Ca 2+ Content and Expression of an Acidocalcisomal Calcium Pump Are Elevated in Intracellular Forms of Trypanosoma cruzi

Abstract

ABSTRACT The survival of a eukaryotic protozoan as an obligate parasite in the interior of a eukaryotic host cell implies its adaptation to an environment with a very different ionic composition from that of its extracellular habitat. This is particularly important in the case of Ca 2+ , the intracellular concentration of which is 3 orders of magnitude lower than the extracellular value. Ca 2+ entry across the plasma membrane is a widely recognized mechanism for Ca 2+ signaling, needed for a number of intracellular processes, and obviously, it would be restricted in the case of intracellular parasites. Here we show that Trypanosoma cruzi amastigotes possess a higher Ca 2+ content than the extracellular stages of the parasite. This correlates with the higher expression of a calcium pump, the gene for which was cloned and sequenced. The deduced protein product (Tca1) of this gene has a calculated molecular mass of 121,141 Da and exhibits 34 to 38% identity with vacuolar Ca 2+ -ATPases of Saccharomyces cerevisiae and Dictyostelium discoideum , respectively. The tca1 gene suppresses the Ca 2+ hypersensitivity of a mutant of S. cerevisiae that has a defect in vacuolar Ca 2+ accumulation. Indirect immunofluorescence and immunoelectron microscopy analysis indicate that Tca1 colocalizes with the vacuolar H + -ATPase to the plasma membrane and to intracellular vacuoles of T. cruzi . These vacuoles were shown to have the same size and distribution as the calcium-containing vacuoles identified by the potassium pyroantimoniate-osmium technique and as the electron-dense vacuoles observed in whole unfixed parasites by transmission electron microscopy and identified in a previous work (D. A. Scott, R. Docampo, J. A. Dvorak, S. Shi, and R. D. Leapman, J. Biol. Chem. 272:28020–28029, 1997) as being acidic and possessing a high calcium content (i.e., acidocalcisomes). Together, these results suggest that acidocalcisomes are distinct from other previously recognized organelles present in these parasites and underscore the ability of intracellular parasites to adapt to the hostile environment of their hosts.