FUNCTIONAL CHARACTERIZATION OF LEGIONELLA GENUS CORE EFFECTORS

Infection by the human pathogen Legionella pneumophila relies on the translocation of ~300 virulence proteins, termed effectors, which manipulate host cell processes. Recently we sequenced, assembled and characterized the genomes of 38 Legionella species and predicted their effector repertoires using a previously validated machine learning approach. This analysis identified 5,885 predicted effectors. The effector repertoires of the different Legionella species were found to be largely non-overlapping, and only a small set of core effectors were shared by all the Legionella species. To characterize these core effectors, deletion mutants were constructed in the L. pneumophila homologs of these effectors and tested for intracellular growth in the environmental host A. castellanii. Three out of nine core effectors (30%) were found to be partially required for intracellular growth in this host and their growth defects were completely complemented by introducing the effectors on a plasmid. Due to the high degree of similarity between yeast and mammalian cells in pathways shown to be affected by L. pneumophila effectors, we cloned and expressed the core effectors in the yeast Saccharomyces cerevisiae. Four out of the nine core effectors caused different degrees of growth inhibition in yeast, indicating that they affect conserved eukaryotic pathways. To conclude, the high conservation of the core effectors throughout the evolution of the genus indicates that they perform critical functions during infection. The fact that 30% of the core effectors had an intracellular growth phenotype when deleted suggests that the function of the other core effectors may be redundant, at least for intracellular growth in A. castellanii. It is possible, however, that these core effectors carry out essential functions required for growth in other hosts.