Prophage WO genes recapitulate and enhance Wolbachia-induced cytoplasmic incompatibility

Daniel P. LePage, Jason A. Metcalf, Sarah R. Bordenstein, Jungmin On, Jessamyn I. Perlmutter, J. Dylan Shropshire, Emily M. Layton, Lisa J. Funkhouser-Jones, John F. Beckmann, Seth R. Bordenstein
2017 Nature  
The genus Wolbachia is an archetype of maternally inherited intracellular bacteria that infect the germline of numerous invertebrate species worldwide. They can selfishly alter arthropod sex ratios and reproductive strategies to increase the proportion of the infected matriline in the population. The most common reproductive manipulation is cytoplasmic incompatibility, which results in embryonic lethality in crosses between infected males and uninfected females. Females infected with the same
more » ... lbachia strain rescue this lethality. Despite more than 40 years of research 1 and relevance to symbiont-induced speciation 2,3 , as well as control of arbovirus vectors 4-6 and agricultural pests 7 , the bacterial genes underlying cytoplasmic incompatibility remain unknown. Here we use comparative and transgenic approaches to demonstrate that two differentially transcribed, co-diverging genes in the eukaryotic association module of prophage WO 8 from Wolbachia strain wMel recapitulate and enhance cytoplasmic incompatibility. Dual expression in transgenic, uninfected males of Drosophila melanogaster crossed to uninfected females causes embryonic lethality. Each gene additively augments embryonic lethality in crosses between infected males and uninfected females. Lethality associates with embryonic defects that parallel those of wild-type cytoplasmic incompatibility and is notably rescued by wMel-infected embryos in all cases. The discovery of cytoplasmic incompatibility factor genes cifA and cifB pioneers genetic studies of prophage WOinduced reproductive manipulations and informs the continuing use of Wolbachia to control dengue and Zika virus transmission to humans. We hypothesized that the genes responsible for cytoplasmic incompatibility (CI) (Extended Data Fig. 1a ) are present in all CI-inducing Wolbachia strains and absent or divergent in non-CI strains; we also predicted that these genes are expressed in the gonads of infected insects. To elucidate CI candidates, we determined the core genome shared by the CI-inducing Wolbachia strains wMel (from D. melanogaster), wRi (from Drosophila simulans), wPip (Pel strain from Culex pipiens), and wRec (from Drosophila recens), while excluding the pan-genome of the mutualistic strain wBm (from Brugia malayi). This yielded 113 gene families representing 161 unique wMel genes ( Fig. 1a and Supplementary Table 1) . We further narrowed this list by comparing it with (1) homologues of genes previously determined by comparative genomic hybridization to be absent or divergent in the strain wAu 9 , a non-CI strain, (2) homologues to genes highly expressed at the RNA level in wVitA-infected Nasonia vitripennis ovaries, and (3) homologues detected at the protein level in wPip (Buckeye)-infected C. pipiens ovaries. We included ovarian data with the reasoning that CI genes might be generally expressed in infected reproductive tissues, or that the CI induction and rescue genes might be the same. Remarkably, only two genes, wMel locus tags WD0631 and WD0632, were shared among all four gene subsets ( Fig. 1b and to A.M.F. Imaging was performed in part through the use of the Vanderbilt University Medical Center Cell Imaging Shared Resource (supported by NIH grants CA68485, DK20593, DK58404, DK59637, and EY08126). We thank K. Jernigan and P. Snider for help with preliminary studies, and A. Brooks for assistance with figure preparation. Author Contributions D.P.L. performed gene expression and hatch rate assays, embryo cytology, and assayed for transgene and infection status of flies. J.A.M. performed comparative genomics analyses, generated transgenic flies, and drafted the manuscript. Sarah R.B. performed evolutionary and bioinformatic analyses. J.O. performed hatch rates, assayed sex ratios, collected flies for all experiments, and assayed for transgene and infection status of flies. J.I.P. conducted younger brother effect experiments and performed embryo cytology. J.D.S. performed hatch rate assays, collected flies for parallel embryo cytology, and assayed for transgene and infection status of flies. E.M.L. collected flies and performed hatch rate assays. L.J.F.-J. obtained the wVitA transcriptome.
doi:10.1038/nature21391 pmid:28241146 pmcid:PMC5358093 fatcat:ue5vrpuynvat3mw2qlrhcx2uwq