Bosch Navarrete, C

Castillo Acosta, Víctor M.

González-Pacanowska, D.

Ruiz Pérez, LM

Valente, M

Yagüe Capilla, M

English

González-Pacanowska, Dolores

Digital.CSIC

560 INMUNOLOGíA E INFECCIOSASI Congreso de Investigadores del PTS. Granada 13-15 Feb. 2019ESSENTIALITY AND ROLE OF NOVELNUCLEOTIDASES INVOLVED INTRYPANOSOMA BRUCEI PYRIMIDINEHOMEOSTASISYagüe Capilla M, Castillo Acosta VM, Valente M, BoschNavarrete C, Ruiz Pérez LM, González Pacanowska DInstituto de Parasitología y Biomedicina López Neyra. Consejo Superior deInvestigaciones Científicas. Granada, SpainCorrespondencia: miyaca31@ipb.csic.esModalidad: PósterNucleotide metabolism has been an area of great interest for the dis-covery of novel targets against many diseases since a balanced pool ofdeoxyribonucleotides is required for correct DNA replication and repair.Particularly relevant for cell survival is the maintenance of a balanceddUTP/dTTP ratio in the pyrimidine pool, as several DNA polymerasescannot distinguish between the two nucleotides and incorporate indiscrimi-nately one or the other depending on their availability. We have previouslyshown that thymidine kinase (TbTK) has a major role in the maintenanceof the dUTP/dTTP ratio and the response to genotoxic agents in bloods-tream forms of Trypanosoma brucei. We reported that TbTK is essentialfor parasite viability, both in vitro and in vivo thus demonstrating thatphosphorylation of deoxyuridine and/or thymidine is important for themaintenance of the dTTP pool even in the absence of a source of extra-cellular pyrimidines. These observations indicated a role of the enzymein de novo synthesis and pointed towards the existence of an intracellu-lar deoxynucleoside pool available for phosphorylation. In this work, wehave aimed at characterizing nucleotidases that could be involved in thegeneration of intracellular metabolites important for thymidylate de novobiosynthesis. Interestingly, we have identified in Trypanosoma brucei anHD domain containing nucleotidase highly similar to human SAMHD1.The absence of the protein in knock-out mutants results in a loss of cellINMUNOLOGíA E INFECCIOSAS 561viability and disturbances in cell cycle progression, which could be com-pletely reverted by the supplementation of thymidine or deoxyuridine. Inaddition, the lack of the HD nucleotidase could also be rescued by the ex-pression of the human dCMP deaminase, which catalyses the deaminationof dCMP to dUMP. Metabolomic analysis by mass spectrometry demons-trated that the absence of the enzyme gives rise to severe perturbations inpyrimidine metabolism. Thus, altogether our results suggest that this HDdomain containing nucleotidase is a vital contributor to dTTP biosynthe-sis and we propose this class of enzymes as relevant players in nucleotidehomeostasis in trypanosomes.

Essentiality and role of novel nucleotidases involved in Trypanosoma brucei pyrimidine homeostasis

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Essentiality and role of novel nucleotidases involved in Trypanosoma brucei pyrimidine homeostasis

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