Na+-dependent and Na+-independent mechanisms for inorganic phosphate uptake in Trypanosoma rangeli

Trypanosoma rangeli is dependent on the presence of exogenous orthophosphate (Pi) for maximal growth and ecto-phosphatase activity is responsible for Pi supply under low Pi. Here we investigated the mechanisms of Pi uptake.

We investigated the kinetics of 32Pi transport, its Na+ and H+ dependence, its correlation with the Na+-ATPase and H+-ATPase, and gene expression of the Na+:Pi cotransporter and Na+-ATPase.

T. rangeli grown under limiting Pi transports this anion to the cytosol in the absence and presence of Na+, suggesting that influx is mediated by both Na+-independent and Na+-dependent transporters. Cloning studies demonstrated that this parasite expresses a Pi transporter not previously studied in trypanosomatids. The H+ ionophore, carbonylcyanide-p-trifluoromethoxyphenylhydrazone, decreased both components of 32Pi influx by 80-95%. The H+-ATPase inhibitor, bafilomycin A1, inhibited the Na+-independent mechanism. Furosemide, an inhibitor of ouabain-insensitive Na+-ATPase, decreased both uptake mechanisms of 32Pi to the same extent, whereas ouabain had no effect, indicating that the former is the pump responsible for inwardly directed Na+ and the electric gradients required by the transporters. Parasite growth in high Pi had a lower Pi influx than that found in those grown in low Pi, without alteration in TrPho89 expression, showing that turnover of the transporters is stimulated by Pi starvation.

Two modes of Pi transport, one coupled to Na+-ATPase and other coupled to H+-ATPase seem to be responsible for Pi acquisition during development of T. rangeli.

This study provides the first description of the mechanism of Pi transport across the plasma membrane of trypanosomatids.