TbVps15 plays a role in vesicular transport and cytokinesis in Trypanosoma brucei

TAMARA STERNLIEB; MILENA MASSIMINO STEPÑIKA; KILDARE MIRANDA; PATRICIO GENTA; GUILLERMO D. ALONSO; ALEJANDRA C. SCHOIJET; BARRERA, NADIA M.; DE SOUZA, WANDERLEY

Ciudad Autónoma de Buenos Aires

Congreso; Reunión Conjunta de Sociedades de Biociencias. XXIX Reunión Anual de la Sociedad Argentina de Protozoología; 2017

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. Previously, we have reported the cloning and characterization of both TcVps34 and TcVps15 in Trypanosoma cruzi and demonstrated that in this parasite these proteins function as a complex.In addition, we showed that TcVps34 plays a prominent role in vital processes for T. cruzi survival such as osmoregulation, acidification, endocytosis and vesicular trafficking, and that both TcVps34 and TcVps15 are involved in autophagy in this parasite. 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 microscopy 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. Uptake of dextran, transferrin and ConcanavalinA were impaired. Finally, TbVps15 silencing affected both general kinase activity and PI3K activity, supporting the hypothesis that both proteins are conforming a complex as occurs in other organisms. In summary, we propose that T. brucei TbVps15 has a role in the maintenance of cytokinesis, endocytosis and intracellular trafficking in this parasite.