Leucine zipper EF hand-containing transmembrane protein 1 (Letm1) and uncoupling proteins 2 and 3 (UCP2/3) contribute to two distinct mitochondrial Ca 2+ uptake pathways

Abstract

Cytosolic Ca 2+ signals are transferred into mitochondria over a huge concentration range. In our recent work we described uncoupling proteins 2 and 3 (UCP2/3) to be fundamental for mitochondrial uptake of high Ca 2+ domains in mitochondria-ER junctions. On the other hand, the leucine zipper EF handcontaining transmembrane protein 1 (Letm1) was identified as a mitochondrial Ca 2+/H + antiporter that achieved mitochondrial Ca 2+ sequestration at small Ca 2+ increases. Thus, the contributions of Letm1 and UCP2/3 to mitochondrial Ca 2+ uptake were compared in endothelial cells. Knock-down of Letm1 did not affect the UCP2/3-dependent mitochondrial uptake of intracellularly released Ca 2+ but strongly diminished the transfer of entering Ca 2+ into mitochondria, subsequently, resulting in a reduction of store-operated Ca 2+ entry (SOCE). Knock-down of Letm1 and UCP2/3 did neither impact on cellular ATP levels nor the membrane potential. The enhanced mitochondrial Ca 2+signals in cells overexpressing UCP2/3 rescued SOCE upon Letm1 knock-down. In digitonin-permeabilized cells, Letm1 exclusively contributed to mitochondrial Ca 2+ uptake at low Ca 2+ conditions. Neither the Letm1- nor the UCP2/3-dependent mitochondrial Ca 2+ uptake was affected by a knock-down of mRNA levels of mitochondrial calcium uptake 1 (MICU1), a protein that triggers mitochondrial Ca 2+ uptake in HeLa cells. Our data indicate that Letm1 and UCP2/3 independently contribute to two distinct, mitochondrial Ca 2+ uptake pathways in intact endothelial cells. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.