The Interplay between Folding-facilitating Mechanisms inTrypanosoma cruziEndoplasmic Reticulum

Abstract

Lectin (calreticulin [CRT])-N-glycan-mediated quality control of glycoprotein folding is operative in trypanosomatid protozoa but protein-linked monoglucosylated N-glycans are exclusively formed in these microorganisms by UDP-Glc:glycoprotein glucosyltransferase (GT)-dependent glucosylation. The gene coding for this enzyme in the human pathogen Trypanosoma cruzi was identified and sequenced. Even though several of this parasite glycoproteins have been identified as essential components of differentiation and mammalian cell invasion processes, disruption of both GT-encoding alleles did not affect cell growth rate of epimastigote form parasites and only partially affected differentiation and mammalian cell invasion. The cellular content of one of the already identified T. cruzi glycoprotein virulence factors (cruzipain, a lysosomal proteinase) only showed a partial (5-20%) decrease in GT null mutants in spite of the fact that Ͼ90% of all cruzipain molecules interacted with CRT during their folding process in wild-type cells. Although extremely mild cell lysis and immunoprecipitation procedures were used, no CRT-cruzipain interaction was detected in GT null mutants but secretion of the proteinase was nevertheless delayed because of a lengthened interaction with Grp78/BiP probably caused by the detected induction of this chaperone in GT null mutants. This result provides a rationale for the absence of a more drastic consequence of GT absence. It was concluded that T. cruzi endoplasmic reticulum folding machinery presents an exquisite plasticity that allows the parasite to surmount the absence of the glycoprotein-specific folding facilitation mechanism.

Deletion of the Glucosidase II Gene in Trypanosoma brucei Reveals Novel N-Glycosylation Mechanisms in the Biosynthesis of Variant Surface Glycoprotein

“…We were also able to successfully differentiate the null mutant into procyclic form T. brucei in vitro (data not shown), suggesting that the gene is also not required for differentiation to, or survival of the procyclic form of the parasite. In this regard, our results are similar to those of Parodi and co-workers (34,39), who have shown that, in the related American parasite T. cruzi, other components of the ER glucosylation-dependent quality control system (i.e. calreticulin and UGGT) can be deleted with only moderate effects on parasite growth, differentiation, and infectivity.…”

The evolution of N-glycan-dependent endoplasmic reticulum quality control factors for glycoprotein folding and degradation

“…UGGTs that glucosylate N-glycans of misfolded proteins were demonstrated in three ways (7,8). First, genome project strains of Entamoeba and Trichomonas were grown and labeled with [2] 3 H-Man in the presence or absence of 200 g/ml of the glucosidase II inhibitor castanospermine (42).…”

Sugar Nucleotide Pools of Trypanosoma brucei , Trypanosoma cruzi , and Leishmania major

“…UDP-Glc is also found in all three parasites and is the glucosyl donor for the unfolded glycoprotein glucosyltransferase (UGGT) involved in glycoprotein quality control in the endoplasmic reticulum (42,117). UGGT is known to function in all three parasites (17,51,68,80). UDP-Glc is also the presumed donor for the synthesis of base J (␤-D-glucosylhydroxymethyluracil), a rare deoxynucleotide found in telomere-proximal DNA in the bloodstream form of T. brucei and in the leishmania and T. cruzi (121).…”