No abstract

The soil-dwelling social amoeba Dictyostelium discoideum feeds on bacteria. Each meal is a potential infection because some bacteria have evolved mechanisms to resist predation. To survive such a hostile environment, D. discoideum has in turn evolved efficient antimicrobial responses that are intertwined with phagocytosis and autophagy, its nutrient acquisition pathways. The core machinery and antimicrobial functions of these pathways are conserved in the mononuclear phagocytes of mammals, which mediate the initial, innate-immune response to infection. In this review, we discuss the advantages and relevance of D. discoideum as a model phagocyte to study cell-autonomous defenses. We cover the antimicrobial functions of phagocytosis and autophagy and describe the processes that create a microbicidal phagosome: acidification and delivery of lytic enzymes, generation of reactive oxygen species, and the regulation of Zn2+, Cu2+, and Fe2+ availability. High concentrations of metals poison microbes while metal sequestration inhibits their metabolic activity. We also describe microbial interference with these defenses and highlight observations made first in D. discoideum. Finally, we discuss galectins, TNF receptor-associated factors, tripartite motif-containing proteins, and signal transducers and activators of transcription, microbial restriction factors initially characterized in mammalian phagocytes that have either homologs or functional analogs in D. discoideum.

Autophagy is a eukaryotic catabolic process also participating in cell-autonomous defence. Infected host cells generate double-membrane autophagosomes that mature in autolysosomes to engulf, kill and digest cytoplasmic pathogens. However, several bacteria subvert autophagy and benefit from its machinery and functions. Monitoring infection stages by genetics, pharmacology and microscopy, we demonstrate that the ESX-1 secretion system of Mycobacterium marinum, a close relative to M. tuberculosis, upregulates the transcription of autophagy genes, and stimulates autophagosome formation and recruitment to the mycobacteria-containing vacuole (MCV) in the host model organism Dictyostelium. Antagonistically, ESX-1 is also essential to block the autophagic flux and deplete the MCV of proteolytic activity. Activators of the TORC1 complex localize to the MCV in an ESX-1-dependent manner, suggesting an important role in the manipulation of autophagy by mycobacteria. Our findings suggest that the infection by M. marinum activates an autophagic response that is simultaneously repressed and exploited by the bacterium to support its survival inside the MCV.

Phagocytic cells capture and kill most invader microbes within the bactericidal phagosome, but some pathogens subvert killing by damaging the compartment and escaping to the cytosol. To prevent the leakage of pathogen virulence and host defence factors, as well as bacteria escape, host cells have to contain and repair the membrane damage, or finally eliminate the cytosolic bacteria. All eukaryotic cells engage various repair mechanisms to ensure plasma membrane integrity and proper compartmentalization of organelles, including the Endosomal Sorting Complex Required for Transport (ESCRT) and autophagy machineries. We show that during infection of Dictyostelium discoideum with Mycobacterium marinum, the ESCRT-I component Tsg101, the ESCRT-III protein Snf7/Chmp4/Vps32 and the AAA-ATPase Vps4 are recruited to sites of damage at the Mycobacterium-containing vacuole. Interestingly, damage separately recruits the ESCRT and the autophagy machineries. In addition, the recruitment of Vps32 and Vps4 to repair sterile membrane damage depends on Tsg101 but appears independent of Ca2+. Finally, in absence of Tsg101, M. marinum accesses prematurely the cytosol, where the autophagy machinery restricts its growth. We propose that ESCRT has an evolutionary conserved function to repair small membrane damage and to contain intracellular pathogens in intact compartments.

Autophagy is a fast-moving field with an enormous impact on human health and disease. Understanding the complexity of the mechanism and regulation of this process often benefits from the use of simple experimental models such as the social amoeba Dictyostelium discoideum. Since the publication of the first review describing the potential of D. discoideum in autophagy, significant advances have been made that demonstrate both the experimental advantages and interest in using this model. Since our previous review, research in D. discoideum has shed light on the mechanisms that regulate autophagosome formation and contributed significantly to the study of autophagy-related pathologies. Here, we review these advances, as well as the current techniques to monitor autophagy in D. discoideum. The comprehensive bioinformatics search of autophagic proteins that was a substantial part of the previous review has not been revisited here except for those aspects that challenged previous predictions such as the composition of the Atg1 complex. In recent years our understanding of, and ability to investigate, autophagy in D. discoideum has evolved significantly and will surely enable and accelerate future research using this model.