Dataset Information

Proteomic comparison of uninfected and wMelPop infected Aag2 cells

ABSTRACT: Wolbachia endosymbionts are widespread intracellular, maternally inherited bacteria which manipulate the host to favour their spread through a population. Cytoplasmic incompatibility (CI) and viral suppression are two such manipulations driven by uncharacterised mechanisms. To gain insight into potential molecular mechanisms responsible for these effects, we performed the first in-depth proteomic characterisation of the host response to Wolbachia infection. We find that the presence of the Wolbachia wMelPop in Aedes aegypti mosquito cells alters levels of proteins involved in cell cycle control, DNA replication, autophagy, vesicular trafficking, iron homeostasis, amino acid degradation, purine metabolism, lipid metabolism and immunity. The majority of the cell cycle/DNA replication proteins were downregulated in the presence of Wolbachia, in particular, a member of the anoctamin family was strongly down regulated which may be related to the chromosomal segregation defects observed in CI crosses. We found clear evidence for perturbed lipid metabolism, Wolbachia infected cells expressed higher levels of Apolipoprotein D and the cholesterol efflux transporter ABCA1, while the LDL receptor and fatty acid synthase were downregulated. ABCA1 was also upregulated at the mRNA level in adult Wolbachia infected Aedes aegypti mosquitoes. Wolbachia therefore perturbs cholesterol homeostasis causing the host cell to respond to an apparent cholesterol excess which may contribute to viral inhibition.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Aedes Aegypti

SUBMITTER: Vincent Geoghegan

LAB HEAD: Steven P. Sinkins

PROVIDER: PXD003429 | Pride | 2017-09-19

REPOSITORIES: Pride

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Publications

Perturbed cholesterol and vesicular trafficking associated with dengue blocking in Wolbachia-infected Aedes aegypti cells.

Geoghegan Vincent V Stainton Kirsty K Rainey Stephanie M SM Ant Thomas H TH Dowle Adam A AA Larson Tony T Hester Svenja S Charles Philip D PD Thomas Benjamin B Sinkins Steven P SP

Nature communications 20170913 1

Wolbachia are intracellular maternally inherited bacteria that can spread through insect populations and block virus transmission by mosquitoes, providing an important approach to dengue control. To better understand the mechanisms of virus inhibition, we here perform proteomic quantification of the effects of Wolbachia in Aedes aegypti mosquito cells and midgut. Perturbations are observed in vesicular trafficking, lipid metabolism and in the endoplasmic reticulum that could impact viral entry a ...[more]

PMID: 28904344

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Project description:Globally invasive Aedes aegypti mosquitoes disseminate numerous arboviruses that impact human health. One promising method to control Ae. aegypti populations is transinfection with the intracellular bacterium Wolbachia pipientis, a symbiont that naturally infects ~40-52% of insects but is normally absent from Ae. aegypti. Transinfection of Ae. aegypti with the wMel Wolbachia strain induces cytoplasmic incompatibility (CI), allowing infected individuals to rapidly invade native populations. Further, wMel Wolbachia-infected females display refractoriness to medically relevant arboviruses. Thus, wMel Wolbachia-infected Ae. aegypti are being released in several areas to replace native populations, thereby suppressing disease transmission by this species. Wolbachia is reported to have minimal effects on Ae. aegypti fertility, but its influence on other reproductive processes is unknown. Female insects undergo several post-mating physiological and behavioral changes required for optimal fertility. Post-mating responses (PMRs) in female insects are typically elicited by receipt of male seminal fluid proteins (SFPs) transferred with sperm during mating, but can be modified by other factors, such as adult age, nutritional status, and microbiome composition. To assess how Wolbachia infection influences Ae. aegypti female PMRs, we collected wMel Wolbachia-infected Ae. aegypti from the field in Medellín, Colombia and introduced the bacterium into our laboratory strain. We found that Wolbachia influences female fecundity, fertility, and re-mating incidence. Further, we observed that Wolbachia significantly extends longevity of virgin females. Changes in female PMRs are not due to defects in sperm transfer by infected males, or sperm storage by infected females. Using proteomic methods to examine the seminal proteome of infected males, we found that Wolbachia infection has a moderate effect on SFP composition. However, we identified 125 Wolbachia proteins that are paternally transferred to females by infected males. Surprisingly, the CI factor proteins (Cifs), were not detected in the ejaculates of Wolbachia-infected males. Our findings indicate that Wolbachia infection of Ae. aegypti alters female post-mating responses, potentially influencing control programs that utilize Wolbachia-infected individuals.

Project description:The Drosophila-Wolbachia system is being used to study the molecular nature of the interactions between a host and a symbiont. This system offers a unique opportunity for such a study since the Drosophila genome sequence is available, several Wolbachi genomes will also be available soon and there are at least three known Wolbachia strains infecting Drosophila: a) mod+ strain that induces cytoplasmic incompatibility, b) mod- strain that cannot induce cytoplasmic incompatibility, and c) popcorn strain, a virulent strain which reduces in half the adult lifespan of Drosophila due to its massive proliferation in adult brain. The Drosophila-Wolbachia interaction manifests itself in 3 main ways; first, destruction of the CNS in infected adults, second, induction of some kind of modification or imprinting in the male germ-line resulting in an early failure in embryonic development, (cytoplasmic incompatability (CI)) and third, modification of the female germ-line resulting in resistance to modified sperm. We are interested in identifying Drosophila genes with changes in expression due to Wolbachia infection. We have generated a series of isogenic fly lines (those being used in the IGF P-element project) which we have infected with Wolbachia strains, infection is readily cured by growth on medium containing tetracycline. Thus, we have equivalent genetic background with and without the parasite. We have tested all of the transgenic lines for the level of CI and find strain-specific levels ranging from 0-50%. We also have a strain of D. simulans that shows over 95% CI. Plan: For our initial experiments we wish to make 4 comparisons, in all cases 2 day old males will be collected and for each comparison we will isolate 3 independent biological replicates: Melanogaster no CI [tet] x Melanogaster no CI [+wol] Melanogaster high CI [tet] x Melanogaster high CI [+wol] We will therefore identify genes with changed expression levels in the male upon Wolbachia infection by comparing the melanogaster strains with high or no CI in the presence of tetracycline and Wolbachia. We also hope to identify similar genes in simulans (where we expect the magnitude of the effect to be larger), differences between melanogaster and simulans are controlled for in the mel v sim comparison.

Dataset Information

Proteomic comparison of uninfected and wMelPop infected Aag2 cells

Publications

Perturbed cholesterol and vesicular trafficking associated with dengue blocking in Wolbachia-infected Aedes aegypti cells.

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