Biology subjects

Saeij, J. P.

Publications and source records attributed to Saeij, J. P..

5 recordsLinked to original sources

bioRxiv2024

Hypermigration of macrophages through the concerted action of GRA effectors on NF-κB/p38 signaling and host chromatin accessibility potentiates Toxoplasma dissemination

Mononuclear phagocytes facilitate the dissemination of the obligate intracellular parasite Toxoplasma gondii. Here, we report how a set of secreted parasite effector proteins from dense granule organelles (GRA) orchestrates dendritic cell-like chemotactic and pro-inflammatory activation of parasitized macrophages. These effects enabled efficient dissemination of the type II T. gondii lineage, a highly prevalent genotype in humans. We identify novel functions for effectors GRA15 and GRA24 in promoting CCR7-mediated macrophage chemotaxis by acting on NF-{kappa}B and p38 MAPK signaling pathways, respectively, with contributions of GRA16/18 and counter-regulation by effector TEEGR. Further, GRA28 boosted chromatin accessibility and GRA15/24/NF-{kappa}B-dependent transcription at the Ccr7 gene locus in primary macrophages. In vivo, adoptively transferred macrophages infected with wild-type T. gondii outcompeted macrophages infected with a GRA15/24 double mutant in migrating to secondary organs in mice. The data show that T. gondii, rather than being passively shuttled, actively promotes its dissemination by inducing a finely regulated pro-migratory state in parasitized human and murine phagocytes via co-operating polymorphic GRA effectors. ImportanceIntracellular pathogens can hijack cellular functions of infected host cells to their advantage, for example, for intracellular survival and for dissemination. However, how microbes orchestrate the hijacking of complex cellular processes, such as host cell migration, remains poorly understood. As such, the common parasite Toxoplasma gondii actively invades immune cells of humans and other vertebrates and modifies their migratory properties. Here, we show that the concerted action of a number of secreted effector proteins from the parasite, principally GRA15 and GRA24, act on host cell signaling pathways to activate chemotaxis. Further, the protein effector GRA28 selectively acted on chromatin accessibility in the host cell nucleus to selectively boost host gene expression. The joint activities of effectors culminated in pro-migratory signaling within the infected phagocyte. We provide a molecular framework delineating how T. gondii can orchestrate a complex biological phenotype, such as the migratory activation of phagocytes to boost dissemination.

microbiology↗

bioRxiv2023

Toxoplasma Membrane Inositol Phospholipid Binding Protein TgREMIND Is Essential for Secretory Organelle Function and Host Infection

Apicomplexan parasites have specialized secretory organelles called rhoptries, micronemes, and dense granules that are essential for host infection. Here, we show that TgREMIND, a Toxoplasma gondii protein containing a membrane phospholipid interacting domain, is required for the biogenesis of rhoptries and dense granules. TgREMIND contains a Fes/CIP4 homology-Bin/Amphiphysin/Rvs (F-BAR) domain at the N-terminus, known to promote cell membrane bending, and a novel uncharacterized domain that we named REMIND for regulator of membrane interacting domain at the C-terminus. TgREMIND binds to PIP2 lipid species and both F-BAR and REMIND domains are necessary to ensure proper biological activities in vitro and in cellulo. Conditional depletion of TgREMIND results in the absence of dense granules and abnormal transparent rhoptries, leading to a severe inhibition of parasite motility, host invasion, and dissemination. Thus, our study demonstrates that TgREMIND is essential for the proper functioning of key secretory organelles required for successful infection by Toxoplasma.

cell biology↗

bioRxiv2020

Toxoplasma GRA15 limits parasite growth in IFNγ-activated fibroblasts through TRAF ubiquitin ligases

