Immediately prior to invasion Toxoplasma gondii tachyzoites release a large number of micronemal proteins (TgMICs) that participate in host cell attachment and penetration. The TgMIC4-MIC1-MIC6 complex was the first to be identified in T. gondii and has been recently shown to be critical in invasion. This study establishes that the N-terminal throm-bospondin type I repeat-like domains (TSR1-like) from TgMIC1 function as an independent adhesin as well as promoting association with TgMIC4. Using the newly solved three-dimensional structure of the C-terminal domain of TgMIC1 we have identified a novel Galectin-like fold that does not possess carbohydrate binding properties and redefines the architecture of TgMIC1. Instead, the TgMIC1 Galectin-like domain interacts and stabilizes TgMIC6, which provides the basis for a highly specific quality control mechanism for successful exit from the early secretory compartments and for subsequent trafficking of the complex to the micronemes.Toxoplasma gondii is a protozoan parasite of the phylum Apicomplexa, which infects virtually all warm-blooded animals and invades almost any cell type. Host cell invasion by this obligate intracellular parasite is an active process initiated by the formation of a tight association/junction with the host cell plasma membrane and leading to the creation of a parasitophorous vacuole. Contact with the host cell results in an increase in parasite intracellular calcium ions, which trigger apical organelles called micronemes to discharge their contents (1). Several micronemal proteins act as ligands for host cell receptors (2), while TgMIC2 and other transmembrane proteins establish a connection with the parasite actinomyosin system via their cytoplasmic tail (3), thus providing the motive force for penetration (4). It is becoming increasingly apparent that many microneme proteins are found in stable adhesive complexes, which are formed in the endoplasmic reticulum, and normally comprise an escorter protein, which is responsible for correct micronemal targeting, and one or more soluble effector proteins. The first such complex to be discovered in T. gondii was TgMIC4-MIC1-MIC6, in which TgMIC6 fulfils the role of the escorter protein, whereas TgMIC1 and TgMIC4 function as adhesins (5). Although TgMIC4-MIC1-MIC6 and the recently identified micronemal complex, TgMIC3-MIC8 (5, 6), are individually dispensable, the generation of double knock-outs for TgMIC1 and TgMIC3 renders the parasites avirulent in vivo, demonstrating functional synergy between these complexes (7). Deletion of the mic1 gene in T. gondii also confirmed the specific and critical role played by TgMIC1 in host cell attachment and invasion in vitro.Micronemal proteins have a modular structure with common themes in domain organization, for example many possess thrombospondin type-1 repeat domains (TSR1), 4 apple (or PAN) domains, and epidermal growth factor-like (EGF) domains (8). A schematic representation of the organization within the TgMIC4-MIC1-MIC6 complex is depicted in Fig. 1. Tg...

Interleukin-25 (IL-25) and group 2 innate lymphoid cells (ILC2s) defend the host against intestinal helminth infection and are associated with inappropriate allergic reactions. IL-33–activated ILC2s were previously found to augment protective tissue-specific pancreatic cancer immunity. Here, we showed that intestinal IL-25–activated ILC2s created an innate cancer-permissive microenvironment. Colorectal cancer (CRC) patients with higher tumor IL25 expression had reduced survival and increased IL-25R–expressing tumor-resident ILC2s and myeloid-derived suppressor cells (MDSCs) associated with impaired antitumor responses. Ablation of IL-25 signaling reduced tumors, virtually doubling life expectancy in an Apc mutation–driven model of spontaneous intestinal tumorigenesis. Mechanistically, IL-25 promoted intratumoral ILC2s, which sustained tumor-infiltrating MDSCs to suppress antitumor immunity. Therapeutic antibody-mediated blockade of IL-25 signaling decreased intratumoral ILC2s, MDSCs, and adenoma/adenocarcinoma while increasing antitumor adaptive T cell and interferon-γ (IFN-γ)–mediated immunity. Thus, the roles of innate epithelium-derived cytokines IL-25 and IL-33 as well as ILC2s in cancer cannot be generalized. The protumoral nature of the IL-25–ILC2 axis in CRC highlights this pathway as a potential therapeutic target against CRC.

Background: TgMIC4 is an important microneme effector protein from Toxoplasma gondii. Results: The structure of TgMIC4 together with carbohydrate microarray analyses reveal a broad specificity for galactoseterminating sequences. Conclusion: Lectin activity within the fifth apple domain of TgMIC4 is reminiscent of the mammalian galectin family. Significance: TgMIC4 may contribute to parasite dissemination within the host or down-regulation of the immune response.

The obligate intracellular parasite Toxoplasma gondii, a member of the phylum Apicomplexa that includes Plasmodium spp., is one of the most widespread parasites and the causative agent of toxoplasmosis. Adhesive complexes composed of microneme proteins (MICs) are secreted onto the parasite surface from intracellular stores and fulfil crucial roles in host-cell recognition, attachment and penetration. Here, we report the high-resolution solution structure of a complex between two crucial MICs, TgMIC6 and TgMIC1. Furthermore, we identify two analogous interaction sites within separate epidermal growth factor-like (EGF) domains of TgMIC6-EGF2 and EGF3-and confirm that both interactions are functional for the recognition of host cell receptor in the parasite, using immunofluorescence and invasion assays. The nature of this new mode of recognition of the EGF domain and its abundance in apicomplexan surface proteins suggest a more generalized means of constructing functional assemblies by using EGF domains with highly specific receptor-binding properties.

Microneme protein 4 is involved in cell binding by the important parasite Toxoplasma gondii. We present here the backbone and side-chain assignments of the first two apple domains together with a new graphical aid for their assignment using NMRView.