Virulent and Avirulent Strains of Toxoplasma gondii Which Differ in Their Glycosylphosphatidylinositol Content Induce Similar Biological Functions in Macrophages

Niehus, Sebastian; Smith, Terry; Azzouz, Nahid; Campos, Marco Antônio; Dubremetz, Jean‐François; Gazzinelli, Ricardo T.; Schwarz, Ralph Τ.; Debierre‐Grockiego, Françoise

doi:10.1371/journal.pone.0085386

Cited by 14 publications

(10 citation statements)

References 60 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…6A). In addition, there is strain variation with regard to the presence of the phosphoethanolamine and ␣Glc termini in GIPLs (66). Besides serving a structural role, GPI-anchors and possibly GIPLs are recognized by host cell TLR2 and probably galectins as co-receptors (21).…”

Section: Gpi-anchor and Gipl-glycogenesmentioning

confidence: 99%

CRISPR/Cas9 and glycomics tools for Toxoplasma glycobiology

Gas-Pascual

Ichikawa

Sheikh

et al. 2019

Journal of Biological Chemistry

View full text Add to dashboard Cite

Infection with the protozoan parasite Toxoplasma gondii is a major health risk owing to birth defects, its chronic nature, ability to reactivate to cause blindness and encephalitis, and high prevalence in human populations. Unlike most eukaryotes, Toxoplasma propagates in intracellular parasitophorous vacuoles, but like nearly all other eukaryotes, Toxoplasma glycosylates many cellular proteins and lipids and assembles polysaccharides. Toxoplasma glycans resemble those of other eukaryotes, but species-specific variations have prohibited deeper investigations into their roles in parasite biology and virulence. The Toxoplasma genome encodes a suite of likely glycogenes expected to assemble N-glycans, O-glycans, a C-glycan, GPI-anchors, and polysaccharides, along with their precursors and membrane transporters. To investigate the roles of specific glycans in Toxoplasma, here we coupled genetic and glycomics approaches to map the connections between 67 glycogenes, their enzyme products, the glycans to which they contribute, and cellular functions. We applied a double-CRISPR/Cas9 strategy, in which two guide RNAs promote replacement of a candidate gene with a resistance gene; adapted MS-based glycomics workflows to test for effects on glycan formation; and infected fibroblast monolayers to assess cellular effects. By editing 17 glycogenes, we discovered novel Glc 0-2-Man 6-GlcNAc 2-type N-glycans, a novel HexNAc-GalNAc-mucin-type O-glycan, and Tn-antigen; identified the glycosyltransferases for assembling novel nuclear O-Fuc-type and cell surface Glc-Fuc-type O-glycans; and showed that they are important for in vitro growth. The guide sequences, editing constructs, and mutant strains are freely available to researchers to investigate the roles of glycans in their favorite biological processes. Toxoplasma gondii is a worldwide, obligate intracellular apicomplexan parasite that can infect most nucleated cells of warm-blooded animals (1), with up to 80% of some human populations being seropositive (2). Toxoplasmosis, the disease caused by Toxoplasma, is associated with encephalitis and blindness in individuals whose parasites are reactivated, as can occur in AIDS and other immunosuppressed patients (3). In utero infections can cause mental retardation, blindness, and death (4). Toxoplasma is transmitted by digesting parasites from feline feces (as oocysts) or undercooked meat (as tissue cysts). Once in the host, parasites convert to the tachyzoite form that disseminates to peripheral tissues (e.g. brain, retina, and muscle). The resulting immune response and/or drugs can control tachyzoite replication, but the parasite survives by encysting into slowly growing bradyzoites. Sporadically, burst of cysts allows the parasites to convert to tachyzoites, whose unchecked growth results in cell and tissue damage (5, 6). Currently, no Toxoplasma vaccine exists, anti-toxoplasmosis drugs have severe side effects, and resistance is developing to these drugs (7-11). As individuals remain infected for life, new anti-Toxoplasma drugs a...

show abstract

Section: Gpi-anchor and Gipl-glycogenesmentioning

confidence: 99%

CRISPR/Cas9 and glycomics tools for Toxoplasma glycobiology

Gas-Pascual

Ichikawa

Sheikh

et al. 2019

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…6A). In addition, there is strain variation with regard to the presence of the phosphoethanolamine and αGlc termini in GIPLs (Niehus et al, 2014). Besides serving a structural role, GPI anchors and possibly GIPLs are recognized by host cell TLR2 and probably galectins as co-receptors (Debierre-Grockiego et al, 2010).…”

