Interferon-Inducible E3 Ligase RNF213 Facilitates Host-Protective Linear and K63-Linked Ubiquitylation of Toxoplasma gondii Parasitophorous Vacuoles

Hernandez, Dulcemaria; Walsh, Stephen; Sanchez, Luz Saavedra; Dickinson, Mary S; Coers, Jörn

doi:10.1128/mbio.01888-22

Cited by 23 publications

(33 citation statements)

References 79 publications

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“…Ubiquitination of the PVM is a marker for parasite clearance in IFNγ-activated human cells (Clough et al ., 2016; Yao et al ., 2021; Hernandez et al ., 2022; Matta et al ., 2022). We therefore assessed if deletion of GRA57, GRA70 or GRA71 leads to differences in ubiquitination of the PVM.…”

Section: Resultsmentioning

confidence: 99%

“…Recent work has found that RNF213, an IFNγ-induced host E3 ubiquitin ligase, mediates the majority of ubiquitin recruitment to both RH and PRU Toxoplasma vacuoles in HFFs (Hernandez et al ., 2022; Matta et al ., 2022); however, the target of this ligase is currently unknown. The observation that GRA57/GRA70/GRA71 deletion leads to reduced ubiquitination of the PV poses the intriguing question of whether the complex is required for RNF213 recruitment, is a direct target of ubiquitination, or is responsible for correct positioning of an RNF213 target at the PV.…”

Section: Discussionmentioning

confidence: 99%

“…Compared to type II & type III parasites, type I parasites are more resistant to ubiquitination and clearance in both HUVECs and HeLa cells. Recent evidence suggests a major function for the E3 ligase RNF213 in ubiquitination of the PVM and enhancing parasite clearance, in multiple human cell types and in a strain-independent manner (Hernandez et al ., 2022; Matta et al ., 2022). The importance of each E3 ligase and ubiquitin linkage therefore depends on the exact combination of host species, parasite strain and cell type studied, but nevertheless ubiquitination has emerged as a key process in the regulation of Toxoplasma clearance.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

A heterotrimeric complex ofToxoplasmaproteins promotes parasite survival in interferon gamma stimulated human cells

Lockyer

Torelli

Butterworth

et al. 2022

Preprint

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Toxoplasma gondii secretes protein effectors to subvert the human immune system sufficiently to establish a chronic infection. Relative to murine infections, little is known about which parasite effectors disarm human immune responses. Here we used targeted CRISPR screening to identify secreted protein effectors required for parasite survival in IFNγ-activated human cells. Independent screens were carried out using two Toxoplasma strains which differ in virulence in mice, leading to the identification of effectors required for survival in IFNγ-activated human cells. We identify the secreted protein GRA57 and two other proteins, GRA70 and GRA71, that together form a complex which enhances the ability of parasites to persist in IFNγ-activated human foreskin fibroblasts (HFFs). Components of the protein machinery required for export of Toxoplasma proteins into the host cell were also found to be important for parasite resistance to IFNγ in human cells, but these export components function independently of the identified protein complex. Host-mediated ubiquitination of the parasite vacuole has previously been associated with increased parasite clearance from human cells, but we find that vacuoles from GRA57, GRA70 and GRA71 knockout strains are surprisingly less ubiquitinated by the host cell. We hypothesise that deletion of this trimeric complex renders parasites hypersensitive to remaining ubiquitination, resulting in increased parasite clearance.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

A heterotrimeric complex ofToxoplasmaproteins promotes parasite survival in interferon gamma stimulated human cells

Lockyer

Torelli

Butterworth

et al. 2022

Preprint

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show abstract

“…Quite a few host ubiquitin ligases, such as leucine-rich repeat and sterile α-motif containing 1 (LRSAM1), Parkin, Ring finger protein 166 (RNF166), RNF213, Ariadne RING-BetweenRING-RING (RBR) E3-ubiquitin protein ligase 1 (ARIH1), SMAD-specific E3-ubiquitin protein ligase 1 (Smurf1), and Skip-Cullin-F-box protein 2 containing complex (SCF FBXO2 ) are reported to decorate pathogen or pathogen-containing vacuoles with a variety of ubiquitin chain topologies (8,9,(11)(12)(13)(14)(15)(16)(17)(18). Notably, in particular, LRSAM1 through auto-ubiquitination possibly generates a robust ubiquitin signal around the bacteria to recruit the autophagic machinery (19,20).…”

