A plasma membrane-type Ca2+-ATPase co-localizes with a vacuolar H+-pyrophosphatase to acidocalcisomes of Toxoplasma gondii

Abstract

Ca2+-ATPases are likely to play critical roles in the biochemistry of Toxoplasma gondii, since these protozoa are obligate intracellular parasites and the Ca2+ concentration in their intracellular location is three orders of magnitude lower than in the extracellular medium. Here, we report the cloning and sequencing of a gene encoding a plasma membrane-type Ca2+-ATPase (PMCA) of T.gondii (TgA1). The predicted protein (TgA1) exhibits 32-36% identity to vacuolar Ca2+-ATPases of Trypanosoma cruzi, Saccharomyces cerevisiae, Entamoeba histolytica and Dictyostelium discoideum. Sequencing of both cDNA and genomic DNA from T.gondii indicated that TgA1 contains two introns near the C-terminus. A hydropathy profile of the protein suggests 10 transmembrane domains. TgA1 suppresses the Ca2+ hypersensitivity of a mutant of S.cerevisiae that has a defect in vacuolar Ca2+ accumulation. Indirect immunofluorescence and immunoelectron microscopy analysis indicate that TgA1 localizes to the plasma membrane and co-localizes with the vacuolar H+-pyrophosphatase to intracellular vacuoles identified morphologically and by X-ray microanalysis as the acidocalcisomes. This vacuolar-type Ca2+-ATPase could play an important role in Ca2+ homeostasis in T. gondii.