title: Proteins anchored via glycosylphosphatidylinositol and solubilizing phospholipases in Trypanosoma cruzi.
abstract: The presence of GPI anchors and phospholipases capable of solubilizing them in Trypanosoma cruzi has been investigated in epimastigotes, metacyclic trypomastigotes from axenic cultures and tissue culture trypomastigotes. The GPI anchored proteins in epimastigote forms are scarce when compared to their abundance in the parasite forms which can infect mammals, and GPI-solubilizing phospholipases C have been found in all life cycles stages. In epimastigote and metacyclic forms, the activity is found in the soluble fraction upon cell lysis, whereas in tissue cultured trypomastigotes it is membrane bound and, being mostly sensitive to p-chloromercuriphenylsulfonate, resembles closely the GPI specific phospholipase of Trypanosoma brucei. Sequential immunoprecipitations with monoclonal antibodies and anti-CRD indicated the presence of several sub-populations among the surface proteins of metacyclic trypomastigotes, five of these belonging to the GPI-anchored 90 kD family. Among this family, the epitopes recognized by MAb-1G7 are present in three members, one of them also expressing the 3F6 epitope. There are 2 members recognized only by MAb-3F6 but not by MAb-1G7, one of them being probably galactosylated on the GPI since it can be immunoprecipitated by anti-CRD. Very strangely, the epitope recognized by the MAb-WIC29.26 was always present on the gp72, as originally described, but under certain circumstances appeared cryptic on one of the 90 kD species. During epimastigote transformation into metacyclic trypomastigotes in vitro, the ability of the GPI of the 1G7-antigen to be solubilized by phospholipase C and D varies depending on the age of the culture and presence or absence of fetal calf serum. Different patterns of solubilization were also obtained for 1G7-Ag, depending on whether the test is performed with parasite lysates or with antigen affinity purified from them. Our data indicate that the phospholipase C resistance observed does not arise from acylation on the inositol, as previously described for acetylcholinesterase from human erythrocytes, being rather due to factors which either modify the GPI or affect the action of the phospholipases. Previously unreported resistance to glycosylphosphatidylinositol-specific phospholipase D has been observed both to glycosylphosphatidylinositol-specific phospholipase D has been observed both to 1G7-Ag and 10D8-Ag, the GPI-anchored mucynlike protein which is acceptor of sialic acid in metacyclic forms. Our findings are discussed in the light of the presently known structures of GPI in this parasite, and imaginative speculation on biological roles for the GPI phospholipase system in T. cruzi is also provided.
authors.affiliation: Department of Microbiology, Immunology and Parasitology, Escola Paulista de Medicina, São Paulo, SP, Brazil.
authors.forename: M L
authors.initials: ML
authors.lastname: de Almeida
authors.forename: N
authors.initials: N
authors.lastname: Heise
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan / Q000276:immunology / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D010740:Phospholipases / Q000378:metabolism*; D011233:Precipitin Tests / Q000379:methods; D015800:Protozoan Proteins / Q000378:metabolism*; D012995:Solubility; D014346:Trypanosoma brucei brucei / Q000378:metabolism; D014349:Trypanosoma cruzi / Q000201:enzymology / Q000254:growth & development / Q000276:immunology / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan; D017261:Glycosylphosphatidylinositols; D015800:Protozoan Proteins; D010740:Phospholipases; D010738:Type C Phospholipases