Section: Introductionmentioning

confidence: 99%

An innate pathogen sensing strategy involving ubiquitination of bacterial surface proteins

et al. 2023

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Sensing of pathogens by ubiquitination is a critical arm of cellular immunity. However, universal ubiquitination targets on microbes remain unidentified. Here, using in vitro, ex vivo, and in vivo studies, we identify the first protein-based ubiquitination substrates on phylogenetically diverse bacteria by unveiling a strategy that uses recognition of degron-like motifs. Such motifs form a new class of intra-cytosolic pathogen-associated molecular patterns (PAMPs). Their incorporation enabled recognition of nonubiquitin targets by host ubiquitin ligases. We find that SCF FBW7 E3 ligase, supported by the regulatory kinase, glycogen synthase kinase 3β, is crucial for effective pathogen detection and clearance. This provides a mechanistic explanation for enhanced risk of infections in patients with chronic lymphocytic leukemia bearing mutations in F-box and WD repeat domain containing 7 protein. We conclude that exploitation of this generic pathogen sensing strategy allows conservation of host resources and boosts antimicrobial immunity.

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“…Virulent type I strains are largely resistant to this pathway, while types II and III are susceptible (27), although the basis for this difference is unknown. Recent studies identified the E3 ubiquitin ligase RNF213 as a candidate for mediating early ubiquitination of T. gondii- containing vacuoles in IFN-γ treated A549 cells (30). The IFN-γ signaling and noncanonical ATG pathways are partially connected by ISG15, which is required for maximal recruitment of ATG adaptors and also interacts with RNF213 (28).…”

Section: Introductionmentioning

confidence: 99%

Molecular Basis for Interferon-mediated Pathogen Restriction in Human Cells

Matta

Kohio

Chandra

et al. 2022

Preprint

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To define novel mechanisms for cellular immunity to the intracellular pathogenToxoplasma gondii, we performed a genome-wide CRISPR loss-of-function screen to provide an unbiased assessment of genes important for IFN-γ-dependent growth restriction. We revealed a previously unknown role for the tumor suppressor NF-2/Merlin for maximum induction of Interferon Stimulated Genes (ISG), which are positively regulated by the transcription factor IRF-1. We then performed an additional focused ISG-targeted CRISPR screen that identified the host E3 ubiquitin ligase RNF213 as essential for IFN-γ mediated control ofT. gondii. RNF213 mediated ubiquitination of targets on the parasite-containing vacuole and growth restriction in response to IFN-γ in a variety of cell types, thus identifying a conserved factor that plays a prominent role in human cells. Surprisingly, growth inhibition did not require the autophagy protein ATG5, indicating that RNF213 initiates restriction independent of a non-canonical autophagy pathway that has previously been implicated in control ofT. gondii. RNF213 was also important for control of unrelated intracellular pathogens in human cells treated with IFN, as shown here forMycobacterium tuberculosisand Vesicular Stomatitis Virus. Collectively, our findings establish RNF213 as a critical component of cell-autonomous immunity to a broad spectrum of intracellular pathogens in human cells.

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Interferon-Inducible E3 Ligase RNF213 Facilitates Host-Protective Linear and K63-Linked Ubiquitylation of Toxoplasma gondii Parasitophorous Vacuoles

Cited by 23 publications

References 79 publications

A heterotrimeric complex ofToxoplasmaproteins promotes parasite survival in interferon gamma stimulated human cells

A heterotrimeric complex ofToxoplasmaproteins promotes parasite survival in interferon gamma stimulated human cells

An innate pathogen sensing strategy involving ubiquitination of bacterial surface proteins

Molecular Basis for Interferon-mediated Pathogen Restriction in Human Cells

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