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J Biol Chem. 2018 Dec 4. pii: jbc.RA118.005357. doi: 10.1074/jbc.RA118.005357. [Epub ahead of print]

Protein O-fucosyltransferase 2-mediated O-glycosylation of the adhesin MIC2 is dispensable for Toxoplasma gondii tachyzoite infection.

Author information

1
The Walter and Eliza Hall Institute, Australia.
2
The Walter and Eliza Hall Institute.
3
University of Michigan.
4
Department of Microbiology & Immunology, University of Michigan Medical School; Medical Science Bldg. II, Rm 5740B, United States.
5
Division of Infection and Immunity, The Walter and Eliza Hall Institute of Medical Research, Australia.
6
Chemical Biology, The Walter and Eliza Hall Institute of Medical Research, Australia.
7
Department of Microbiology and Immunology, University of Melbourne, Australia.

Abstract

Toxoplasma gondii is a ubiquitous, obligate intracellular eukaryotic parasite that causes congenital birth defects, disease in immunocompromised individuals, and blindness. Protein glycosylation plays an important role in the infectivity and evasion of immune responses of many eukaryotic parasites and is also of great relevance to vaccine design. Here, we demonstrate that micronemal protein 2 (MIC2), a motility-associated adhesin of T. gondii, has highly glycosylated thrombospondin repeat (TSR) domains. Using affinity-purified MIC2 and MS/MS analysis along with enzymatic digestion assays, we observed that at least seven C-linked and three O-linked glycosylation sites exist within MIC2, with > 95% occupancy at these O-glycosylation sites. We found that the addition of O-glycans to MIC2 is mediated by a protein O-fucosyltransferase 2 homologue (TgPOFUT2) encoded by the TGGT1_273550 gene. Even though POFUT2 homologs are important for stabilizing motility-associated adhesins and for host infection in other apicomplexan parasites, loss of TgPOFUT2 in T. gondii had only a modest impact on MIC2 levels and the wider parasite proteome. Consistent with this, both plaque formation and tachyzoite invasion were broadly similar in the presence or absence of TgPOFUT2. These findings indicate that TgPOFUT2 O-glycosylates MIC2 and that this glycan, in contrast to previous findings in another study, is dispensable in T. gondii tachyzoites and for T. gondii infectivity.

KEYWORDS:

Toxoplasma gondii; fucosyltransferase; glycosylation; mass spectrometry (MS); proteomics

PMID:
30514763
DOI:
10.1074/jbc.RA118.005357
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