is the agent of Chagas disease， and the finding that this parasite possesses a mitochondrial calcium uniporter (TcMCU) with characteristics similar to that of mammalian mitochondria was fundamental for the discovery of the molecular nature of MCU in eukaryotes. We report here that ablation of ， or its paralog ， by clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 led to a marked decrease in mitochondrial Ca uptake without affecting the membrane potential of these cells， whereas overexpression of each gene caused a significant increase in the ability of mitochondria to accumulate Ca While knockout (KO) epimastigotes were viable and able to differentiate into trypomastigotes， infect... More
is the agent of Chagas disease， and the finding that this parasite possesses a mitochondrial calcium uniporter (TcMCU) with characteristics similar to that of mammalian mitochondria was fundamental for the discovery of the molecular nature of MCU in eukaryotes. We report here that ablation of ， or its paralog ， by clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 led to a marked decrease in mitochondrial Ca uptake without affecting the membrane potential of these cells， whereas overexpression of each gene caused a significant increase in the ability of mitochondria to accumulate Ca While knockout (KO) epimastigotes were viable and able to differentiate into trypomastigotes， infect host cells， and replicate normally， ablation of resulted in epimastigotes having an important growth defect， lower rates of respiration and metacyclogenesis， more pronounced autophagy changes under starvation， and significantly reduced infectivity. Overexpression of ， in contrast to what was proposed for its mammalian ortholog， did not result in a dominant negative effect on TcMCU. The finding of a mitochondrial calcium uniporter (MCU) in was essential for the discovery of the molecular nature of this transporter in mammals. In this work， we used the CRISPR/Cas9 technique that we recently developed for to knock out two components of the uniporter: MCU， the pore subunit， and MCUb， which was proposed as a negative regulator of MCU in human cells. In contrast to what occurs in human cells， MCU is not essential， while MCUb is essential for growth， differentiation， and infectivity; has a bioenergetic role; and does not act as a dominant negative subunit of MCU.
