Sialic Acid Glycobiology Unveils Trypanosoma cruzi Trypomastigote Membrane Physiology

LANTOS A; RUIZ DÍAZ P; BOSSI M; BERTOZZI C; LANTOS A; RUIZ DÍAZ P; BOSSI M; BERTOZZI C; CARLEVARO G; CÁMARA MD L; YU H; MUCCI J*; ARAOZ B; CARLEVARO G; BUSCAGLIA CA; CÁMARA MD L; CHEN X; YU H; CAMPETELLA O*; MUCCI J*; ARAOZ B; BUSCAGLIA CA; CHEN X; CAMPETELLA O*

PLOS PATHOGENS

PUBLIC LIBRARY SCIENCE

Lugar: San Francisco; Año: 2016 vol. 12 p. 1 - 29

1553-7366

Juan Mucci and Oscar Campetella are corresponding authors and co-senior authors.Trypanosoma cruzi, the flagellate protozoan agent of Chagas disease or Americantrypanosomiasis, is unable to synthesize sialic acids de novo. Mucins and transsialidase(TS) are substrate and enzyme, respectively, of the glycobiological systemthat scavenges sialic acid from the host in a crucial interplay for T. cruzi life cycle. The acquisition of the sialyl residue allows the parasite to avoid lysis by serum factors andto interact with the host cell. A major drawback to study the sialylation kinetics andturnover of the trypomastigote glycoconjugates is the difficulty to identify and follow therecently acquired sialyl residues. To tackle this issue, we followed an unnatural sugarapproach as bioorthogonal chemical reporters where the use of azidosialyl residuesallowed identifying the acquired sugar. Advanced microscopy techniques together with biochemical methods were used to study the trypomastigote membrane from itsglycobiological perspective. Main sialyl acceptors were identified as mucins bybiochemical procedures and protein markers. Together with determining their sheddingand turnover rates, we also report that several membrane proteins including TS and itssubstrates, both glycosylphosphatidylinositol-anchored proteins, are separatelydistributed on parasite surface and contained in different and highly stable membranemicrodomains. Notably, labeling for α(1,3)Galactosyl residues only partially colocalizewith sialylated mucins indicating that two species of glycosylated mucins do exist,which are segregated at the parasite surface. Moreover, sialylated mucins wereincluded in lipid-raft-domains whereas TS molecules are not. The location of thesurface-anchored TS resulted too far off as to be capable to sialylate mucins, a roleplayed by the shed TS instead. Phosphatidylinositol-phospholipase-C activity isactually not present in trypomastigotes. Therefore, shedding of TS occurs viamicrovesicles instead of as a fully soluble form.