Macrophages Rapidly Transfer Pathogens from Lipid Raft Vacuoles to Autophagosomes

Abstract

Macrophages activate autophagy as an immediate response to Legionella pneumophila infection, but what marks the pathogen phagosome as a target for the autophagy machinery is not known. Because a variety of bacteria, parasites, viruses, and toxins that associate with the endoplasmic reticulum enter host cells by a cholesterol-dependent route, we tested the hypothesis that autophagy is triggered when microbes engage components of lipid raft domains. As the intracellular respiratory pathogen L. pneumophila or the extracellular uropathogen FimH(+) Escherichia coli entered macrophages by a cholesterol-sensitive mechanism, they immediatezly resided in vacuoles rich in glycosylphosphatidylinositol moieties and the autophagy enzyme Atg7. As expected for autophagosomes, the vacuoles sequentially acquired the endoplasmic reticulum protein BiP, the autophagy markers Atg8 and monodansyl-cadaverine, and the lysosomal protein LAMP-1. A robust macrophage response to the pathogens was cholesterol-dependent, since fewer Atg7-rich vacuoles were observed when macrophages were pretreated with methyl-beta-cyclodextrin or filipin. A model in which macrophages exploit autophagy to capture pathogens within the lipid raft pathway for antigen presentation prior to disposal in lysosomes is discussed.

Francisella Targets Cholesterol-Rich Host Cell Membrane Domains for Entry into Macrophages

“…There is some evidence that such a transfer may be substantially facilitated by a previous enclosure in a cholesterol-rich vesicle (65).…”

Nanoparticles as a Novel Class of Autophagy Activators

“…23 This strategy can have an impact simultaneously on two steps in pathogenesis: additional autophagy activation, and targeted enzyme delivery to the lyso/phagosomal compartment. NPs, containing antimicrobial substances, can be used when persisting microorganisms compromise phago/lysosomal fusion 6,24 in parallel with the excessive autophagy activation; 25 but in this case NPs should not increase the autophagic activity. Moreover, targeted delivery of NPs to the microbe-containing phagosome is potentially possible.…”

Depletion of the Ubiquitin-binding Adaptor Molecule SQSTM1/p62 from Macrophages Harboring cftr ΔF508 Mutation Improves the Delivery of Burkholderia cenocepacia to the Autophagic Machinery

“…A11008). Nuclei were stained with the nucleic acid dye DAPI (6,35). Samples were analyzed with an Olympus Fluoview FV10i confocal microscope at the Ohio State University, Department of Microbial Infection and Immunity.…”

“…Confocal Microscopy-Immunofluorescence experiments for colocalization with autophagy markers were performed as described previously (34,6). Rabbit anti-LC3 (Abgent, catalog no.…”

“…Autophagy is a conserved physiological process that eliminates non-functional organelles and recycles cytosolic components to generate nutrients during periods of stress or starvation (5,6). Autophagy also targets cytosolic long-lived proteins and organelles for lysosomal degradation in eukaryotic cells and plays a role in innate immunity (7).…”