Wolbachia is an intracellular bacterium that infects a remarkable range of insect hosts. Insects such as mosquitos act as vectors for many devastating human viruses such as Dengue, West Nile, and Zika. Remarkably, Wolbachia infection provides insect hosts with resistance to many arboviruses thereby rendering the insects ineffective as vectors. To utilize Wolbachia effectively as a tool against vector-borne viruses a better understanding of the host-Wolbachia relationship is needed. To investigate Wolbachia-insect interactions we used the Wolbachia/Drosophila model that provides a genetically tractable system for studying host-pathogen interactions. We coupled genome-wide RNAi screening with a novel high-throughput fluorescence in situ hybridization (FISH) assay to detect changes in Wolbachia levels in a Wolbachia-infected Drosophila cell line JW18. 1117 genes altered Wolbachia levels when knocked down by RNAi of which 329 genes increased and 788 genes decreased the level of Wolbachia. Validation of hits included in depth secondary screening using in vitro RNAi, Drosophila mutants, and Wolbachia-detection by DNA qPCR. A diverse set of host gene networks was identified to regulate Wolbachia levels and unexpectedly revealed that perturbations of host translation components such as the ribosome and translation initiation factors results in increased Wolbachia levels both in vitro using RNAi and in vivo using mutants and a chemical-based translation inhibition assay. This work provides evidence for Wolbachia-host translation interaction and strengthens our general understanding of the Wolbachia-host intracellular relationship.
1Wolbachia is an intracellular bacterium that infects a remarkable range of insect hosts. 2Insects such as mosquitos act as vectors for many devastating human viruses such as 3 Dengue, West Nile, and Zika. Remarkably, Wolbachia infection provides insect hosts with 4 resistance to many arboviruses thereby rendering the insects ineffective as vectors. To 5 utilize Wolbachia effectively as a tool against vector-borne viruses a better understanding 6 of the host-Wolbachia relationship is needed. To investigate Wolbachia-insect 7 interactions we used the Wolbachia/Drosophila model that provides a genetically 8 tractable system for studying host-pathogen interactions. We coupled genome-wide RNAi 9 screening with a novel high-throughput fluorescence in situ hybridization (FISH) assay to 10 detect changes in Wolbachia levels in a Wolbachia-infected Drosophila cell line JW18. 11 1117 genes altered Wolbachia levels when knocked down by RNAi of which 329 genes 12 increased and 788 genes decreased the level of Wolbachia. Validation of hits included in 13 depth secondary screening using in vitro RNAi, Drosophila mutants, and Wolbachia-14 detection by DNA qPCR. A diverse set of host gene networks was identified to regulate 15 Wolbachia levels and unexpectedly revealed that perturbations of host translation 16 components such as the ribosome and translation initiation factors results in increased 17 Wolbachia levels both in vitro using RNAi and in vivo using mutants and a chemical-based 18 translation inhibition assay. This work provides evidence for Wolbachia-host translation 19 interaction and strengthens our general understanding of the Wolbachia-host intracellular 20 relationship. 21 22 23 Author summary 24Insects such as mosquitos act as vectors to spread devastating human diseases such as 25Dengue, West Nile, and Zika. It is critical to develop control strategies to prevent the 26 transmission of these diseases to human populations. A novel strategy takes advantage 27 of an endosymbiotic bacterium Wolbachia pipientis. The presence of this bacterium in 28 insect vectors prevents successful transmission of RNA viruses. The degree to which 29 viruses are blocked by Wolbachia is dependent on the levels of the bacteria present in 30 the host such that higher Wolbachia levels induce a stronger antiviral effect. In order to 31 use Wolbachia as a tool against vector-borne virus transmission a better understanding 32 of host influences on Wolbachia levels is needed. Here we performed a genome-wide 33RNAi screen in a model host system Drosophila melanogaster infected with Wolbachia 34 to identify host systems that affect Wolbachia levels. We found that host translation can 35influence Wolbachia levels in the host. 36 37 Wolbachia-host interactions can be studied in the genetically tractable Drosophila 63 melanogaster system which allows for the systematic dissection of host signaling 64 pathways that interact with the bacteria using the wide array of genetic and genomic tools 65 available. The Drosophila system enables rapid unbias...
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