Coupling Auxin-Inducible Degron System with Ultrastructure Expansion Microscopy to Accelerate the Discovery of Gene Function in Toxoplasma gondii

Pacheco, Nicolas Dos Santos; Soldati‐Favre, Dominique

doi:10.1007/978-1-0716-1681-9_8

Cited by 20 publications

(22 citation statements)

References 12 publications

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“…Recent progress on genetic manipulation of the parasites has allowed identifying novel factors conferring fitness and virulence to T. gondii in vivo [ 215–217 ]. Genetic tools, when coupled to computational modeling [ 218 ] or novel imaging techniques [ 219 , 220 ], can also reveal new potential drug targets. Besides, new in vivo or in vitro differentiation models [ 102 , 221–223 ] offer interesting perspectives for investigating different developmental stages that were largely understudied until now.…”

Section: Discussionmentioning

confidence: 99%

The pathogenicity and virulence of Toxoplasma gondii

Sanchez

Besteiro

2021

Virulence

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Toxoplasma gondii is a parasitic protist infecting a wide group of warm-blooded animals, ranging from birds to humans. While this infection is usually asymptomatic in healthy individuals, it can also lead to severe ocular or neurological outcomes in immunocompromised individuals or in developing fetuses. This obligate intracellular parasite has the ability to infect a considerable range of nucleated cells and can propagate in the intermediate host. Yet, under the pressure of the immune system it transforms into an encysted persistent form residing primarily in the brain and muscle tissues. Encysted parasites, which are resistant to current medication, may reactivate and give rise to an acute infection. The clinical outcome of toxoplasmosis depends on a complex balance between the host immune response and parasite virulence factors. Susceptibility to the disease is thus determined by both parasite strains and host species. Recent advances on our understanding of host cell-parasite interactions and parasite virulence have brought new insights into the pathophysiology of T. gondii infection and are summarized here.

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Section: Discussionmentioning

confidence: 99%

The pathogenicity and virulence of Toxoplasma gondii

Sanchez

Besteiro

2021

Virulence

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“…The latter often relies on conditional regulation strategies to characterise function ( Figure 8 ). Less reducing screening methods, such as coupling tagging and conditional mutant generation with high-content fluorescent imaging Li et al., 2022 ; Smith et al., 2022 , or ultrastructure expansion microscopy (U-ExM) ( Dos Santos Pacheco and Soldati-Favre, 2021 ) would allow testing a greater number of candidates and increase the descriptive detail of each, thus making the best use of screening outputs (see Imaging and Figure 8 ). Using unrestricted screens to classify mutants based on observations from high-content imaging would reduce the waste and/or duplication associated with reducing screening methods.…”

Section: Functional Screeningmentioning

confidence: 99%

“…Stimulated emission depletion (STED) ( Willig et al., 2006 ), stochastic optical reconstruction microscopy (STORM) ( Rust et al., 2006 ), and structured illumination microscopy (SIM) ( Vangindertael et al., 2018 ) have all been used in the context of Kinetoplastid and Apicomplexan research. Expansion microscopy (ExM) ( Chen et al., 2015b ) and ultrastructure expansion microscopy (U-ExM) ( Gambarotto et al., 2021 ) have also been incorporated with great success ( Amodeo et al., 2021 ; Bertiaux et al., 2021 ; Dos Santos Pacheco and Soldati-Favre, 2021 ; Gorilak et al., 2021 ; Kalichava and Ochsenreiter, 2021 ; Liffner and Absalon, 2021 ; Tomasina et al., 2021 ). Despite their value, important bottlenecks for using super-resolution in a high-throughput manner are the time required for image acquisition, and the volume of data produced for quantitation.…”

Section: Imagingmentioning

confidence: 99%

Paving the Way: Contributions of Big Data to Apicomplexan and Kinetoplastid Research

Kent

Briggs²,

Colon³

et al. 2022

Front. Cell. Infect. Microbiol.

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In the age of big data an important question is how to ensure we make the most out of the resources we generate. In this review, we discuss the major methods used in Apicomplexan and Kinetoplastid research to produce big datasets and advance our understanding of Plasmodium, Toxoplasma, Cryptosporidium, Trypanosoma and Leishmania biology. We debate the benefits and limitations of the current technologies, and propose future advancements that may be key to improving our use of these techniques. Finally, we consider the difficulties the field faces when trying to make the most of the abundance of data that has already been, and will continue to be, generated.

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Section: Introductionmentioning

confidence: 99%

“…Since 2015 there has been a steady growth of the publication involving this technique, especially in protozoan parasites, like Giardia lamblia, Plasmodium falciparum , and Toxoplasma gondii (Tillberg and Chen 2019; dos Santos Pacheco and Soldati-Favre 2021; Gorilak et al 2021; Tomasina et al 2021). The use of ultrastructure expansion microscopy was first demonstrated in Kinetoplastea localizing a component of the mitochondrial genome segregation machinery in Trypanosoma brucei (Amodeo et al 2021; Kalichava and Ochsenreiter 2021) but it has not been reported in T. cruzi .…”

Section: Introductionmentioning

confidence: 99%

Ultrastructure Expansion Microscopy (U-ExM) in Trypanosoma cruzi: localization of tubulin isoforms and isotypes

Alonso

2022

Preprint

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Ultrastructure Expansion Microscopy (U-ExM) is a recently developed technique that enables the increase of the spatial resolution within a cell or a tissue for microscopic imaging by physically expanding the sample. I report, for the first time, a detailed protocol validating the use of U-ExM in Trypanosoma cruzi and quantifying the expansion factors of different subcellular compartments. I was able to determine the localization patterns of different tubulin isoforms, such as alpha-tubulin and beta-tubulin. Also, I immunolocalized acetylated and tyrosinated alpha-tubulin isotypes in epimastigotes and use mitochondrial cell-permeable dyes to identify this organelle. Finally, U-ExM was also performed in trypomastigotes and amastigotes validating this technique in all life cycle stages of T. cruzi.

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Coupling Auxin-Inducible Degron System with Ultrastructure Expansion Microscopy to Accelerate the Discovery of Gene Function in Toxoplasma gondii

Cited by 20 publications

References 12 publications

The pathogenicity and virulence of Toxoplasma gondii

The pathogenicity and virulence of Toxoplasma gondii

Paving the Way: Contributions of Big Data to Apicomplexan and Kinetoplastid Research

Ultrastructure Expansion Microscopy (U-ExM) in Trypanosoma cruzi: localization of tubulin isoforms and isotypes

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