill AMP mutant flies [39], E. coli 1106 suggested to be affected by the Dro gene [22,38] , and P. burhodogranariea strain B where the Dro gene was shown to contribute to defence alongside other AMPs [6] .…”

“…Unlike the example with Diptericins and P. rettgeri , other AMPs also contribute collectively to defence against En. cloacae [23].…”

Ingestion ofBacillus cereusspores dampens the immune response to favor bacterial persistence

“…To assess the involvement of the AMPs in Btk SA-11 persistence, we used a fly strain in which the 14 AMP genes were deleted (ΔAMP 14 ) (Carboni et al, 2021;Hanson et al, 2019). We quantified the Btk SA-11 load in the midgut of wild type (Canton S) and ΔAMP 14 flies.…”

“…Flies were reared on our standard laboratory medium (Nawrot-Esposito et al, 2020) Dredd F64 (Leulier et al, 2000) Dif ] (Bloomington #36559); ΔAMP14 (gift from B. Lemaitre) (Carboni et al, 2022); w; AttD-Gal4 UAS-cherry (gift from Leopold Kurz) (Tavignot et al, 2017); w; DptA-cherry (gift from Leopold Kurz, Bloomington #55706); UAS-DUOX RNAI (Bloomington #38907); UAS-REL RNAI (Bloomington #33661); UAS-PGRP-LB RNAI (Bloomington #67236); UAS-PGRP-SC2 RNAI (Bloomington #56915); w; myo1A-Gal4; tubGal80ts UAS-GFP/TM6b (gift from Nicolas Tapon) (Shaw et al, 2010). The w; Rel E20 , Dif 1 homozygous viable double mutant was obtained using classic mendelian genetic crosses.…”