Carvalho, Ana Simão Pinto de
Functional Characterization of Putative Wolbachia Effector Proteins / Ana Simão Pinto de Carvalho
Functional Characterization of Putative Wolbachia Effector Proteins / Ana Simão Pinto de Carvalho . - Lisboa : NOVA Medical School, 2020 . - viii, 52 p. . -
Wolbachia is a widespread endosymbiont found in many arthropods and nematodes. Wolbachia induces different phenotypes depending on the combination of strain and host. One of the most striking phenotypes is antiviral protection conferred by Wolbachia to the infected host. This phenotype was observed not only in Drosophila melanogaster, but also in artificially infected mosquitoes, which are currently being released to fight against the spread of dengue and Zika. However, little is known about the mechanisms behind Wolbachia-mediated antiviral protection. Wolbachia encodes for an unusually high number of ankyrin-repeat proteins, which are used by prokaryotes to interact with host machinery. Moreover, one genomic region encoding for eight genes, called Octomom, is known to regulate Wolbachia proliferation. Additionally, it is known that Wolbachia titers correlate to the strength of Wolbachia-induced phenotypes. Ankyrin repeats and Octomom genes are candidate effector proteins, however, since Wolbachia is genetically intractable, we can only study these candidates using heterologous expression systems. In this project, we aimed at establishing a pipeline to functionally characterize Wolbachia effectors. To do so, we expressed the harmonized coding DNA sequences of candidate effectors in Drosophila melanogaster, both in cell culture and in whole flies. We were able to observe the subcellular localization of the candidate effectors in different cellular environments. Expression in whole flies infected with Wolbachia was used to assess the impact of expressing the putative effector in Wolbachia titers and transmission. By expressing the effector in flies with and without Wolbachia we will also assess for impact in antiviral protection. We have thus established a series of protocols to characterize Wolbachia putative effectors Wolbachia
Drosophila
DED Signaling Adaptor Proteins
Academic Dissertation
Portugal
616
Functional Characterization of Putative Wolbachia Effector Proteins / Ana Simão Pinto de Carvalho . - Lisboa : NOVA Medical School, 2020 . - viii, 52 p. . -
Wolbachia is a widespread endosymbiont found in many arthropods and nematodes. Wolbachia induces different phenotypes depending on the combination of strain and host. One of the most striking phenotypes is antiviral protection conferred by Wolbachia to the infected host. This phenotype was observed not only in Drosophila melanogaster, but also in artificially infected mosquitoes, which are currently being released to fight against the spread of dengue and Zika. However, little is known about the mechanisms behind Wolbachia-mediated antiviral protection. Wolbachia encodes for an unusually high number of ankyrin-repeat proteins, which are used by prokaryotes to interact with host machinery. Moreover, one genomic region encoding for eight genes, called Octomom, is known to regulate Wolbachia proliferation. Additionally, it is known that Wolbachia titers correlate to the strength of Wolbachia-induced phenotypes. Ankyrin repeats and Octomom genes are candidate effector proteins, however, since Wolbachia is genetically intractable, we can only study these candidates using heterologous expression systems. In this project, we aimed at establishing a pipeline to functionally characterize Wolbachia effectors. To do so, we expressed the harmonized coding DNA sequences of candidate effectors in Drosophila melanogaster, both in cell culture and in whole flies. We were able to observe the subcellular localization of the candidate effectors in different cellular environments. Expression in whole flies infected with Wolbachia was used to assess the impact of expressing the putative effector in Wolbachia titers and transmission. By expressing the effector in flies with and without Wolbachia we will also assess for impact in antiviral protection. We have thus established a series of protocols to characterize Wolbachia putative effectors Wolbachia
Drosophila
DED Signaling Adaptor Proteins
Academic Dissertation
Portugal
616
