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Infection by dsRNA viruses is associated with enhanced sporulation efficiency in Saccharoymces cerevisiae

Citation

Travers Cook, Thomas; Skirgaila, Christina; Martin, Oliver; Buser, Claudia (2022), Infection by dsRNA viruses is associated with enhanced sporulation efficiency in Saccharoymces cerevisiae, Dryad, Dataset, https://doi.org/10.5061/dryad.x0k6djhm8

Abstract

Upon starvation, diploid cells of the facultative sexual yeast Saccharoymces cerevisiae undergo sporulation, forming four metabolically quiescent and robust haploid spores encased in a degradable ascus. All endosymbionts, whether they provide net benefits or costs, utilise host resources; in yeast, this should induce an earlier onset of sporulation. Here, we tested whether the presence of endosymbiotic dsRNA viruses (M satellite and L-A helper) correspond with higher sporulation rate of their host, S. cerevisiae. We find that S. cerevisiae hosting both the M and L-A viruses (so-called “killer yeasts”) have significantly higher sporulation efficiency than those without. We also found that the removal of the M virus did not reduce sporulation frequency, possibly because the L-A virus still utilises host resources with and without the M virus. Our findings indicate that either virulent resource use by endosymbionts induces sporulation, or that viruses are spread more frequently to sporulating strains. Further exploration is required to distinguish cause from effect.

Methods

Study system

In total, 11 S. cerevisiae strains – five non-killers (DBVPG1985; DBVPG 6302, DBVPG4410, DBVPG4460, DBVPG6223) and six killers (NCYC 190, Y-2429, YJM1307, NCYC1001, Ca7, Ex198) – were used (Rodriguez-Cousino et al. 2013, Peter et al. 2018, Fredericks et al. 2021). Of the killer strains, three belonged to the K1 killer-type, two to K2 and one to Klus. For each killer strain sample, a corresponding M virus-free strain was created from treatment with elevated temperature (Wickner 1974), anisomycin (Fredericks et al. 2021) or cycloheximide (Fink and Styles 1972) to destroy genetic material of the M virus. This procedure is commonly referred to as “curing” the strains. Killer strains and their cured equivalent were donated and cured with the aforementioned methods by the groups of Paul Rowley and Nieves Rodríguez-Cousiño (Fredericks et al. 2021; Rodríguez-Cousiño et al. 2013). For a control, all non-killer strains were additionally independently exposed to elevated temperature (a widely utilised curing method) prior to the experiment (Wickner 1974), as this would allow us to establish whether curing of the killer factor per se or simply a curing process can influence sporulation efficiency. This also leads to a full factorial design of cured non-killers, non-killers, cured-killers and killers. Background information on each strain can be found in the respective collections of each strain.

Sporulation

Three independent replicates per S. cerevisiae strain were cultivated on standard YPD media (1% Yeast Extract, 2% Peptone, 10% Glucose) at 25°C overnight, and subsequently the optical densities (OD) of the growing yeast cells were measured (SpectraMax 19, software: SoftMax Pro 6.2.2). When the OD was between 0.6-0.8, 50 ml of yeast were washed according to the protocol of Knight and Goddard (2016), with the following modifications: washed twice with 3 ml of sterile water. 0.5 ml of each yeast strain was plated on sporulation plates (40 mm plates, sporulation media: 1% Potassium Acetate, 0.1% Yeast Extract, 0.05% Glucose, 2% Agar, 25 mg/L Zinc Sulphate). Yeast plates were inoculated for 7 days at 25° C. Finally, we evaluated 100 cells for each strain replicate under the microscope and classified their sporulation status to calculate the sporulation efficiency, defined as the proportion of cells in a culture that have sporulated (Knight and Goddard 2016).

References

Fink, G., and C. A. Styles. 1972. Curing of a killer factor in Saccharomyces cerevisiae. Proceedings of the National Academy of Sciences 69:2846-2849.

Fredericks, L. R., M. D. Lee, A. M. Crabtree, J. M. Boyer, E. A. Kizer, N. T. Taggart, C. R. Roslund, S. S. Hunter, C. B. Kennedy, C. G. Willmore, N. M. Tebbe, J. S. Harris, S. N. Brocke, and P. A. Rowley. 2021. The species-specific acquisition and diversification of a K1-like family of killer toxins in budding yeasts of the Saccharomycotina. PLoS Genet 17:e1009341.

Knight, S. J., and M. R. Goddard. 2016. Sporulation in soil as an overwinter survival strategy in Saccharomyces cerevisiae. FEMS Yeast Res 16:fov102.

Peter, J., M. De Chiara, A. Friedrich, J.-X. Yue, D. Pflieger, A. Bergström, A. Sigwalt, B. Barre, K. Freel, A. Llored, C. Cruaud, K. Labadie, J.-M. Aury, B. Istace, K. Lebrigand, P. Barbry, S. Engelen, A. Lemainque, P. Wincker, G. Liti, and J. Schacherer. 2018. Genome evolution across 1,011 Saccharomyces cerevisiae isolates. Nature 556:339-344.

Rodriguez-Cousino, N., P. Gomez, and R. Esteban. 2013. L-A-lus, a new variant of the L-A totivirus found in wine yeasts with Klus killer toxin-encoding Mlus double-stranded RNA: possible role of killer toxin-encoding satellite RNAs in the evolution of their helper viruses. Appl Environ Microbiol 79:4661-4674.

Wickner, R. B. 1974. “Killer character” of Saccharomyces cerevisiae: curing by growth at elevated temperature. Journal of Bacteriology 117:1356-1357.

Funding

ETH Zürich Foundation, Award: ETH-23 20-1