Characterizing host translation inhibition by Legionella pneumophila

Project Summary Translation is a dynamic and energetically demanding fundamental process requiring numerous factors that promote and regulate protein synthesis. Dysregulation of translation often occurs in many human diseases. For example, many pathogens promote their own replication by targeting these translation factors, the ribosome, and regulatory mechanisms to undermine translation in their host cell. One such bacterium is Legionella pneumophila, the causative agent of the severe pneumonia called “Legionnaires’ disease”. The incidence of Legionella infections in the United States has increased in recent years, necessitating a better understanding of how this bacterium interacts with its host cell. Legionella translocates hundreds of toxic proteins, termed effectors, that subvert many biochemical processes within its host. Notably, at least eleven of these effectors inhibit translation. These effectors exert their function through a range of mechanisms, and most appear to be enzymes that modify translation factors to inactivate them while others mark proteins for degradation. Despite several studies describing the mechanisms of some of these translation-inhibiting effectors, it is unclear why Legionella employs them. This is driven by the fact that the mechanisms of some effectors remain elusive but also because their seemingly redundant nature has made it difficult to isolate their physiological relevance. This proposal seeks to close the gaps in knowledge regarding the translation-inhibiting effectors by using both specific and broad approaches. First, I will determine the mechanism of one of these effectors, SidL. Based on strong preliminary data, I propose a set of experiments to characterize its function and identify its molecular target. Second, I propose a comprehensive temporal analysis of translation inhibition during the Legionella infection cycle. Legionella has a ~24-hour infection cycle and some studies suggest that the translation-inhibiting effectors are temporally regulated; however, a careful characterization of when each effector works precludes a deeper understanding of them. Together, these two approaches will establish a framework to understand the physiological relevance of the translation-inhibiting effectors of Legionella. Furthermore, because these effectors target host cell factors, this work can reveal novel insights into our own cell biology.