Regulation of Legionella pathogenesis by meta-effector-effector interactions exemplified by the LubX-SidH pair

Abstract

Legionella pneumophila is a Gram-negative bacterial pathogen that causes Legionnaires' disease, a severe form of pneumonia in humans. It utilizes a multiprotein complex called the Dot/Icm secretion system to translocate a diverse set of bacterial proteins known as effectors into host cells during infection. L. pneumophila's effector arsenal comprises over 330 proteins, accounting for more than 10% of this bacterium's proteome. The functional redundancy and sheer number of L. pneumophila effectors complicate the characterization of their specific functions during infection, which remains poorly understood. Recent research has revealed that Legionella bacteria have a unique mechanism of controlling the activity of other effectors by utilizing a subset of effectors called meta-effectors. These meta-effectors modify the function of other effectors by targeting them within the host cell. One such example is LubX, a meta-effector encoded by the lpg2830 gene in L. pneumophila, which targets another effector, SidH (encoded by lpg2829), for ubiquitination and degradation by host proteasome. A large-scale analysis of effector-meta-effector interactions has identified nineteen such pairs, indicating that this type of regulation plays a crucial role in Legionella's pathogenic strategy. This study investigates the functional diversification of SidH across different Legionella strains, its interaction with the meta-effector LubX, and its potential mechanisms underlying SidH-mediated toxicity. A combination of structural analysis, in vitro experiments, and in cellulo assays was employed to investigate the characteristics of the SidH protein. The study also evaluated the interactions between SidH and LubX and the toxicity of SidH orthologs across Legionella species using a yeast model system. The C-terminus portion of SidH adopts an alpha-helical fold with no structural similarity to other characterized proteins and co-precipitates with host Rab11A. The research findings also indicate that LubX specifically ubiquitinates the N-terminal region of SidH, and LubX mutants, which lack E3 ubiquitin ligase function, are incapable of ubiquitinating the N-terminal region of SidH. The results provide insights into the functional diversification of SidH and the species-specific mechanisms of effector-meta-effector interactions in bacterial pathogenesis, highlighting the importance of meta-effectors in the pathogenic strategy and emphasizing the need for both effector interaction and functional regulation.