Authors

A Gossner¹; A Raper²; M Hassan³;  ¹ University of Edinburgh, UK;  ² The Roslin Institute, Universilty of Edinburgh, UK;  ³ Roslin Institute, University of Edinburgh, UK

Discussion

Abstract A third of the world’s human population is thought to be chronically infected with Toxoplasma gondii despite the robust immune response to the parasite in immune competent individuals. As with many intracellular pathogens interferon gamma (IFN) production has been shown to be central to the immune response to Toxoplasma and control of infection of this protozoan parasite, which is able to infect almost all warmblooded vertebrates. Murine studies of innate immunity to Toxoplasma have identified pathways involved in the IFN response to parasite infection. Despite this the response in humans is less defined and there are fundamental differences between the human and murine innate immune systems, such as TLR11 and TLR12, both involved in resistance to the parasite in mice are absent in humans. Despite these differences IFN has a major role in the induction of cell-autonomous responses by the human innate immune system, through activation of a large transcriptional sequence of events resulting in the heightened expression of interferon stimulated genes (ISGs). Among the many ISGs expressed by IFN stimulated cells, GTPases such as guanylatebinding proteins (GBPs) and immunity-related GTPases (IRGs) have been shown to be important in the immune response to Toxoplasma. While the role of IFN, a type II IFN, has been extensively studied and shown to be important in immunity against Toxoplasma, other IFNs are less well studied. IFNβ, a type I IFN, has been shown to restrict Toxoplasma growth in both mouse and human macrophages. Mice lacking the functional Interferon alpha and beta receptor subunit 1 (IFNAR1), a type I IFN receptor, have increased susceptibility to Toxoplasma infection. While there is activation of overlapping pathways by type I and II IFNs there are also specific type I and II ISGs, resulting in the activation of different signalling pathways. A potential role for type I IFNs in response to the parasite was investigated using a type I IFN inducible ISGKnockout library. Several genes restricting Toxoplasma growth in type I IFN stimulated THP-1 cells, a human monocytic cell line, were identified. These IFN inducible genes included sorting nexin-5 (SNX5), coagulation factor II receptor-like 2 (F2RL2), myc-associated factor X (MAX) and small RNA Binding Exonuclease Protection Factor La (SSB), all previously unknown restrictors of Toxoplasma growth. To characterise their mode of action we generated CRISPR knockout THP1 cell clones for each gene. These THP1 KO cells will be used to functionally characterise how the selected ISGs restrict Toxoplasma growth in THP-1 cells in a type I IFN-induced response.