Identification of novel defense factors using Luciferase expressing Listeria by RNAi screening.

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Innate immune system is one of the first-line defense system that is highly conserved from plants, insects and mammals. Peptidoglycan recognition protein (PGRP)-LE firstly identified in our laboratory is functioning on both intracellular and extracellular space and responsible for the activation of one of the innate immune signaling pathway, Imd pathway, and melanin synthesis pathway. Furthermore, PGRP-LE plays a crucial role to eliminate intracellular pathogens using one of the general protein degradation system called "Autophagy", which is independent on the Toll and Imd pathway. Therefore, to identify further novel host defense factors invoved in this multi-functional PGRP-LE, I set up the system where the growth of Listeria can be monitored by constitutive Luciferase expressing Listeria strains. Using this Listeria strain, the infection condition where we can see the effect of PGRP-LE was scrutinized. As an alternative strategy, I also conducted DNA microarray and identified a novel antimicrobial-like peptide named Listericin. Further analysis of Listericin using protein purification, RNAi and overexpression studies demonstrated that the expression of Listericin is cooperatively regulated by both PGRP-LE and the JAK-STAT pathway, as a secreted form, it exibits bacteriocidal activities against gram-negative bacteria and Listeria strain. Furthermore, in vivo approach using Listericin overexpressing transgenic flies showed that they exhibit resistance against Listeria infection in the surival test. Taken together with these results, we can show that Listericin is a novel antimicrobial peptide on Drosophila innate immune response.