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Thioester-containing proteins participate in the antibacterial immune defense of Drosophila against the pathogen Photorhabdus Open Access

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Thioester-containing proteins (TEPs) are present in many animal species ranging from deuterostomes to protostomes, which indicates their evolutionary importance in immune function. Phylogenetically, insect TEPs share sequence similarity with mammalian alpha-2 macroglobulins. Here I characterized the functions of TEPs in the immune response of Drosophila against two different Photorhabdus bacteria, P. luminescens and P. asymbiotica. In particular, I investigated several critical cellular and humoral immune responses as well as pathophysiological effects in loss-of-function tep mutants. I show that certain tep mutants survive infection with Photorhabdus bacteria significantly better during the initial hours of infection and have lower pathogen burden compared to their background controls. I also display that Tep2, Tep4 and Tep6 genes are transcriptionally upregulated in wild-type flies in response to Photorhabdus infection, mainly in the hemocytes and fat body tissues. The data signifies that TEP2, TEP4 and TEP6 are involved in immune signaling pathway activation in Drosophila in response to Photorhabdus infection. After estimating changes in hemocyte numbers and viability in the three tep mutants, I report that tep4 and tep6 mutants have significantly fewer live and dead hemocytes compared to background control flies in response to Photorhabdus infection. In addition, tep2 mutants exhibit significantly higher numbers of hemocytes after P. luminescens infection, but not after P. asymbiotica challenge. They also have more live hemocytes compared to background control flies responding to challenge with either Photorhabdus species. I further document that TEP2, TEP4 and TEP6 participate in phagocytosis, phenoloxidase and melanization response in Drosophila flies infected by Photorhabdus.Both tep2 and tep4 mutants display higher metabolic activity compared to background control flies upon infection with the pathogens. By performing Q-PCR on gut and fat body tissues, I find that tep mutants have significantly less pathogen burden compared to the background control flies. Furthermore, I report significant reduction in the transcript levels of Dronc, an apoptosis marker, in the tep mutants compared to the background control flies infected with Photorhabdus. These results indicate that the inactivation of Tep2 and Tep4 genes leads to reduced inflammation as well as metabolic depression in the flies upon infection with Photorhabdus bacteria.In conclusion, my work has shown for the first time that certain TEP molecules regulate humoral and cellular aspects of the Drosophila immune response against infection with Photorhabdus. Changes in the activation of immune signaling in flies with inactivated Tep genes result in lower levels of inflammation, higher metabolic activity and lower pathogen persistence, which leads to the increased survival during the initial hours of infection. These findings generate novel insights into the immune role of TEP molecules as regulators and effectors of the D. melanogaster antibacterial immune response and uncover a previously unknown layer of the innate immune system.

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