Biochemical and phylogenetic analyses of phosphatidylinositol production in Angomonas deanei, an endosymbiont-harboring trypanosomatid

Allan C de Azevedo-Martins, João MP Alves, Fernando Garcia de Mello, Ana Tereza R Vasconcelos, Wanderley de Souza, Marcelo Einicker-Lamas, Maria Cristina M Motta

The endosymbiosis in trypanosomatids is characterized by co-evolution between one bacterium and its host protozoan in a mutualistic relationship, thus constituting an excellent model to study organelle origin in the eukaryotic cell. In this association, an intense metabolic exchange is observed between both partners: the host provides energetic molecules and a stable environment to a reduced wall symbiont, while the bacterium is able to interfere in host metabolism by enhancing phospholipid production and completing essential biosynthesis pathways, such as amino acids and hemin production. The bacterium envelope presents a reduced cell wall which is mainly composed of cardiolipin and phosphatidylcholine, being the latter only common in intracellular prokaryotes. Phosphatidylinositol (PI) is also present in the symbiont and host cell membranes. This phospholipid is usually related to cellular signaling and to anchor surface molecules, which represents important events for cellular interactions. In order to investigate the production of PI and its derivatives in symbiont bearing trypanosomatids, aposymbiotic and wild type strains of Angomonas deanei, as well as isolated symbionts, were incubated with [(3)H]myo-inositol and the incorporation of this tracer was analyzed into inositol-containing molecules, mainly phosphoinositides and lipoproteins. Gene searches and their phylogenies were also performed in order to investigate the PI synthesis in symbiontbearing trypanosomatids. Our results showed that the bacterium did not incorporate the tracer and that both strains produced similar quantities of PI and its derivatives, indicating that the symbiont does not influence the production of these metabolites. Gene searches related to PI synthesis revealed that the trypanosomatid genome contains an inositol transporter, PI synthase and the myo-inositol synthase. Thus, the host is able to produce PI either from exogenous myo-inositol (inositol transporter) or from myo-inositol synthesized de novo. Phylogenetic analysis using other organisms as references indicated that, in trypanosomatids, the genes involved in PI synthesis have a monophyletic origin. In accordance with experimental data, sequences for myo-inositol transport or for myo-inositol and PI biosynthesis were not found in the symbiont. Altogether, our results indicate that the bacterium depends on the host to obtain PI.