A chloroplast-localized mitochondrial calcium uniporter transduces osmotic stress in Arabidopsis.

Chloroplasts are integral to sensing biotic and abiotic stress in plants, but their role in transducing Ca-mediated stress signals remains poorly understood. Here we identify cMCU, a member of the mitochondrial calcium uniporter (MCU) family, as an ion channel mediating Ca flux into chloroplasts in vivo. Using a toolkit of aequorin reporters targeted to chloroplast stroma and the cytosol in cMCU wild-type and knockout lines, we provide evidence that stress-stimulus-specific Ca dynamics in the chloroplast stroma correlate with expression of the channel.

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

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.

Evolutionary insight into the ionotropic glutamate receptor superfamily of photosynthetic organisms.

Photosynthetic eukaryotes have a complex evolutionary history shaped by multiple endosymbiosis events that required a tight coordination between the organelles and the rest of the cell. Plant ionotropic glutamate receptors (iGLRs) form a large superfamily of proteins with a predicted or proven non-selective cation channel activity regulated by a broad range of amino acids. They are involved in different physiological processes such as C/N sensing, resistance against fungal infection, root and pollen tube growth and response to wounding and pathogens.

A voltage-dependent chloride channel fine-tunes photosynthesis in plants.

In natural habitats, plants frequently experience rapid changes in the intensity of sunlight. To cope with these changes and maximize growth, plants adjust photosynthetic light utilization in electron transport and photoprotective mechanisms. This involves a proton motive force (PMF) across the thylakoid membrane, postulated to be affected by unknown anion (Cl(-)) channels.

Calcium Flux across Plant Mitochondrial Membranes: Possible Molecular Players.

Plants, being sessile organisms, have evolved the ability to integrate external stimuli into metabolic and developmental signals. A wide variety of signals, including abiotic, biotic, and developmental stimuli, were observed to evoke specific spatio-temporal Ca(2+) transients which are further transduced by Ca(2+) sensor proteins into a transcriptional and metabolic response. Most of the research on Ca(2+) signaling in plants has been focused on the transport mechanisms for Ca(2+) across the plasma- and the vacuolar membranes as well as on the components involved in decoding of cytoplasmic Ca(2+) signals, but how intracellular organelles such as mitochondria are involved in the process of Ca(2+) signaling is just emerging.

Physiology of intracellular potassium channels: A unifying role as mediators of counterion fluxes?

Plasma membrane potassium channels importantly contribute to maintain ion homeostasis across the cell membrane. The view is emerging that also those residing in intracellular membranes play pivotal roles for the coordination of correct cell function. In this review we critically discuss our current understanding of the nature and physiological tasks of potassium channels in organelle membranes in both animal and plant cells, with a special emphasis on their function in the regulation of photosynthesis and mitochondrial respiration.

Ion Channels in Plant Bioenergetic Organelles, Chloroplasts and Mitochondria: From Molecular Identification to Function.

Recent technical advances in electrophysiological measurements, organelle-targeted fluorescence imaging, and organelle proteomics have pushed the research of ion transport a step forward in the case of the plant bioenergetic organelles, chloroplasts and mitochondria, leading to the molecular identification and functional characterization of several ion transport systems in recent years. Here we focus on channels that mediate relatively high-rate ion and water flux and summarize the current knowledge in this field, focusing on targeting mechanisms, proteomics, electrophysiology, and physiological function. In addition, since chloroplasts evolved from a cyanobacterial ancestor, we give an overview of the information available about cyanobacterial ion channels and discuss the evolutionary origin of chloroplast channels.

The EF-Hand Ca2+ Binding Protein MICU Choreographs Mitochondrial Ca2+ Dynamics in Arabidopsis.

Plant organelle function must constantly adjust to environmental conditions, which requires dynamic coordination. Ca(2+) signaling may play a central role in this process. Free Ca(2+) dynamics are tightly regulated and differ markedly between the cytosol, plastid stroma, and mitochondrial matrix.

Alternative splicing-mediated targeting of the Arabidopsis GLUTAMATE RECEPTOR3.5 to mitochondria affects organelle morphology.

Since the discovery of 20 genes encoding for putative ionotropic glutamate receptors in the Arabidopsis (Arabidopsis thaliana) genome, there has been considerable interest in uncovering their physiological functions. For many of these receptors, neither their channel formation and/or physiological roles nor their localization within the plant cells is known. Here, we provide, to our knowledge, new information about in vivo protein localization and give insight into the biological roles of the so-far uncharacterized Arabidopsis GLUTAMATE RECEPTOR3.

MICU1 and MICU2 finely tune the mitochondrial Ca2+ uniporter by exerting opposite effects on MCU activity.

Mitochondrial calcium accumulation was recently shown to depend on a complex composed of an inner-membrane channel (MCU and MCUb) and regulatory subunits (MICU1, MCUR1, and EMRE). A fundamental property of MCU is low activity at resting cytosolic Ca(2+) concentrations, preventing deleterious Ca(2+) cycling and organelle overload. Here we demonstrate that these properties are ensured by a regulatory heterodimer composed of two proteins with opposite effects, MICU1 and MICU2, which, both in purified lipid bilayers and in intact cells, stimulate and inhibit MCU activity, respectively.

A thylakoid-located two-pore K+ channel controls photosynthetic light utilization in plants.

The size of the light-induced proton motive force (pmf) across the thylakoid membrane of chloroplasts is regulated in response to environmental stimuli. Here, we describe a component of the thylakoid membrane, the two-pore potassium (K(+)) channel TPK3, which modulates the composition of the pmf through ion counterbalancing. Recombinant TPK3 exhibited potassium-selective channel activity sensitive to Ca(2+) and H(+).

