Inorganic phosphate uptake in Trypanosoma cruzi is coupled to K+ cycling and to active Na+ extrusion

Abstract Background Orthophosphate (P i ) is a central compound in the metabolism of all organisms, including parasites. There are no reports regarding the mechanisms of P i acquisition by Trypanosoma cruzi . Methods 32 P i influx was measured in T. cruzi epimastigotes. The expression of P i transporter genes and the coupling of the uptake to Na + , H + and K + fluxes were also investigated. The transport capacities of different evolutive forms were compared. Results Epimastigotes grew significantly more slowly in 2 mM than in 50 mM P i . Influx of P i into parasites grown under low P i conditions took place in the absence and presence of Na + . We found that the parasites express TcPho84, a H + :P i -symporter, and TcPho89, a Na + :P i -symporter. Both P i influx mechanisms showed Michaelis–Menten kinetics, with a one-order of magnitude higher affinity for the Na + -dependent system. Collapsing the membrane potential with carbonylcyanide- p -trifluoromethoxyphenylhydrazone strongly impaired the influx of P i . Valinomycin (K + ionophore) or SCH28028 (inhibitor of (H +  + K + )ATPase) significantly inhibited P i uptake, indicating that an inwardly-directed H + gradient energizes uphill P i entry and that K + recycling plays a key role in P i influx. Furosemide, an inhibitor of the ouabain-insensitive Na + -ATPase, decreased only the Na + -dependent P i uptake, indicating that this Na + pump generates the Na + gradient utilized by the symporter. Trypomastigote forms take up P i inefficiently. Conclusions P i starvation stimulates membrane potential-sensitive P i uptake through different pathways coupled to Na + or H + /K + fluxes. General significance This study unravels the mechanisms of P i acquisition by T. cruzi , a key process in epimastigote development and differentiation to trypomastigote forms.