Toxoplasma  GRA15 and GRA24 are important activators of the host innate immune response in the absence of TLR11

Abstract

The murine innate immune response against Toxoplasma gondii is predominated by the interaction of TLR11/12 with Toxoplasma profilin. However, mice lacking Tlr11 or humans, who do not have functional TLR11 or TLR12, still elicit a strong innate immune response upon Toxoplasma infection. The parasite factors that determine this immune response are largely unknown. Herein, we investigated two dense granule proteins (GRAs) secreted by Toxoplasma, GRA15 and GRA24, for their role in stimulating the innate immune response in Tlr11 -/mice and in human cells, which naturally lack TLR11/TLR12. Our results show that GRA15 and GRA24 synergistically shape the early immune response and parasite virulence in Tlr11 -/mice, with GRA15 as the predominant effector. Nevertheless, acute virulence in Tlr11 -/mice is still dominated by allelic combinations of ROP18 and ROP5, which are effectors that determine evasion of the immunity-related GTPases. In human macrophages, GRA15 and GRA24 play a major role in the induction of IL12, IL18 and IL1β secretion. We further show that GRA15/GRA24-mediated IL12, IL18 and IL1β secretion activates IFNγ secretion by peripheral blood mononuclear cells (PBMCs), which controls Toxoplasma proliferation. Taken together, our study demonstrates the important role of GRA15 and GRA24 in activating the innate immune response in hosts lacking TLR11.

Influence of the Host and Parasite Strain on the Immune Response During Toxoplasma Infection

“…Indeed, mice deficient in components of the inflammasome sensors NLRP1, NLRP3, ASC, caspase 1 and caspase 11, or downstream cytokine IL18 or its receptor (IL18R) all showed increased susceptibility to Toxoplasma infection (Gorfu et al, 2014 ). In line with this, in human PBMCs, both IL18 and IL1β secreted through the activation of the NLRP3 inflammasome pathway are responsible for the secretion of IFNγ and IL12 (Mukhopadhyay et al, 2020a ). Furthermore, apart from murine models, inflammasome activation is also important to restrict parasite growth in human and rat macrophages (Sergent et al, 2005 ; Witola et al, 2011 ; Cavaillès et al, 2014 ; Cirelli et al, 2014 ; Gov et al, 2017 ; Fisch et al, 2019a ).…”

“…In mammals or mice lacking TLR11, activation of the immune system upon Toxoplasma infection is dependent on cytoplasmic sensors such as NLRP1/3, the adaptor protein apoptosis associated speck-like protein containing a CARD (ASC) and the caspases 1 and 4, all of which lead to the secretion of IL1β and IL18, a process known as inflammasome activation (Witola et al, 2011 ; Gov et al, 2013 , 2017 ; Mukhopadhyay et al, 2020a ). In mice lacking TLR11, and therefore DC-derived IL12, a large amount of IFNγ is secreted by neutrophils and CD8 T cells via pro-inflammatory monocyte and macrophage dependent inflammasome driven secretion of IL1β and IL18 (Sturge et al, 2013 ; López-Yglesias et al, 2019 ) ( Figure 1 ).…”

“…Although DC-derived IL12 is important for secretion of IFNγ in vivo , it alone is not sufficient to protect the host, as Myd88 −/− mice that were administered recombinant IL12 post-infection did not survive infection (Hou et al, 2011 ). The pro-inflammatory cytokines IL18 and IL1β secreted by monocytes and macrophages upon inflammasome activation also enhance IFNγ production and, together with IL12, can thereby contribute to host resistance against Toxoplasma (Hunter et al, 1995 ; Cai et al, 2000 ; Gov et al, 2013 , 2017 ; Mukhopadhyay et al, 2020a ). Furthermore, while mice lacking either MyD88 or IL12p40 succumb rapidly to infection, mice that lack Myd88 specifically in DCs or macrophage lineages do not, indicating that there are other factors and cytokines that regulate IL12 and IFNγ secretion (Scanga et al, 2002 ; Hou et al, 2011 ).…”

“…Quantitative trait locus (QTL) analysis of host gene expression levels after infection with F1 progenies derived from IIxIII crosses led to the discovery that ROP16 (encoding a secreted kinase) (Saeij et al, 2007 ) and GRA15 (Rosowski et al, 2011 ) determine most Toxoplasma strain differences in the modulation of the inflammatory response in mice. Through the NFκB pathway, type II GRA15 (GRA15 II ) induces the production of inflammatory cytokines such as IL12 and IL1β, and elicits classically activated macrophages (M1) (Jensen et al, 2011 ; Gov et al, 2013 ; Mukhopadhyay et al, 2020a ). These M1 macrophages are generally good at killing intracellular pathogens but their secretion of pro-inflammatory cytokines can cause pathology through an excessive activation of NK, Th1, and Th17 cells, and by inducing pro-apoptotic pathways (Murray et al, 2014 ).…”

“…All three major clonal populations of Toxoplasma express GRA24, however, it is in type II strains where GRA24, together with GRA15 and in the absence of active ROP16, can induce a heightened inflammatory immune response. GRA15 and GRA24-induced IL12 secretion by macrophages is achieved through the activation of the cREL NFκB pathway (Mukhopadhyay et al, 2020a ). GRA15 and GRA24 together induce the secretion of IL18 and IL12, resulting in higher IFNγ secretion and lower parasite burden, while infection with parasites lacking these two effectors results in significantly lower IFNγ secretion and concomitantly higher peritoneal parasite burden in comparison to wild-type parasites (Mercer et al, 2020 ; Mukhopadhyay et al, 2020a ).…”