The mitochondrial calcium uniporter is a multimer that can include a dominant-negative pore-forming subunit.

Mitochondrial calcium uniporter (MCU) channel is responsible for Ruthenium Red-sensitive mitochondrial calcium uptake. Here, we demonstrate MCU oligomerization by immunoprecipitation and Förster resonance energy transfer (FRET) and characterize a novel protein (MCUb) with two predicted transmembrane domains, 50% sequence similarity and a different expression profile from MCU. Based on computational modelling, MCUb includes critical amino-acid substitutions in the pore region and indeed MCUb does not form a calcium-permeable channel in planar lipid bilayers.

Regulation of photosynthesis by ion channels in cyanobacteria and higher plants.

Photosynthesis converts light energy into chemical energy, and supplies ATP and NADPH for CO2 fixation into carbohydrates and for the synthesis of several compounds which are essential for autotrophic growth. Oxygenic photosynthesis takes place in thylakoid membranes of chloroplasts and photosynthetic prokaryote cyanobacteria. An ancestral photoautotrophic prokaryote related to cyanobacteria has been proposed to give rise to chloroplasts of plants and algae through an endosymbiotic event.

Molecular targets of antimicrobial photodynamic therapy identified by a proteomic approach.

Antimicrobial photodynamic therapy (PDT) is a promising tool to combat antibiotic-resistant bacterial infections. During PDT, bacteria are killed by reactive oxygen species generated by a visible light absorbing photosensitizer (PS). We used a classical proteomic approach that included two-dimensional gel electrophoresis and mass spectrometry analysis, to identify some proteins of Staphylococcus aureus that are damaged during PDT with the cationic PS meso-tetra-4-N-methyl pyridyl porphine (T4).

A forty-kilodalton protein of the inner membrane is the mitochondrial calcium uniporter.

Mitochondrial Ca(2+) homeostasis has a key role in the regulation of aerobic metabolism and cell survival, but the molecular identity of the Ca(2+) channel, the mitochondrial calcium uniporter, is still unknown. Here we have identified in silico a protein (named MCU) that shares tissue distribution with MICU1 (also known as CBARA1), a recently characterized uniporter regulator, is present in organisms in which mitochondrial Ca(2+) uptake was demonstrated and whose sequence includes two transmembrane domains. Short interfering RNA (siRNA) silencing of MCU in HeLa cells markedly reduced mitochondrial Ca(2+) uptake.

ATP-sensitive cation-channel in wheat (Triticum durum Desf.): identification and characterization of a plant mitochondrial channel by patch-clamp.

Indirect evidence points to the presence of K(+) channels in plant mitochondria. In the present study, we report the results of the first patch clamp experiments on plant mitochondria. Single-channel recordings in 150 mM potassium gluconate have allowed the biophysical characterization of a channel with a conductance of 150 pS in the inner mitochondrial membrane of mitoplasts obtained from wheat (Triticum durum Desf.

Dual localization of plant glutamate receptor AtGLR3.4 to plastids and plasmamembrane.

Bioinformatic approaches have allowed the identification in Arabidopsis thaliana of twenty genes encoding for homologues of animal ionotropic glutamate receptors (iGLRs). Some of these putative receptor proteins, grouped into three subfamilies, have been located to the plasmamembrane, but their possible location in organelles has not been investigated so far. In the present work we provide multiple evidence for the plastid localization of a glutamate receptor, AtGLR3.

Characterization of a plant glutamate receptor activity.

Bioinformatic approaches have allowed the identification of twenty genes, grouped into three subfamilies, encoding for homologues of animal ionotropic glutamate receptors (iGLRs) in the Arabidopsis thaliana model plant. Indirect evidence suggests that plant iGLRs function as non-selective cation channels. In the present work we provide biochemical and electrophysiological evidences for the chloroplast localization of glutamate receptor(s) of family 3 (iGLR3) in spinach.

A novel potassium channel in photosynthetic cyanobacteria.

Elucidation of the structure-function relationship of a small number of prokaryotic ion channels characterized so far greatly contributed to our knowledge on basic mechanisms of ion conduction. We identified a new potassium channel (SynK) in the genome of the cyanobacterium Synechocystis sp. PCC6803, a photosynthetic model organism.

Evidences for interaction of PsbS with photosynthetic complexes in maize thylakoids.

The PsbS subunit of Photosystem II (PSII) has received much attention in the past few years, given its crucial role in photoprotection of higher plants. The exact location of this small subunit in thylakoids is also debated. In this work possible interaction partners of PsbS have been identified by immunoaffinity and immunoprecipitation, performed with mildly solubilized whole thylakoid membrane.

Localization of a putative ClC chloride channel in spinach chloroplasts.

Seven genes seem to encode for putative ClC chloride channels (AtClC-a to AtClC-g) in Arabidopsis thaliana. Their function and localization is still largely unknown. AtClC-f shares considerable sequence similarity with putative ClC channel proteins from Synechocystis, considered to represent the precursor of chloroplasts.

Light- and pH-dependent structural changes in the PsbS subunit of photosystem II.

In higher plants, the PsbS subunit of photosystem II (PSII) plays a crucial role in pH- and xanthophyll-dependent nonphotochemical quenching of excess absorbed light energy, thus contributing to the defense mechanism against photoinhibition. We determined the amino acid sequence of Zea mays PsbS and produced an antibody that recognizes with high specificity a region of the protein located in the stroma-exposed loop between the second and third putative helices. By means of this antiserum, the thylakoid membranes of various higher plant species revealed the presence of a 42-kDa protein band, indicating the formation of a dimer of the 21-kDa PsbS protein.