Host actin cytoskeleton is often targeted by pathogenic bacteria through the secretion of effectors. Legionella pneumophila virulence relies on the injection of the largest known arsenal of bacterial proteins, over 300 Dot/Icm type 4 secretion system effectors, into the host cytosol. Here, we define the functional interactions between VipA and LegK2, two effectors with antagonistic activities towards actin polymerization that have been proposed to interfere with the endosomal pathway. We confirmed the prominent role of LegK2 effector in Legionella infection, as the deletion of legK2 results in defects in the inhibition of actin polymerization at the Legionella-containing vacuole, as well as in endosomal escape of bacteria and subsequent intracellular replication. More importantly, we observed the restoration of the ΔlegK2 mutant defects, upon deletion of vipA gene, making LegK2/VipA a novel example of effector-effector suppression pair that targets the actin cytoskeleton and whose functional interaction impacts L. pneumophila virulence. We demonstrated that LegK2 and VipA do not modulate each other’s activity in a “metaeffector” relationship. Instead, the antagonistic activities of the LegK2/VipA effector pair would target different substrates, Arp2/3 for LegK2 and G-actin for VipA, to temporally control actin polymerization at the LCV and interfere with phagosome maturation and endosome recycling, thus contributing to the intracellular life cycle of the bacterium. Strikingly, the functional interaction between LegK2 and VipA is consolidated by an evolutionary history that has refined the best effector repertoire for the benefit of L. pneumophila virulence.

中文翻译：

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军团菌效应器对对宿主肌动蛋白聚合的双重控制

宿主肌动蛋白细胞骨架通常通过效应物的分泌而被病原菌靶向。嗜肺军团菌的毒力依赖于将已知最大的细菌蛋白库（超过 300 个点/Icm 4 型分泌系统效应子）注射到宿主细胞质中。在这里，我们定义了 VipA 和 LegK2 之间的功能相互作用，这两个效应器对肌动蛋白聚合具有拮抗活性，已被提议干扰内体途径。我们证实了LegK2效应子在军团菌感染中的重要作用，因为legK2的缺失会导致含有军团菌的液泡中肌动蛋白聚合的抑制缺陷，以及细菌的内体逃逸和随后的细胞内复制。更重要的是，我们观察到删除vipA基因后ΔlegK2突变体缺陷的恢复，使 LegK2/VipA 成为效应器-效应器抑制对的新例子，其靶向肌动蛋白细胞骨架，其功能相互作用影响嗜肺军团菌毒力。我们证明 LegK2 和 VipA 不会以“元效应器”关系调节彼此的活动。相反，LegK2/VipA效应器对的拮抗活性将针对不同的底物，LegK2的Arp2/3和VipA的G-肌动蛋白，以暂时控制LCV处的肌动蛋白聚合并干扰吞噬体成熟和内体回收，从而有助于细菌的细胞内生命周期。引人注目的是，LegK2 和 VipA 之间的功能相互作用通过进化历史得以巩固，该进化历史为嗜肺军团菌毒力的益处而完善了最佳效应器库。