Section: Results and Dicussionmentioning

confidence: 99%

Enabling tools forToxoplasmaglycobiology

Gas-Pascual¹,

Ichikawa²,

Sheikh³

et al. 2018

Preprint

View full text Add to dashboard Cite

Abbreviations: AAL, Aleuria aurantia agglutinin or lectin; Dol-P-sugar, a conjugate of Glc or Man to dolichol phosphate; EIC, extracted ion chromatogram; FBS, fetal bovine serum; GPI, glycophosphatidylinositol; GT, glycosyltransferase; HAc, acetic acid; HdH, a disaccharide consisting of a hexose and a deoxy-hexose residue; HFF, human foreskin fibroblasts; hTERT, HFF immortalized with human telomerase reverse transcriptase; H6N2, an oligosaccharide consisting of 6 hexose (Hex) and 2 Nacetylhexosamine (HexNAc) residues; NHEJ, non-homologous end-joining; nLC-MS/MS, nano-liquid chromatography coupled to multi-dimensional mass spectrometry; mAb, monoclonal antibody; pp-αGalNAcT, UDP-GalNAc-dependent polypeptide α-N-acetyl-D-galactosaminyltransferase; Tn antigen, a structure corresponding to αGalNAc-S/T; TSR, thrombospondin type 1 repeats ABSTRACT Infection by the protozoan parasite Toxoplasma gondii is a major health risk owing to its chronic nature, ability to reactivate to cause blindness and encephalitis, and high prevalence in human populations. Like nearly all eukaryotes, Toxoplasma glycosylates many of its proteins and lipids and assembles polysaccharides. Unlike most eukaryotes, Toxoplasma divides and differentiates in vacuoles within host cells. While their glycans resemble canonical models, they exhibit species-specific variations that have inhibited deeper investigations into their roles in parasite biology and virulence. The genome of Toxoplasma encodes a suite of likely glycogenes expected to assemble a range of N-glycans, O-glycans, a C-glycan, GPI-anchors, and polysaccharides, along with their requisite precursors and membrane transporters. To facilitate genetic approaches to investigate the roles of specific glycans, we mapped probable connections between 59 glycogenes, their enzyme products, and the glycans to which they contribute. We adapted a double-CRISPR/Cas9 strategy and a mass spectrometry-based glycomics workflow to test these relationships, and conducted infection studies in fibroblast monolayers to probe cellular functions. Through the validated disruption of 17 glycogenes, we also discovered novel Glc0-2-Man6-GlcNAc2-type N-glycans, GalNAc2-and Glc-Fuc-type O-glycans, and a nuclear O-Fuc type glycan. We describe the guide sequences, disruption constructs, and mutant strains, which are freely available to practitioners for application in the context of the relational map to investigate the roles of glycans in their favorite biological processes.

show abstract

“…Differences between lipid types and quantities directly influence C. pseudotuberculosis pathogenicity and its capacity to form granulomas [22]. Analyses of cell wall lipids and glycolipids have been conducted with the objective of identifying and differentiating bacterial and protozoan genera, species, and strains [22][23][24]. In the current work, lipids derived from the cell walls of two C. pseudotuberculosis strains were purified with the objective of identifying any differences between them that may exist.…”

Section: Discussionmentioning

confidence: 99%

Cell wall glycolipids from Corynebacterium pseudotuberculosis strains with different virulences differ in terms of composition and immune recognition

et al. 2020

View full text Add to dashboard Cite

Caseous lymphadenitis (CLA) is an infectious disease caused by Corynebacterium pseudotuberculosis in small ruminants and is characterized by the development of granulomas in the lymph nodes, spleen, liver, and lungs. Although little is known about the host-pathogen relationship of this bacterium, it was previously reported that the pathogen's lipids are important for its taxonomic classification and survival inside macrophages. However, there are no studies regarding the composition of these molecules. In this study, cell wall glycolipids from two C. pseudotuberculosis strains presenting different virulence profiles were purified and its composition was characterized. A difference was observed between the electrophoretic and chromatogram profiles for cell wall components from the two strains, mainly among molecules with low molecular weights. IgM from sheep with acute CLA recognized antigens with an estimated molecular weight of 11 kDa of the low-pathogenicity strain, while low-molecular weight antigens from the high-pathogenicity strain presented a lower recognition by these antibodies. Mass spectrometry analysis showed that the cell wall of the high-pathogenicity strain contained glycolipids with high amounts of unsaturated fatty acids and glycerophosphoinositols, which may contribute to the capacity of this strain to cause severe disease. In conclusion, it is indicated that cell wall non-protein antigens can play a key role in C. pseudotuberculosis virulence.

show abstract

Virulent and Avirulent Strains of Toxoplasma gondii Which Differ in Their Glycosylphosphatidylinositol Content Induce Similar Biological Functions in Macrophages

Cited by 14 publications

References 60 publications

CRISPR/Cas9 and glycomics tools for Toxoplasma glycobiology

CRISPR/Cas9 and glycomics tools for Toxoplasma glycobiology

Enabling tools forToxoplasmaglycobiology

Cell wall glycolipids from Corynebacterium pseudotuberculosis strains with different virulences differ in terms of composition and immune recognition

Contact Info

Product

Resources

About