Multifaceted roles of the lysosome in the establishment and expansion of tuberculous infection

Abstract

Infection with Mycobacterium tuberculosis, the causative agent of TB, begins with the deposition of bacteria into the lung alveolus where they establish infection in host macrophages. M. tuberculosis dwells within host macrophages for much of its life cycle, until it again becomes extracellular prior to transmission into a new host. M. tuberculosis’ use of the macrophage as an expansion niche is surprising because these immune cells are normally tasked with the elimination of invading bacteria by trafficking them to degradative lysosomal compartments. Using the optically transparent and genetically tractable zebrafish model of mycobacterial infection with M. marinum, I have evaluated the diverse roles played by the macrophage lysosome at two stages of infection: first, during the initial interaction between infecting mycobacteria and first-responder macrophages; and second, during the stage of infection when bacteria are released from the macrophage and grow extracellularly. When newly infecting bacteria are phagocytosed by macrophages, their survival within these broadly microbicidal cells is thought to depend upon their ability to block phagosome-lysosome fusion and subsequent acidification. This notion persists despite multiple studies in cultured macrophages, spanning several decades, reporting a substantial proportion of viable mycobacteria in phagolysosomes. This work shows that while lysosomal trafficking in the macrophage can slow bacterial replication, it does not kill the pathogen. For its part, the mycobacterium survives within the macrophage lysosome through the action of a highly conserved virulence factor that mediates acid tolerance in vitro. A concurrent forward genetic screen identified a zebrafish mutant that is hypersusceptible to M. marinum infection and manifests hallmarks of human lysosomal storage diseases. The mutant is deficient in the lysosomal hydrolases Cathepsin B and L, although its hypersusceptibility does not stem from defective intramacrophage restriction of mycobacterial growth. Rather the mutant’s hypersusceptibility is attributed to defective macrophage migration owing to accumulated undegraded cell products that accumulate through normal homeostatic macrophage function. Similarly, macrophage migration defects underlie hypersusceptibility to mycobacterial infection following disruption of the zebrafish orthologs of genes implicated in human lysosomal storage disorders. All of these migration defects share a common mechanism in disrupted endocytic recycling caused by the lysosomal accumulation of undigestible cargo. Macrophage immotility resulting from lysosomal storage defects causes hypersusceptibility by compromising migration to dying infected cells in the granuloma, resulting in its breakdown with extracellular release of bacteria. Lysosomal storage defects also disrupt macrophage participation in the earliest stage of infection when resident macrophages migrate to and phagocytose newly arriving mycobacteria. The alveolar (lung resident) macrophages of human smokers display lysosomal accumulations of tobacco smoke and fail to migrate to M. tuberculosis. This may explain the increased susceptibility of human smokers to TB. This work defines the limited protection afforded to the host by lysosomal trafficking in the restriction of intramacrophage mycobacterial replication, and elucidates the consequences of lysosomal dysfunction, which results in host susceptibility through a completely distinct mechanism of macrophage incapacitation following the lysosomal accumulation of indigestible cargo.