Physiological Characterization of a Plant Mitochondrial Calcium Uniporter in Vitro and in Vivo

Abstract

Over the recent years, several proteins that make up the mitochondrial calcium uniporter complex (MCUC) mediating Cauptake into the mitochondrial matrix have been identified in mammals, including the channel-forming protein MCU. Although six MCU gene homologs are conserved in the model plant Arabidopsis (Arabidopsis thaliana) in which mitochondria can accumulate Ca, a functional characterization of plant MCU homologs has been lacking. Using electrophysiology, we show that one isoform, AtMCU1, gives rise to a Ca-permeable channel activity that can be observed even in the absence of accessory proteins implicated in the formation of the active mammalian channel. Furthermore, we provide direct evidence that AtMCU1 activity is sensitive to the mitochondrial calcium uniporter inhibitors Ruthenium Red and Gd, as well as to the Arabidopsis protein MICU, a regulatory MCUC component. AtMCU1 is prevalently expressed in roots, localizes to mitochondria, and its absence causes mild changes in Ca dynamics as assessed by in vivo measurements in Arabidopsis root tips. Plants either lacking or overexpressing AtMCU1 display root mitochondria with altered ultrastructure and show shorter primary roots under restrictive growth conditions. In summary, our work adds evolutionary depth to the investigation of mitochondrial Ca transport, indicates that AtMCU1, together with MICU as a regulator, represents a functional configuration of the plant mitochondrial Ca uptake complex with differences to the mammalian MCUC, and identifies a new player of the intracellular Ca regulation network in plants.

The mitochondrial complexome of Arabidopsis thaliana

“…MICU was shown to regulate mitochondrial calcium ion dynamics in vivo , while the MCU protein is one of six Arabidopsis homologs of MCU, a protein forming the pore of the channel complex in mammals (Stael et al ., ; Wagner et al ., ). Two MCU homologs were also detected in the potato mitochondrial proteome (Salvato et al ., ; Rao et al ., ), and a very recent functional characterization of one Arabidopsis MCU homolog (MCU1: At1g09575) showed channel activity in planar lipid bilayers and inhibition by MICU (Teardo et al ., ). In mammals MCU needs to assemble into a homo‐tetrameric or homo‐pentameric transmembrane complex to make up the pore, which needs to associate with regulatory proteins (incl.…”

Advances and current challenges in calcium signaling

“…Because the outer mitochondrial membrane is permeable for Ca 2+ , a similar [Ca 2+ ] is shared by the cytosol and the mitochondrial intermembrane space. However, for import into the matrix, specialized Ca 2+ uniporter complexes (MCUC) have been identified in animals and homologs were found in plants (Foskett & Philipson, 2015;Teardo et al, 2017). In particular, the pore-forming MCU (mitochondrial Ca 2+ uniporter) is well conserved and shows a similar regulatory mechanism involving additional complex proteins (Baughman et al, 2011;Foskett & Philipson, 2015).…”

“…In particular, the pore-forming MCU (mitochondrial Ca 2+ uniporter) is well conserved and shows a similar regulatory mechanism involving additional complex proteins (Baughman et al, 2011;Foskett & Philipson, 2015). One of this regulatory proteins is provided by the mitochondrial Ca 2+ uptake 1 protein (MICU) (Wagner et al, 2015;Teardo et al, 2017). MICUs harbor EF-hand domains and could sense matrix [Ca 2+ ] as a Ca 2+ sensor, restricting its uptake upon Ca 2+ saturation.…”

Chloroplast Calcium Signaling in the Spotlight

“…Recently, two proteins belonging to the family of the mitochondrial calcium uniporter (MCU) have been found to mediate Ca 2+ transport across the mitochondria and chloroplast membranes, respectively AtMCU1 (Teardo et al, 2017) and AtMCU6 (later renamed AtcMCU (Teardo et al, 2019). In animal cells the only isoform, MCU (De Stefani et al, 2011;Baughman, 2011) is responsible for Ca 2+ loading into mitochondria, thus helping recovery of resting [Ca 2+ ] cyt .…”