Genome wide CRISPR screen of T. gondii genes important under interferon gamma pressure in human cells

In both mice and humans, Type II interferon-gamma is crucial for regulation of Toxoplasma gondii (T. gondii) infection, whether in the acute or chronic phases. To thwart this defense, T. gondii secretes protein effectors hindering the host's immune response. T. gondii relies on the MYR translocon complex to deploy soluble dense granule effectors (GRAs) into the host cell cytosol or nucleus. Recent genome-wide loss-of-function screens to identify Toxoplasma genes important for parasite fitness in IFN gamma primed primary human fibroblasts identified MYR translocon components as crucial for parasite resistance to IFN gamma, but notably, they did not determine specific MYR dependent GRA proteins that block IFN gamma signaling. This suggests a likely combinatorial function of multiple effectors. Our study reveals that T. gondii depends on the MYR translocon complex to prevent host cell death and parasite premature egress in human cells stimulated with IFN gamma post-infection, a unique phenotype observed in various human cell lines but not in murine cells. Intriguingly, inhibiting parasite egress did not prevent host cell death. Genome-wide loss-of-function screens uncovered TgIST, GRA16, GRA24, and GRA28 as effectors necessary for a complete block of IFN gamma response. GRA24 and GRA28 directly influence IFN gamma driven transcription, with GRA24's action dependent on its interaction with p38 MAPK, while GRA28 disrupts histone acetyltransferase activity of CBP/p300. Given the intricate nature of the immune response to T. gondii, it appears that the parasite has evolved equally intricate mechanisms for complete subversion of IFN gamma signaling. These mechanisms extend beyond direct interference with the JAK/STAT1 pathway, encompassing manipulation of other signaling pathways as well.