IFN-γ orchestrates the host response against intracellular pathogens. Members of the guanylate binding proteins (GBP) comprise the most abundant IFN-γ-induced transcriptional response. mGBPs are GTPases that are specifically up-regulated by IFN-γ, other proinflammatory cytokines, toll-like receptor agonists, as well as in response to Listeria monocytogenes and Toxoplasma gondii infection. mGBP2 localizes at the parasitophorous vacuole (PV) of T. gondii; however, the molecular function of mGBP2 and its domains in T. gondii infection is not known. Here, we show that mGBP2 is highly expressed in several cell types, including T and B cells after stimulation. We provide evidence that the Cterminal domain is sufficient and essential for recruitment to the T. gondii PV. Functionally, mGBP2 reduces T. gondii proliferation because mGBP2-deficient cells display defects in the replication control of T. gondii. Ultimately, mGBP2-deficient mice reveal a marked immune susceptibility to T. gondii. Taken together, mGBP2 is an essential immune effector molecule mediating antiparasitic resistance.pathogen defense | cell-autonomous immunity | host-pathogen interaction
GBPs are essential for immunity against intracellular pathogens, especially for Toxoplasma gondii control. Here, the molecular interactions of murine GBPs (mGBP1/2/3/5/6), homo- and hetero-multimerization properties of mGBP2 and its function in parasite killing were investigated by mutational, Multiparameter Fluorescence Image Spectroscopy, and live cell microscopy methodologies. Control of T. gondii replication by mGBP2 requires GTP hydrolysis and isoprenylation thus, enabling reversible oligomerization in vesicle-like structures. mGBP2 undergoes structural transitions between monomeric, dimeric and oligomeric states visualized by quantitative FRET analysis. mGBPs reside in at least two discrete subcellular reservoirs and attack the parasitophorous vacuole membrane (PVM) as orchestrated, supramolecular complexes forming large, densely packed multimers comprising up to several thousand monomers. This dramatic mGBP enrichment results in the loss of PVM integrity, followed by a direct assault of mGBP2 upon the plasma membrane of the parasite. These discoveries provide vital dynamic and molecular perceptions into cell-autonomous immunity.DOI:
http://dx.doi.org/10.7554/eLife.11479.001
Members of the murine guanylate-binding protein family (mGBP) are induced by interferon gamma (IFN-γ) and have been shown to be important factors in cell-autonomous immunity toward the intracellular pathogen Toxoplasma gondii. Previously, we identified that mGBP2 mediates disruption of the parasitophorous vacuole membrane (PVM) and directly assaults the plasma membrane of the parasite. Here, we show that mGBP7-deficient mice are highly susceptible to T. gondii infection. This is demonstrated by the loss of parasite replication control, pronounced development of ascites, and death of the animals in the acute infection phase. Interestingly, live-cell microscopy revealed that mGBP7 recruitment to the PVM occurs after mGBP2 recruitment, followed by disruption of the PVM and T. gondii integrity and accumulation of mGBP7 inside the parasite. This study defines mGBP7 as a crucial effector protein in resistance to intracellular T. gondii.
IMPORTANCE Guanylate-binding proteins (GBPs) are induced by the inflammatory cytokine interferon gamma (IFN-γ) and have been shown to be important factors in the defense of the intracellular pathogen Toxoplasma gondii. In previous studies, we showed that members of the mouse GBP family, such as mGBP2 and mGBP7, accumulate at the parasitophorous vacuole of T. gondii, which is the replicatory niche of the parasite. In this study, we show that mice deficient in mGBP7 succumb early after infection with T. gondii, showing a complete failure of resistance to the pathogen. On a molecular level, mGBP7 is found directly at the parasite, likely mediating its destruction.
Chlamydia, the most common sexually transmitted pathogen, is an exquisitely adapted Gram-negative obligate intracellular bacterium. Intracellular Chlamydia trachomatis replicate in a specialized vacuole, termed inclusion, which shields the bacterium from antimicrobial immunity of the host cells and acts as a signalling interface. Previously it was shown that members of the interferon induced guanylate binding protein (mGBP) family, in particular murine GBP1 and mGBP2, were found to accumulate at the bacterial inclusions, similar to previously published recruitment of GBPs to the parasitophorous vacuole of Toxoplasma gondii. Here, we provide a wide comparison of mGBPs roles within the host cell in the context of Chlamydia and Toxoplasma infection. By confocal microscopy on fixed and living infected cells we show localization of mGBP3, mGBP6, mGBP7, mGBP9, and mGBP10, in addition to mGBP1 and mGBP2, at chlamydia inclusions. In time lapse videos using GFP expressing Chlamydia we show rapid and transient dynamics of mGBP9 accumulation onto chlamydia inclusions. Taken together this study reveals a broad activation of mGBP recruitment towards Chlamydia trachomatis inclusions after infection and provides evidence for time limited action of mGBP9 at the chlamydia inclusion.
Guanylate binding proteins (GBPs) are large interferon-inducible GTPases, executing essential host defense activities against Toxoplasma gondii, an invasive intracellular apicomplexan protozoan parasite of global importance. T. gondii establishes a parasitophorous vacuole (PV) which shields the parasite from the host's intracellular defense mechanisms. Murine GBPs (mGBPs) recognize T. gondii PVs and assemble into supramolecular mGBP homo- and heterocomplexes that are required for the disruption of the membrane of PVs eventually resulting in the cell-autonomous immune control of vacuole-resident pathogens. We have previously shown that mGBP2 plays an important role in T. gondii immune control. Here, in order to unravel mGBP2 functions, we report Galectin-9 (Gal9) and cytoskeleton-associated protein 4 (Ckap4) as critical mGBP2 interaction partners engaged for immunity to T. gondii. Interestingly, Gal9 and Ckap4 also accumulate and colocalize with mGBP2 at the T. gondii PV. Furthermore, we could prove the requirement of Gal9 and Ckap4 for growth control of T. gondii by CRISPR/Cas9 mediated gene editing. These discoveries clearly indicate that mGBP2 engages Gal9 and Ckap4 and that Gal9 and Ckap4 are critical factors for the mGBP2 coordinated cell autonomous host defense mechanism against T. gondii.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.