The protozoan parasite Toxoplasma gondii lives inside a vacuole in the host cytoplasm where it is protected from host cytoplasmic innate immune responses. However, IFN{gamma}-dependent cell-autonomous immunity can destroy the vacuole and the parasite inside. Toxoplasma strain differences in susceptibility to human IFN{gamma} exist but the Toxoplasma effector(s) that determine these differences are unknown. We show that in human primary fibroblasts, the polymorphic Toxoplasma secreted effector GRA15 mediates the recruitment of ubiquitin ligases, including TRAF2 and TRAF6, to the vacuole membrane, which enhances recruitment of ubiquitin receptors (p62/NDP52) and ubiquitin-like molecules (LC3B, GABARAP). This ultimately leads to lysosomal degradation of the vacuole. In murine fibroblasts, GRA15-mediated TRAF6 recruitment mediates the recruitment of immunity-related GTPases and destruction of the vacuole. Thus, we have identified how the Toxoplasma effector GRA15 affects cell-autonomous immunity in human and murine cells.

immunology↗

bioRxiv2020

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

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 IL1{beta} secretion. We further show that GRA15/GRA24-mediated IL12, IL18 and IL1{beta} secretion activates IFN{gamma} 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. ImportanceIn mice, the early immune response against Toxoplasma is dominated by TLR11-mediated release of IL-12, which subsequently induces protective IFN{gamma}. Here we show that in Tlr11-/- mice and in human cells, which do not have TLR11, the Toxoplasma GRA15 and GRA24 effectors play an important role in induction of IL12, IL18 and IL1{beta}, and thus in the subsequent protective IFN{gamma} secretion.

immunology↗

bioRxiv2018

Lewis rat NLRP1 inflammasome activation is mediated by three Toxoplasma gondii dense granule proteins

The Lewis rat is the only known warm-blooded animal that has sterile immunity to Toxoplasma. Upon invasion of Lewis rat macrophages Toxoplasma rapidly activates the nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain containing 1 (NLRP1) inflammasome resulting in interleukin (IL)-1{beta} secretion and a form of cell death known as pyroptosis, which prevents Toxoplasma replication. Using a chemical mutagenesis screen we identified Toxoplasma mutants that no longer induced pyroptosis. Whole genome sequencing led to the identification of three Toxoplasma parasitophorous vacuole-localized dense granule proteins, GRA35, GRA42 and GRA43 that are individually required for inflammasome activation in Lewis rat macrophages. Macrophage infection with {Delta}gra35, {Delta}gra42, and {Delta}gra43 parasites leads to greatly reduced cell death and reduced IL-1{beta} secretion. Lewis rat macrophage infected with parasites containing single, double or triple deletion of these GRAs showed similar levels of cell viability suggesting the three GRAs function in the same pathway that activates the inflammasome. Deletion of GRA42 and GRA43 resulted in GRA35, and other GRAs, being retained inside the parasitophorous vacuole instead of being localized to the parasitophorous vacuole membrane. Toxoplasma deficient in GRA35, GRA42 or GRA43 do not establish chronic infection in Lewis rats, but have reduced cyst number in parasite-susceptible F344 rats, in which Toxoplasma does not activate the NLRP1 inflammasome, revealing these GRAs determine parasite in vivo fitness independent of their role in inflammasome activation. Overall, our data suggest that Toxoplasma dense granule proteins that localize to the parasitophorous vacuole membrane are novel mediators of host NLRP1 inflammasome activation.\n\nImportanceInflammasomes are a major component of the innate immune system and responsible for detecting various microbial and environmental danger signals. The Lewis rat has sterile immunity to Toxoplasma because upon invasion of Lewis rat macrophages the parasite rapidly activates the NLRP1 inflammasome resulting in cell death and parasite elimination. The work reported here identified that Toxoplasma GRA35, GRA42 and GRA43 are required for activation of the Lewis rat NLRP1 inflammasome. GRA42 and GRA43 mediate the correct localization of other GRAs, including GRA35, to the parasitophorous vacuole membrane. In addition to their role in inflammasome activation, these three GRAs are also important for parasite in vivo fitness in a Toxoplasma-susceptible rat strain. Thus, these results give new insight into NLRP1 inflammasome activation by Toxoplasma effectors and identified three GRAs that are required for pathogenesis of the parasite.

microbiology↗