CRISPR Screens Identify Toxoplasma Genes That Determine Parasite Fitness in Interferon Gamma-Stimulated Human Cells

mBio, Год журнала: 2023, Номер 14(2)

Опубликована: Март 14, 2023

Toxoplasma infection causes serious complications in immunocompromised individuals and the developing fetus. During infection, certain immune cells release a protein called interferon gamma that activates to destroy parasite or inhibit its growth.

Язык: Английский

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The role of IFN-γ-mediated host immune responses in monitoring and the elimination of Toxoplasma gondii infection

International Immunology, Год журнала: 2024, Номер 36(5), С. 199 - 210

Опубликована: Янв. 3, 2024

Toxoplasma gondii is a pathogenic protozoan parasite of the Apicomplexa family that affects approximately 30% world's population. Symptoms are usually mild in immunocompetent hosts, but it can pose significant health risks to immunosuppressed patients and pregnant women. Current treatment options limited, new therapies vaccines needed. The innate immune system first recognize T. infection activates pro-inflammatory cytokines chemokines promote acquired immunity. IL-12/IFN-γ axis particularly important, when this pathway inhibited, becomes uncontrolled lethal. In mice, receptors such as Toll-like receptor 11 (TLR11), TLR12, chemokine involved recognition modulation responses. humans, where TLR11 TLR12 absent, other mechanisms have been reported sensing infection. Immune cells activated response produce interleukin (IL)-12, which stimulates proliferation natural killer T promotes production interferon (IFN)-γ. Several IFN-γ-induced anti-T. defense inhibit growth. These include nitric oxide (NO) production, indoleamine 2,3-dioxygenase, destruction parasitophorous vacuoles by IFN-γ-inducible immunity related GTPase groups (IRGs GBPs). Recent studies focusing on diversity IRGs rodents effector molecules suggest host pathogen evasion co-evolved. Furthermore, has suggested cysts not simply dormant during chronic This review summarizes recent findings innate, adaptive, cell-autonomous

Язык: Английский

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A CRISPR view on genetic screens in Toxoplasma gondii

Current Opinion in Microbiology, Год журнала: 2025, Номер 83, С. 102577 - 102577

Опубликована: Янв. 8, 2025

Genome editing technologies, such as CRISPR-Cas9, have revolutionised the study of genes in a variety organisms, including unicellular parasites. Today, CRISPR-Cas9 technology is vastly applied high-throughput screens to investigate interactions between Apicomplexan parasite Toxoplasma gondii and its hosts. In vitro vivo T. performed naive restrictive conditions led discovery essential fitness-conferring genes, well factors important for virulence dissemination. Recent studies adapted screening based on phenotypes unrelated survival. These advances were achieved by using conditional systems coupled with imaging, single-cell RNA sequencing phenotypic selection. Here, we review state-of-the-art technologies focus gondii, highlighting strengths, current limitations future avenues development, application other species.

Язык: Английский

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High-throughput identification of Toxoplasma gondii effector proteins that target host cell transcription

Cell Host & Microbe, Год журнала: 2023, Номер 31(10), С. 1748 - 1762.e8

Опубликована: Окт. 1, 2023

Intracellular pathogens and other endosymbionts reprogram host cell transcription to suppress immune responses recalibrate biosynthetic pathways. This reprogramming is critical in determining the outcome of infection or colonization. We combine pooled CRISPR knockout screening with dual host-microbe single-cell RNA sequencing, a method we term perturb-seq, identify molecular mediators these transcriptional interactions. Applying perturb-seq intracellular pathogen Toxoplasma gondii, are able previously uncharacterized effector proteins directly infer their function from transcriptomic data. show that TgGRA59 contributes export parasite into an effector, TgSOS1, necessary for sustained STAT6 signaling thereby evasion persistence. Together, this work demonstrates tool can be broadly adapted interrogate interactions reveal mechanisms evasion.

Язык: Английский

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Genome-wide CRISPR screen identifies genes synthetically lethal with GRA17, a nutrient channel encoding gene in Toxoplasma

PLoS Pathogens, Год журнала: 2023, Номер 19(7), С. e1011543 - e1011543

Опубликована: Июль 27, 2023

Toxoplasma gondii is a parasite that replicates within specialized compartment called the parasitophorous vacuole (PV), which surrounded by PV membrane (PVM). To obtain essential nutrients, must transport molecules across PVM, process mediated secreted proteins GRA17 and GRA23. These form pores in PVM through small can diffuse out of PV. GRA23 are synthetically lethal, suggesting at least one pore type for survival. In ‘nutrient sensitized’ Δ gra17 strain it likely other genes become essential, because they mediate nutrient acquisition from host or involved trafficking to PVM. identify these genes, genome-wide loss-of-function screen was performed wild-type parasites, identified multiple were sick/lethal with . Several correct localization GRAs, including GRA17/GRA23, One top hits, GRA72, predicted on its deletion led formation enlarged “bubble vacuoles” reduced molecule permeability, similar what previously observed parasites. Furthermore, gra72 parasites had vitro growth virulence mice. findings suggest absence GRA17, play role proper (and GRAs) determine host-derived

Язык: Английский

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IFNs in host defence and parasite immune evasion during Toxoplasma gondii infections

