Modulation of innate immunity by Toxoplasma gondii virulence effectors

Abstract

Toxoplasma gondii is a common parasite of animals and humans and can cause serious opportunistic infections. However, the majority of infections are asymptomatic, possibly because the organism has co-evolved with its many vertebrate hosts and has developed multiple strategies to persist asymptomatically for the lifetime of the host. Over the past two decades, infection studies in the mouse, combined with forward-genetics approaches aimed at unravelling the molecular basis of infection, have revealed that T. gondii virulence is mediated, in part, by secretion of effector proteins into the host cell during invasion. Here, we review recent advances that illustrate how these virulence factors disarm innate immunity and promote survival of the parasite.

Figure 1. The complex life cycle of T. gondii

Cats are the definitive host where sexual replication takes place. Following replication within enterocytes of the gut (a process known as merogony), male and female gametes are formed within the host cell, as described previously . Fusion of gametes leads to the formation of diploid oocysts that are shed in cat faeces and undergo meiosis in the environment to yield eight haploid progeny. Oocysts are capable of surviving in the environment for long periods of time and can contaminate food and water, providing a route of infection for intermediate hosts. In the intermediate host (shown here as rodents) asexual replication occurs. Acute infection is characterized by fast replicating tachyzoites that disseminate throughout the body. Differentiation to slow-growing bradyzoites within tissue cysts leads to long-term chronic infection. Ingestion of tissue cysts via omnivorous or carnivorous feeding can lead to transmission to other intermediate hosts or to cats, which re-initiates the sexual phase of the life cycle. Many animals serve as intermediate hosts, including farm animals. Humans become infected by eating undercooked meat containing tissue cysts or by the ingestion of oocysts in contaminated water ^,. Although most infections are mild, toxoplasmosis can cause serious symptoms in the brain and other organs (as indicated) in immunocompromised patients, as well as in the developing foetus following congenital infection.

Figure 2. Innate immune responses to Toxoplasma gondii during infection

a) Early in infection, the first cells to respond are dendritic cells (DCs) and monocytes/macrophages. Interaction of T. gondii profilin with TLR11 on DCs is important for the production of IL-12. In addition to stimulating IL-12 production, macrophages also induce TNF-α, a cofactor in antimicrobial activity, in response to the detection of GPI anchored proteins via TLR2 and TLR4. b) The immune response results in the production of IFN-γ from NK cells through the innate response and eventually from CD4⁺ and CD8⁺ T cells as the adaptive response ensues. IL-10 and IL-27 are key to modulating these pathways and prevent the overproduction of T_H1 cytokines. c) Production of IFN-γ during the innate and adaptive phases is responsible for activating cells to control parasite infection. IFN-γ propagates a signal through a surface receptor (IFN-γR) to activate STAT1, a nuclear transcription factor that controls the expression of many genes. In response to STAT1 activity, nitric oxide (NO) and reactive oxygen species (ROS) are upregulated in monocytes/macrophages, both of which contribute to the control of intracellular parasites. Both haematopoetic and nonhaematopoetic cells also upregulate two families of defence proteins called immunity related GTPases (IRGs) and guanylate binding proteins (GBPs), which are recruited to pathogen-containing vacuoles and are involved in parasite clearance. The function of IRGs and GBPs depends on the autophagy protein Atg5.

Figure 3. Structure and function of ROP2 family members

a) Model of the kinase domain of ROP18 based on homology to the X-ray crystal structure of the pseudokinase ROP8 (pdb 3BYV). The two N-terminal helices (green) are important for stabilization of the kinase domain (red) and are also involved in auto-activation of ROP18. The N- and C-lobes of the kinase domain are labelled, while the N-terminal extension involved in membrane tethering, is absent from the structure. Provided by Ray Hui, Structural Genomics Consortium, Toronoto. b) Domain structure of ROP2 family members showing the signal peptide and prodomain, both of which are processed prior to secretion (arrows). The N-terminal half of the mature protein (green) contains a series of three amphipathic alpha helical regions with low sequence complexity (grey boxes) that mediate membrane targeting. The C-terminal region contains a S/T kinase domain (red). Localization of full-length ROP18 on the membrane of the parasite containing vacuole is mediated by the N-terminal amphipathic region (green). Deletion of this region disrupts vacuole association of the kinase domain (red), as shown for the truncated version of ROP18.

Figure 4. The role of T. gondii virulence factors in modulating immune signalling in the host

ROP16_I/III (pink) phosphorylates STAT3 and STAT6 resulting in prolonged activation of these two transcription factors and subsequent upregulation of IL-4 while antagonizing induction of IL-12. The dense granule protein GRA15_II (green) activates TRAF6, which in turn activates IKK, leading to phosphorylation of IκB and the release of NFκB following proteasomal degradation. NFκB migrates to the nucleus and drives the production of IL-12. ROP18_I (red) is localized on the cytoplasmic side of the parasite containing vacuole (PV). ROP18_I has been shown to phosphorylate immunity related GTPases (IRGs), thus blocking their accumulation on the vacuole and protecting the parasite from destruction. ROP18_I also phosphorylates the host transcription factor ATF6β, which is involved in the unfolded protein response and may also be important for efficient antigen presentation by DCs. Genetic evidence suggests that ROP5_I/III (purple), a pseudokinase important for virulence, regulates the functions of ROP18_I. Allele types are shown in subscript (i.e. ROP_I denotes the allele in type I strains).