title: Making Home Sweet and Sturdy: Toxoplasma gondii ppGalNAc-Ts Glycosylate in Hierarchical Order and Confer Cyst Wall Rigidity.
abstract: The protozoan intracellular parasite Toxoplasma gondii forms latent cysts in the central nervous system (CNS) and persists for the lifetime of the host. This cyst is cloaked with a glycosylated structure called the cyst wall. Previously, we demonstrated that a mucin-like glycoprotein, CST1, localizes to the cyst wall and confers structural rigidity on brain cysts in a mucin-like domain-dependent manner. The mucin-like domain of CST1 is composed of 20 units of threonine-rich tandem repeats that are O-GalNAc glycosylated. A family of enzymes termed polypeptide N-acetylgalactosaminyltransferases (ppGalNAc-Ts) initiates O-GalNAc glycosylation. To identify which isoforms of ppGalNAc-Ts are responsible for the glycosylation of the CST1 mucin-like domain and to evaluate the function of each ppGalNAc-T in the overall glycosylation of the cyst wall, all five ppGalNAc-T isoforms were deleted individually from the T. gondii genome. The ppGalNAc-T2 and -T3 deletion mutants produced various glycosylation defects on the cyst wall, implying that many cyst wall glycoproteins are glycosylated by T2 and T3. Both T2 and T3 glycosylate the CST1 mucin-like domain, and this glycosylation is necessary for CST1 to confer structural rigidity on the cyst wall. We established that T2 is required for the initial glycosylation of the mucin-like domain and that T3 is responsible for the sequential glycosylation on neighboring acceptor sites, demonstrating hierarchical glycosylation by two distinct initiating and filling-in ppGalNAc-Ts in an intact organism.
  
  
  
  Toxoplasma gondii is an obligate intracellular parasite that infects a third of the world's population. It can cause severe congenital disease and devastating encephalitis in immunocompromised individuals. We identified two glycosyltransferases, ppGalNAc-T2 and -T3, which are responsible for glycosylating cyst wall proteins in a hierarchical fashion. This glycosylation confers structural rigidity on the brain cyst. Our studies provide new insights into the mechanisms of O-GalNAc glycosylation in T. gondii.
authors.affiliation: Department of Pathology, Albert Einstein College of Medicine, Bronx, New York, USA.
authors.forename: Tadakimi
authors.initials: T
authors.lastname: Tomita
authors.affiliation: Department of Pathology, Albert Einstein College of Medicine, Bronx, New York, USA.
authors.forename: Tatsuki
authors.initials: T
authors.lastname: Sugi
authors.affiliation: Department of Pathology, Albert Einstein College of Medicine, Bronx, New York, USA.
authors.forename: Rama
authors.initials: R
authors.lastname: Yakubu
authors.affiliation: Department of Pathology, Albert Einstein College of Medicine, Bronx, New York, USA.
authors.forename: Vincent
authors.initials: V
authors.lastname: Tu
authors.affiliation: Department of Pathology, Albert Einstein College of Medicine, Bronx, New York, USA.
authors.forename: Yanfen
authors.initials: Y
authors.lastname: Ma
authors.affiliation: Department of Pathology, Albert Einstein College of Medicine, Bronx, New York, USA louis.weiss@einstein.yu.edu.|Department of Medicine, Albert Einstein College of Medicine, Bronx, New York, USA.
authors.forename: Louis M
authors.initials: LM
authors.lastname: Weiss
mesh_terms: D017353:Gene Deletion; D006023:Glycoproteins / Q000378:metabolism*; D006031:Glycosylation; D017350:N-Acetylgalactosaminyltransferases / Q000235:genetics / Q000378:metabolism*; D015800:Protozoan Proteins / Q000378:metabolism*; D033761:Spores, Protozoan / Q000737:chemistry* / Q000254:growth & development; D014122:Toxoplasma / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D000097763:Polypeptide N-acetylgalactosaminyltransferase
chemical_list: D006023:Glycoproteins; D017350:N-Acetylgalactosaminyltransferases; D015800:Protozoan Proteins; D000097763:Polypeptide N-acetylgalactosaminyltransferase