Functional characterization of Drosophila melanogaster immune effectors

Drosophila melanogaster, commonly known as the "fruit fly", is a genetically tractable model organism widely used to study biological processes, notably the innate immune system. The advent of novel genome editing technologies, such as the CRISPR-Cas9 system, has allowed researchers to overcome technical challenges associated with conventional genetical approaches. This advancement has opened up novel opportunities to delete short genes and generate multiple knockouts, allowing the functional study of numerous uncharacterized genes. In this thesis, we take advantage of this biotechnological breakthrough to investigate three novel classes of immune-related genes. The first part of this thesis focuses on the functional characterization of a family of antimicrobial peptides named the Cecropins. We generated a fly line lacking the four Cecropin genes, named dCecA-C. Using the dCecA-C deficiency alone or in combination with other antimicrobial peptide mutations, we showed that Cecropins contribute to defense against certain Gram-negative bacteria and fungi. Our work provides the first genetic demonstration of a role for Cecropins in vivo. The second part of this thesis aims at characterizing the molecular function of a family of stress-induced peptides named the Turandots. We generated a mutant fly line lacking 6 Turandots (A,B,C,Z,M and X) and showed that this deletion increases fly susceptibility to environmental stresses due to tracheal apoptosis. The high exposure of phosphatidylserines, a negatively charged phospholipid, on the surface of tracheal cells sensitizes them to antimicrobial peptide activity. Turandots are secreted into the hemolymph of flies and subsequently bind to host cells exposing high levels of phosphatidylserines, masking them from cationic pore forming AMPs. This study provides the first demonstration of a role for Turandots in immune resilience by mitigating antimicrobial peptide toxicity to host tissues. The third part of this PhD thesis aims at functional characterization of the role of Drosophila immune-induced Dnases. To investigate this, we generated a null mutant line for Dnase II gene. Dnase II seems to play a role in disease tolerance, as Dnase II mutant flies are susceptible to systemic bacterial infection, without any increase in pathogen load. Our preliminary results suggest a role for Dnase II in the cellular immune response. Hemocytes of DNAse II deficient larvae are unable to digest phagocytosed apoptotic DNA after injury, leading to immune activation. This activation can be abolished by STING knock-out, suggesting that immunity is activated through STING following detection by a yet-unidentified cytosolic DNA sensor.Collectively, this thesis provides new insights on key innate immune effectors of Drosophila melanogaster, revealing their roles in fighting pathogens and increasing resilience, by protecting the host from deleterious effects of the immune system.