In Situ Compositional Analysis of Acidocalcisomes in Trypanosoma cruzi

Abstract

We measured the elemental content of different compartments in Trypanosoma cruzi epimastigotes using quick freezing, ultracryomicrotomy, and electron probe microanalysis. Vacuoles identified by high electron density contained (in units of mmol/kg dry weight ؎ S.E.) large amounts of phosphorus (1390 ؎ 13), magnesium (646 ؎ 19), calcium (171 ؎ 5), sodium (161 ؎ 18), and zinc (148 ؎ 6). No other compartment had appreciable calcium or zinc content. Iron (128 ؎ 16 mmol/kg) was detected only in vacuoles distinct from the electron-dense vacuoles and other organelles. Incubation of cells for 70 min in culture medium in the presence of ionomycin plus nigericin led to a very significant 3-or 2-fold increase in potassium in the electron-dense vacuoles and the iron-rich vacuoles, respectively, with no significant change in the other elements investigated. This indicated the acidic nature of the vacuoles and demonstrated that the electron-dense vacuoles correspond to what were described previously as acidocalcisomes, i.e. acidic compartments rich in Ca 2؉ . The acidocalcisomes were investigated by separation of epimastigote fractions on Percoll gradients in combination with Triton WR-1339 treatment. This detergent caused a rapid vacuolation; these vacuoles were shown by electron microscopy to be largely transparent, with a diffuse matrix. Percoll gradient fractionation demonstrated decreases in the density of various organelle markers in detergent-treated cells compared with controls. Large decreases in the density of the acidocalcisome and the mitochondrion were seen, as well as smaller decreases in the density of the other markers. Conventional electron microscopy of epimastigotes loaded with gold-labeled transferrin indicated that the endosomal system was separate from vacuoles that probably corresponded to the calcium-containing organelles detected by electron probe microanalysis. The combined results provide evidence that acidocalcisomes are organelles different from lysosomes or other organelles previously described in these parasites.

Factors Affecting the Level and Localization of the Transferrin Receptor in Trypanosoma brucei

“…A calibration curve at a specimen thickness of 200 nm showed our detection limit to be 20 mM iron, i.e. far less than the level found in the iron-rich organelles of T. cruzi (42). This suggests that iron storage in T. brucei does not involve a similar organelle as in T. cruzi.…”

“…Cytosolic aconitase, the iron sensor in mammalian cells, is not involved in T. brucei iron sensing (5). In T. cruzi iron is stored in specialized organelles detected by electron probe analysis (42). We have used the same techniques to search for iron in T. brucei.…”

In situ structural analysis of Golgi intracisternal protein arrays

“…4A). These vacuoles, called acidocalcisomes, are acidified compartments that store polyphosphate complexed with high concentrations of cations, including calcium, iron, zinc, and copper (46)(47)(48). Thus, an alternative candidate for the trans-Golgi filaments is polyphosphate, the major nonorganic component of acidocalcisomes (49)(50)(51).…”

The reservosome of Trypanosoma cruzi epimastigotes: an organelle of the endocytic pathway with a role on metacyclogenesis

“…However, ultrastructural demonstration that the inner inclusions were not membrane bound, together with cytochemical evidentiation that these vesicles were indeed lipid droplets, revealed the misinterpretation of the former nomenclature, and the name reservosome was then proposed (Soares & De Souza 1988). Reservosomes are morphologically and biochemically distinct from the recently described acidocalcisomes (Scott et al 1997). As reservosomes are a site for protein accumulation, two major questions arise: how proteins arrive there, and what reservosomes are for?…”