Functional diversification despite structural congruence in the HipBST toxin-antitoxin system ofLegionella pneumophila

ABSTRACT Toxin-antitoxin (TA) systems are abundant genetic modules in bacterial chromosomes and on mobile elements. They are often patchily distributed and their physiological functions remain poorly understood. Here, we characterize a TA system in Legionella pneumophila that is highly conserved across Legionella species. This system is distantly related to Escherichia coli HipBST and we demonstrate that it is a functional tripartite TA system (denoted HipBST Lp ). We identify HipBST Lp homologs in diverse taxa, yet in the Gammaproteobacteria these are almost exclusively found in Legionella species. Notably, the toxin HipT Lp was previously reported to be a pathogenic effector protein that is translocated by L. pneumophila into its eukaryotic hosts. Contrary to this, we find no signal of HipT Lp translocation beyond untranslocated control levels and make several observations consistent with a canonical role as a bacterial toxin. We present structural and biochemical insights into the regulation and neutralization of HipBST Lp , and identify key variations between this system and HipBST Ec . Finally, we show that the target of HipT Lp is likely not conserved with any characterized HipA or HipT toxin. This work serves as a unique comparison of a TA system across bacterial species and illustrates the molecular diversity that exists within a single TA family.