title: Host and bacterial factors that regulate LC3 recruitment to Listeria monocytogenes during the early stages of macrophage infection.
abstract: Listeria monocytogenes is a bacterial pathogen that can escape the phagosome and replicate in the cytosol of host cells during infection. We previously observed that a population (up to 35%) of L. monocytogenes strain 10403S colocalize with the macroautophagy marker LC3 at 1 h postinfection. This is thought to give rise to spacious Listeria-containing phagosomes (SLAPs), a membrane-bound compartment harboring slow-growing bacteria that is associated with persistent infection. Here, we examined the host and bacterial factors that mediate LC3 recruitment to bacteria at 1 h postinfection. At this early time point, LC3(+) bacteria were present within single-membrane phagosomes that are LAMP1(+). Protein ubiquitination is known to play a role in targeting cytosolic L. monocytogenes to macroautophagy. However, we found that neither protein ubiquitination nor the ubiquitin-binding adaptor SQSTM1/p62 are associated with LC3(+) bacteria at 1 h postinfection. Reactive oxygen species (ROS) production by the CYBB/NOX2 NADPH oxidase was also required for LC3 recruitment to bacteria at 1 h postinfection and for subsequent SLAP formation. Diacylglycerol is an upstream activator of the CYBB/NOX2 NADPH oxidase, and its production by both bacterial and host phospholipases was required for LC3 recruitment to bacteria. Our data suggest that the LC3-associated phagocytosis (LAP) pathway, which is distinct from macroautophagy, targets L. monocytogenes during the early stage of infection within host macrophages and allows establishment of an intracellular niche (SLAPs) associated with persistent infection.
authors.affiliation: Cell Biology Program; Hospital for Sick Children; Toronto, ON Canada; Institute of Medical Science; University of Toronto; Toronto, ON Canada.
authors.forename: Grace Y
authors.initials: GY
authors.lastname: Lam
authors.forename: Marija
authors.initials: M
authors.lastname: Cemma
authors.forename: Aleixo M
authors.initials: AM
authors.lastname: Muise
authors.forename: Darren E
authors.initials: DE
authors.lastname: Higgins
authors.forename: John H
authors.initials: JH
authors.lastname: Brumell
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D004075:Diglycerides / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000378:metabolism; D054884:Host-Pathogen Interactions; D008089:Listeria monocytogenes / Q000378:metabolism* / Q000648:ultrastructure; D008264:Macrophages / Q000382:microbiology* / Q000648:ultrastructure; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008954:Models, Biological; D019255:NADPH Oxidases / Q000378:metabolism; D010587:Phagocytosis; D010588:Phagosomes / Q000378:metabolism / Q000382:microbiology / Q000648:ultrastructure; D017382:Reactive Oxygen Species / Q000378:metabolism; D000071456:Sequestosome-1 Protein; D054875:Ubiquitination
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D004075:Diglycerides; D006360:Heat-Shock Proteins; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D017382:Reactive Oxygen Species; D000071456:Sequestosome-1 Protein; C101343:Sqstm1 protein, mouse; D019255:NADPH Oxidases
keywords: LC3; LC3-associated phagocytosis; Listeria monocytogenes; autophagy; diacylglycerol; innate immunity; reactive oxygen species; ubiquitin