The major Leishmania donovani chagasi surface glycoprotein in tunicamycin-resistant promastigotes.
Two populations of Leishmania donovani chagasi promastigotes resistant to the lethal effects of tunicamycin (TM), an inhibitor of N-linked glycosylation, were raised. These parasites exhibited altered patterns of glycosylation when compared to wild-type controls. In particular the major surface glycoprotein gp63 was present in membranes of one population of TM-resistant promastigotes (population 1) primarily in a deglycosylated form, which migrated at a lower Mr than wild-type gp63. The deglycosylated protein was proteolytically inactive on substrate-containing gels, in contrast to glycosylated gp63. Assays of promastigote attachment to human macrophages revealed that, despite a proposed role for gp63 protease activity in binding to macrophage receptors, population 1 TM-resistant promastigotes appeared to attach to the CR3 but not to the mannose-fucose receptor. Control promastigotes bound to both receptors. In contrast, a second population of TM-resistant promastigotes (population 2) did not produce gp63 that could be detected on immunoblots, either in a glycosylated or deglycosylated form. The latter TM-resistant promastigotes bound to neither CR3 nor the mannose-fucose receptor, suggesting that expression of gp63 might be important for promastigotes to bind to CR3. LPG was present in immunoblots of both TM-resistant and control populations suggesting this molecule might not account for the differences in attachment. Surprisingly both TM-resistant promastigote populations contained gp63 mRNA by Northern analysis in apparently equal amounts. We conclude that N-glycosylation is probably necessary for the protease function of gp63, but that neither N-glycosylation nor protease activity of gp63 is necessary for L. donovani chagasi promastigotes to bind to CR3. Furthermore the expression of gp63 protein by TM-resistant promastigotes is dependent upon postranscriptional events.
Wilson
M E
ME
Department of Internal Medicine, VA Medical Center, Iowa City, IA.
Hardin
K K
KK
D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D003208:Concanavalin A / Q000378:metabolism; D004351:Drug Resistance; D006031:Glycosylation; D016023:Integrins / Q000235:genetics; D037181:Lectins, C-Type; D007893:Leishmania donovani / Q000187:drug effects* / Q000235:genetics / Q000472:pathogenicity; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins; D008562:Membrane Glycoproteins / Q000235:genetics*; D008666:Metalloendopeptidases; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000378:metabolism; D015800:Protozoan Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic / Q000378:metabolism; D014415:Tunicamycin / Q000494:pharmacology*
D016023:Integrins; D037181:Lectins, C-Type; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins; D008562:Membrane Glycoproteins; D015800:Protozoan Proteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D003208:Concanavalin A; D014415:Tunicamycin; D010447:Peptide Hydrolases; D008666:Metalloendopeptidases; C055146:glycoprotein gp63, Leishmania