Trypanosoma brucei: characterisation of a life cycle stage-specific G-protein

G-proteins have important and diverse roles in cellular signaling in higher eukaryotes but as yet little is known about the involvement of these GTP-binding proteins in protozoan parasites. Bacterial toxin-catalysed ADP-ribosylation, routinely used in the characterisation of G-proteins in higher eukaryotes, was used to determine whether functional G-protein homologues could be detected in Trypanosoma brucei. A 43-kDa polypeptide that was ADP-ribosylated in the presence of pertussis toxin was identified in membrane fractions. This polypeptide was also recognised by an antibody generated against a peptide sequence representing a mammalian GO alpha G-protein subunit. GTP and GTP gamma S (a nonhydrolysable analogue of GTP) abolished the ADP-ribosylation reaction, indicating that the 43-kDa polypeptide also binds GTP. Furthermore, this T. brucei G-protein is present in bloodstream but not procyclic life cycle stages. Using an in situ GTP binding assay, a further group of small-molecular-size GTP-binding proteins which are present in both bloodstream and procyclic stages have been identified. This demonstration of G-proteins in T. brucei suggests possible intracellular mechanisms that may be involved in the transduction of host or other parasite signals.