How did we get here? Insights into mechanisms of immunity-related GTPase targeting to intracellular pathogens

Abstract

The cytokine gamma-interferon activates cell-autonomous immunity against intracellular bacterial and protozoan pathogens by inducing a slew of antimicrobial proteins, some of which hinge upon immunity-related GTPases (IRGs) for their function. Three regulatory IRG clade M (Irgm) proteins chaperone about approximately 20 effector IRGs (GKS IRGs) to localize to pathogen-containing vacuoles (PVs) within mouse cells, initiating a cascade that results in PV elimination and killing of PV-resident pathogens. However, the mechanisms that allow IRGs to identify and traffic specifically to 'non-self' PVs have remained elusive. Integrating recent findings demonstrating direct interactions between GKS IRGs and lipids with previous work, we propose that three attributes mark PVs as GKS IRG targets: the absence of membrane-bound Irgm proteins, Atg8 lipidation, and the presence of specific lipid species. Combinatorial recognition of these three distinct signals may have evolved as a mechanism to ensure safe delivery of potent host antimicrobial effectors exclusively to PVs.

Figure 1.. Structural features of IRGs.

All IRGs contain a C-domain, G-domain, and N-domain. The G-domain contains the nucleotide binding pocket, with GKS IRGs such as Irga6, Irgb6, and Irgb10 containing a canonical GxxxxGKS amino acid sequence in the nucleotide-binding pocket and Irgm proteins containing a non-canonical GxxxxGMS sequence. Homo- and hetero-oligomerization among IRGs occurs via G-domain interactions. Membrane binding requires a conserved αK amphipathic helix in the C-domain. Additionally, Irga6 and Irgb6 possess an N-terminal myristoylation motif, and Irgm1 contains a C-terminal palmitoylation motif allowing for covalent lipid attachments that assist in membrane binding. In contrast, Irgb6 harbors a binding pocket for specific phospholipids such as PI5P. Figure created with BioRender.com

Figure 2.. Three-signal model for IRG recruitment.

Irgm proteins reside on the membranes of host organelles such as mitochondria, lipid droplet (LD), endoplasmic reticulum, and Golgi apparatus. As Irgm proteins inhibit GTPase activity of GKS IRGs, their absence (i.e. “missing self”) on the membranes of vacuoles containing intracellular pathogens such as C. trachomatis (green circles) renders those membranes permissive to GKS IRG recruitment. Conjugation of Atg8 proteins to phosphotidylethanolamine by Atg5/12/16L1 at the PVM comprises a second signal for GKS IRG recruitment. Finally, some IRGs such as Irgb6 specifically bind PV-enriched lipid species such as PI5P and may go on to recruit other IRGs such as Irga6 and Irgb10. Figure created with BioRender.com.