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Abstract
Trypanosoma brucei is the causative agent of African Trypanosomiasis, a deadly disease affecting humans and cattle. The inositol 1,4,5 triphosphate/diacylglycerol (IP3/DAG) pathway regulates important processes in many organisms. T. brucei has an active IP3 receptor, localized to the acidocalcisome, that is essential for infection in mice. T. brucei has a phosphoinositide phospholipase C (TbPI-PLC, Tb927.11.5970) that hydrolyses phosphatidylinositol (PI) and phosphatidylinositol 4,5-bisphosphate (PIP2). However, knockdown of TbPI-PLC expression by RNAi did not affect growth. Here we investigate if a PI-PLC-like protein (TbPI-PLC-like, Tb927.6.2090) is involved in the IP3/DAG pathway. TbPI-PLC-like was identified in a proteomics study of palmitoylated proteins of T. brucei, and it is a highly conserved protein in all Trypanosomatids. The protein has an X catalytic domain but lacks the Y catalytic domain possessing instead a PDZ domain, in addition it has an has an N-terminal myristoylation consensus sequence. Recombinant TbPI-PLC-like does not hydrolyze PI or PIP2 and does not bind to IP3. Knockdown of TbPI-PLC-like expression by RNAi did not change the levels of IP6 in the cells. Taken together these results suggest that TbPI-PLC-like is not involved in the IP3/DAG pathway. We also tested if TbPI-PLC-like was a modulator of TbPI-PLC. Recombinant TbPI-PLC-like did not modulate TbPI-PLC activity and knockdown of TbPI-PLC-like expression by RNAi did not affect the expression of TbPI-PLC. However, it did result in growth inhibition and decreased virulence in vivo indicating that this protein is important for the parasite. Preliminary mass spectrometry analysis of the possible binding partners of TbPI-PLC-like show strong interactions with the translational machinery.