IFNγ inducible GTPases mediate host Resistance against Chlamydia trachomatis via autophagy

Chlamydial infection of the host cell induces Gamma interferon (IFNγ), a central immunoprotector for humans and mice. The major effector mechanism for IFNγ differs substantially between humans and mice: in humans epithelial cells, indoleamine 2,3-dioxygenase (IDO), which catabolizes tryptophan (Trp), limits intracellular chlamydial growth by starvation for this amino acid, whereas in mice IDO does not appear to play a substantial role in the control of chlamydial infection. The primary antichlamydial effector mechanism in mouse cells involves the recently described IFNγ-inducible family of immunity related GTPases (IRG) proteins. The IFNγ-inducible immunity-related GTPase Irga6 was previously shown to play a role in defense against Chlamydia trachomatis in murine cells. Irga6 was identified as an important resistance factor against C. trachomatis infection in IFNγ-induced mouse embryonic fibroblast (MEFs) but not C. muridarum. In the first part of this study, the mechanism by which Irga6 confers resistance to intracellular C. trachomatis was studied. Irga6, Irgd, Irgm2 and Irgm3 accumulated at bacterial vacuoles in MEFs upon stimulation with IFNγ. This accumulation triggered a rerouting of the bacterial vacuoles to autophagosomes that subsequently fuse to lysosomes for elimination. Impairment of lysosomal acidification as well as the use of autophagy-deficient cells hindered the IFNγ-induced antimicrobial effects. While Irgm2, Irgm3 and Irgd still localized to C. trachomatis early inclusions in IFNγ-induced ATG5-/- cells, Irga6 localization was disrupted, indicating a special mechanism of Irga6 recruitment. Further, Irgd, Irgm3 and Irgm2 still localized to C. trachomatis early inclusions in IFNγ-treated autophagy-lacking MEFs except for Irga6, indicating a pivotal role for Irga6 and autophagy in resistance to the microbe. Strikingly, Irga6-deficient (Irga6-/-) MEFs, in which chlamydial growth is enhanced compared to wild type (WT) MEFs, did not respond to IFNγ even though Irgb6, Irgd, Irgm2 and Irgm3 still localized to C. trachomatis early inclusions. In addition, these cells were unable to capture C. trachomatis in autophagosomes, enabling survival of the pathogen. In human macrophages neither toxic nitric oxide (NO) nor Trp depletion appear to play a substantial role in the control of C. trachomatis infection, suggesting that known mechanisms of growth inhibition do not hold in human macrophages. Thus, in the second part of this study, the antichlamydial effector mechanism of IFNγ-inducible guanylate binding proteins (GBPs) in human macrophages was studied. The IFNγ-inducible GBPs were previously shown to play a role in defense against viral replication, regulation of vasculogenesis and have an anti-proliferative effect in endothelial cells. Here, GBP1 and GBP2 were identified as important resistance factors against C. trachomatis infection in IFNγ-induced human macrophages. Further, the mechanism by which IFNγ-inducible GBPs confer partial resistance to C. trachomatis was identified. IFNγ was found to trigger a rerouting of the bacterial vacuoles to lysosomes for elimination. Knock-down of either GBP1 or GBP2 induced resistance against IFNγ-induced killing of C. trachomatis and thus enabled survival of the pathogen. Taken together, these findings identify IFNγ-inducible GTPases as necessary factors in conferring host resistance by remodelling a classically nonfusogenic intracellular pathogen to stimulate fusion with autophagosomes, thereby rerouting the intruder to the lysosomal compartment for destruction.