Frontiers in Immunology, Год журнала: 2024, Номер 15

Опубликована: Фев. 7, 2024

Interferons (IFNs) are a family of cytokines with diverse functions in host resistance to pathogens and immune regulation. Type II IFN, i.e. IFN-γ, is widely recognized as major mediator intracellular pathogens, including the protozoan Toxoplasma gondii . More recently, IFN-α/β, type I IFNs, IFN-λ (type III IFN) have been identified also play important roles during T. infections. This parasite widespread pathogen humans animals, it model organism study cell-mediated responses infection. Its success depends, among other factors, on ability counteract IFN system, both at level IFN-mediated gene expression IFN-regulated effector molecules. Here, review recent advances our understanding molecular mechanisms underlying regulation discuss those that has evolved efficiently evade immunity. Knowledge these fascinating host-parasite interactions their signalling machineries crucial for deeper pathogenesis toxoplasmosis, might identify potential targets parasite-directed or host-directed supportive therapies combat more effectively.

Язык: Английский

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Trx4, a novel thioredoxin protein, is important for Toxoplasma gondii fitness

Parasites & Vectors, Год журнала: 2024, Номер 17(1)

Опубликована: Апрель 4, 2024

Abstract Background To successfully replicate within the host cell, Toxoplasma gondii employs several mechanisms to overcome cell defenses and mitigate harmful effects of free radicals resulting from its own metabolic processes using effectors such as thioredoxin proteins. In this study, we characterize location functions a newly identified in T. , which was named Trx4. Methods We characterized functional role Trx4 Type I RH II Pru strains by gene knockout studied subcellular localization endogenous protein HA tagging CRISPR-Cas9 editing. The enzyme-catalyzed proximity labeling technique, TurboID system, employed identify proteins Results dense granule predominantly expressed parasitophorous vacuole (PV) partially co-localized with GRA1 GRA5. Functional analysis showed that deletion trx4 markedly influenced parasite lytic cycle, impaired invasion capacity both strains. Mutation Trx domains strain revealed two were important for invasion. By utilizing system biotinylate Trx4, substantial number proteins, some are novel, others previously characterized, distributed granules. addition, uncovered three novel Intriguingly, did not affect these Finally, virulence assay demonstrated resulted significant attenuation reduction brain cyst loads mice. Conclusions plays an strain. Combining technique many PV-localized associated These findings suggest versatile mediating occur distinctive intracellular membrane-bound vacuolar compartment. Graphical

Язык: Английский

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CRISPR screens identify genes essential for in vivo virulence among proteins of hyperLOPIT-unassigned subcellular localization in Toxoplasma

mBio, Год журнала: 2024, Номер 15(9)

Опубликована: Июль 31, 2024

ABSTRACT The research field to identify and characterize genes essential for in vivo virulence Toxoplasma gondii has been dramatically advanced by a series of clustered regularly interspaced short palindromic repeats (CRISPR) screens. Although subcellular localizations thousands proteins were predicted the spatial proteomic method called hyperLOPIT, those more than 1,000 remained unassigned, their essentiality was also unknown. In this study, we generated two small-scale gRNA libraries targeting approximately 600 hyperLOPIT-unassigned performed CRISPR As result, identified several that previously unreported. We further characterized candidates, TgGTPase TgRimM, which are localized cytoplasm apicoplast, respectively. Both parasite widely conserved phylum Apicomplexa. Collectively, our current study provides resource estimating with unknown localizations. IMPORTANCE is protozoan causes severe infection immunocompromised patients or newborns. possesses 8,000 genes; however, not fully identified. apicomplexan parasites, including , developed unique organelles do exist other model organisms; thus, determining location important understanding functions. Here, used genetic screens enabled us investigate hundreds during mouse infection. screened many novel confer mice. Among top hits, virulence, Our findings will contribute how apicomplexans adapt host environment cause disease.

Язык: Английский

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GRA12 is a common virulence factor across Toxoplasma gondii strains and mouse subspecies

Nature Communications, Год журнала: 2025, Номер 16(1)

Опубликована: Апрель 16, 2025

Язык: Английский

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A potential cure for tumor‐associated immunosuppression by Toxoplasma gondii

Cancer Reports, Год журнала: 2023, Номер 7(2)

Опубликована: Дек. 18, 2023

Abstract Background Recently, immunotherapy has become very hopeful for cancer therapy. Cancer treatment through excellent specificity and less toxicity than conventional chemoradiotherapy. Pathogens have been used in a long time. The current study aims to evaluate the possibility of Toxoplasma gondii ( T. ) as probable cancers such melanoma, breast, ovarian, lung, pancreatic cancer. Recent findings Nonreplicating type I uracil auxotrophic mutants can stimulate immune responses against tumors by reverse immunosuppression at cellular level. be utilized research T helper 1 (Th1) cell immunity intracellular infections. Avirulent auxotroph vaccine change tumor's improve production cytokines, i.e., Interferon‐gamma (IFN‐γ) Interleukin‐12 (IL‐12) activate tumor‐related Cluster Differentiation 8 (CD8+) cells identify destroy cells. profilin protein, along with secreted proteins, found exhibit promising properties various cancers. These proteins are being studied their potential inhibit tumor growth enhance effectiveness therapies. Their unique mechanisms action make them valuable candidates targeted interventions ovarian cancer, breast lung treatments. Conclusion In summary, underscores significant harnessing , including its diverse array antigens, particularly avirulent form, groundbreaking approach immunotherapy.

Язык: Английский

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