Chip-calorimetric assessment of heat generation during Ca2+ uptake by digitonin-permeabilized Trypanosoma cruzi.

Ca2+ signaling in trypanosomatids is an important component of energy metabolism regulation and therefore, cytosolic Ca2+ concentration is finely regulated by Ca2+ transport through the plasma membrane and Ca2+ uptake and release by intracellular organelles. To maintain intracellular Ca2+ homeostasis with different gradients of the ion within the cellular compartments, there is an energy cost and also energy dissipation in the form of heat. Using an innovative segmented fusion technique of a chip-calorimeter and CRISPR/Cas9 knockout (–KO) Trypanosoma cruzi cell lines, we evaluated the heat generation during Ca2+ uptake by digitonin-permeabilized T. cruzi epimastigotes, a system consisting of Ca2+ uptake predominantly by mitochondria and acidocalcisomes. We used three T. cruzi epimastigotes cell lines: control cells denominated scrambled, cells with the absence of the pyruvate dehydrogenase phosphatase (TcPDP-KO) and cells lacking mitochondrial Ca2+ uptake via the mitochondrial calcium uniporter (TcMCU-KO), that presented, in this respective order, decreasing rates and capacities of Ca2+ uptake. TcPDP-KO cells exhibited the lowest heat production following Ca2+ addition, which may be due to its lower mitochondrial oxidative phosphorylation capacity and lower ATP availability for acidocalcisomal Ca2+ uptake. Scrambled and TcMCU-KO cells exhibited similar Ca2+-induced heat effects, which correlates with a higher ATP-dependent acidocalcisomal Ca2+ uptake in these cells. Our results show evidences that mitochondrial Ca2+ transport via the uniporter is minimally heat dissipative, while ATPase pumps in acidocalcisomes possess a predominant contribution to the heat generated during Ca2+ uptake. [ABSTRACT FROM AUTHOR]