Large-Scale Identification of Wolbachia pipientis Effectors

Abstract

Wolbachia pipientis is an intracellular symbiont of arthropods well known for the reproductive manipulations induced in the host and, more recently, for the ability of Wolbachia to block virus replication in insect vectors. Since Wolbachia cannot yet be genetically manipulated, and due to the constraints imposed when working with an intracellular symbiont, little is known about mechanisms used by Wolbachia for host interaction. Here we employed a bioinformatics pipeline and identified 163 candidate effectors, potentially secreted by Wolbachia into the host cell. A total of 84 of these candidates were then subjected to a screen of growth defects induced in yeast upon heterologous expression which identified 14 top candidates likely secreted by Wolbachia. These predicted secreted effectors may function in concert as we find that their native expression is correlated and is highly upregulated at specific time points during Drosophila development. In addition, the evolutionary histories of some of these predicted effectors are also correlated, suggesting they may function together, or in the same pathway, during host infection. Similarly, most of these predicted effectors are limited to one or two Wolbachia strains—perhaps reflecting shared evolutionary history and strain specific functions in host manipulation. Identification of these Wolbachia candidate effectors is the first step in dissecting the mechanisms of symbiont–host interaction in this important system.

Mi Casa es Su Casa: how an intracellular symbiont manipulates host biology

“…The absence of conventional molecular tools has made it relatively difficult to functionally characterize proteins that Wolbachia may use to manipulate its host, of which there may be many. However, through the use of heterologous systems (Whitaker et al ., ), eukaryotic model systems like S. cerevisiae (Sheehan et al ., ; Beckmann et al ., ; Rice et al ., ) and leveraging the power of genetics in Drosophila (Ote et al ., ; Sheehan et al ., ; LePage et al ., ), we can begin to identify symbiont effectors and host cellular processes affected.…”

“…Two loci ( cifA and cifB ) were identified and transgenically expressed in flies to recapitulate CI. Finally, bioinformatics was combined with a large‐scale yeast growth assay to identify Wolbachia secreted effectors from strain w Mel (Rice et al ., ). As part of that work, 84 candidate effectors were screened and 14 predicted effectors were identified, some encoding Ankyrin repeat domains but many encoding other domains primarily known from eukaryotic systems (Table ).…”

“…As such, they likely interact with host vesicle trafficking pathways and the host cytoskeleton to maintain vacuole integrity, promote division of the vacuole once the bacteria replicate, and allow for cell‐to‐cell transfer (Kumar and Valdivia, ; Creasey and Isberg, ). Wolbachia encode six proteins (WD0073, WD0224, WD0290, WD0353, WD0630 and WD0754) that share homology to Chlamydia tractomatis IncA (Rice et al ., ). In Chlamydia , the IncA proteins play a role in the recruitment of Rab effectors (Rzomp et al ., ) and may facilitate homotypic fusion of the Chlamydia vacuole (termed the inclusion) (Delevoye et al ., ).…”

“…Other w Mel predicted effectors that may facilitate vacuolar integrity include those which encode domains involved in cytoskeleton‐interaction (Table 1; synuclein, HAUS‐augmin3) (Rice et al ., ). Interactions between Wolbachia and host cytoskeletal elements have been well characterized.…”

“…Secretion of WalE1 was demonstrated via a heterologous assay in E. coli (Whitaker et al ., ) and upon transgenic expression, WalE1 was shown to increase Wolbachia load in Drosophila melanogaster females, allowing more efficient transmission of Wolbachia to the next generation (Sheehan et al ., ). Native expression of walE1 mRNA suggests that it is upregulated during host pupation, along with components of the type IV secretion system (Rice et al ., ). Characterization of native WalE1 function, and the host targets of this Wolbachia effector, will elucidate whether this actin bundler facilitates soma to germ line transmission and/or helps to maintain the Wolbachia vacuole.…”