TbVps15 is required for vesicular transport and cytokinesis in Trypanosoma brucei.

The class III phosphatidylinositol 3-kinase (PI3K) Vps34 is an important regulator of key cellular functions, including cell growth, survival, intracellular trafficking, autophagy and nutrient sensing. In yeast, Vps34 is associated with the putative serine/threonine protein kinase Vps15, however, its role in signaling has not been deeply evaluated. Here, we have identified the Vps15 orthologue in Trypanosoma brucei, named TbVps15. Knockdown of TbVps15 expression by interference RNA resulted in inhibition of cell growth and blockage of cytokinesis. Scanning electron microcopy revealed a variety of morphological abnormalities, with enlarged parasites and dividing cells that often exhibited a detached flagellum. Transmission electron microscopy analysis of TbVps15 RNAi cells showed an increase in intracellular vacuoles of the endomembrane system and some cells displayed an enlargement of the flagellar pocket, a common feature of cells defective in endocytosis. Moreover, uptake of dextran, transferrin and Concanavalin A was impaired. Finally, TbVps15 downregulation affected the PI3K activity, supporting the hypothesis that TbVps15 and TbVps34 form a complex as occurs in other organisms. In summary, we propose that TbVps15 has a role in the maintenance of cytokinesis, endocytosis and intracellular trafficking in T. brucei.