Abstract
Previous observations suggested a concomitant relationship between the release of the variant surface glycoprotein (VSG) and the activation of adenylate cyclase in the bloodstream form of the parasitic protozoan Trypanosoma brucei. In order to evaluate this hypothesis, adenylate cyclase activity was measured in live trypanosomes subjected to different treatments known to induce the shedding of the VSG coat, namely low pH and trypsin digestion. In both cases adenylate cyclase activation occurred in parallel with the release of the VSG. The latter was found to be mediated by the glycosylphosphatidylinositol-specific phospholipase C that cleaves the glycosylphosphatidylinositol anchor of the protein (VSG lipase). Furthermore, both adenylate cyclase and VSG release were activated by the incubation of trypanosomes with specific inhibitors of protein kinase C, suggesting a repressive role for protein kinase C on both VSG lipase and adenylate cyclase activities. Significantly, in mutant trypanosomes lacking VSG lipase, adenylate cyclase was activated under conditions where VSG release did not occur. Moreover,VSG release was also found to occur in the absence of activation of the cyclase, as observed in the presence of low concentration of the thiol modifying reagent p-chloromercuriphenylsulfonic acid. These observations provide the first demonstration that release of the VSG in response to cellular stress is mediated by the VSG lipase and that while both release of the VSG and activation of adenylate cyclase occur in response to the same stimuli they are not obligatorily coupled.
Highlights
Trypanosoma brucei, the parasitic protozoan causative of Nagana in the African cattle, is transmitted between mammals by the tsetse fly
As DMG is a potent stimulator of protein kinase C (PKC), we investigated whether the adenylate cyclase activation observed during variant surface glycoprotein (VSG) release would depend on stimulation of PKC by DMG
A series of kinetic studies demonstrated that the release of the VSG by treatments as different as mild acid or trypsin digestion led to a strong activation of adenylate cyclase, confirming similar observations made under other experimental approaches [23,24,25]
Summary
Trypanosoma brucei, the parasitic protozoan causative of Nagana in the African cattle, is transmitted between mammals by the tsetse fly. On the basis of observations made in other eukaryotes, it is probable that activation of cell surface receptors by appropriate ligands may cause the generation of second messengers which trigger changes in the programming of gene expression Typical in this respect is the generation of cAMP produced as a result of the stimulation of adenylate cyclase [2,3,4]. In dividing procyclic and bloodstream forms the activity of adenylate cyclase is down-regulated, since it can be strongly stimulated upon rupturing of the cells [17] In both intact cells and isolated plasma membranes [11], the adenylate cyclase activity of trypanosomes was found to be insensitive to agents known to activate G-protein-responsive adenylate cyclases, such as GTP␥S, Gpp(NH)p, forskolin, cholera, and pertussis toxins. In view of the observations reported for Dictyostelium [20], it may be suggested that the trypanosomal adenylate cyclases differ in their response to specific ligands and are involved in distinct events of transformation and/or proliferation in the parasite life cycle
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