Cardiolipin (CL) is a multifunctional dimeric phospholipid that physically interacts with electron transport chain complexes I, III, and IV, and ATP synthase (complex V). The enzyme ALCAT1 catalyzes the conversion of cardiolipin by incorporating polyunsaturated fatty acids into cardiolipin. The resulting CL species are said to be more susceptible to oxidative damage. This is thought to negatively affect the interaction of cardiolipin and electron transport chain complexes, leading to increased ROS production and mitochondrial dysfunction. Furthermore, it is discussed that ALCAT1 itself is upregulated due to oxidative stress. Here, we investigated the effects of overexpression of ALCAT1 under different metabolic conditions. ALCAT1 is located at the ER and mitochondria, probably at contact sites. We found that respiration stimulated by galactose supply promoted supercomplex assembly but also led to increased mitochondrial ROS levels. Endogeneous ALCAT1 protein expression levels showed a fairly high variability. Artificially induced ALCAT1 overexpression reduced supercomplex formation, further promoted ROS production, and prevented upregulation of coupled respiration. Taken together, our data suggest that the amount of the CL conversion enzyme ALCAT1 is critical for coupling mitochondrial respiration and metabolic plasticity.
- MeSH
- 1-Acylglycerol-3-Phosphate O-Acyltransferase genetics metabolism MeSH
- Cell Respiration MeSH
- Galactose metabolism MeSH
- HeLa Cells MeSH
- Cardiolipins metabolism MeSH
- Humans MeSH
- Membrane Potential, Mitochondrial MeSH
- Mitochondria metabolism MeSH
- Protein Multimerization genetics MeSH
- Multiprotein Complexes metabolism MeSH
- Oxidative Stress MeSH
- Reactive Oxygen Species metabolism MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
The monomeric photosystem I-light-harvesting antenna complex I (PSI-LHCI) supercomplex from the extremophilic red alga Cyanidioschyzon merolae represents an intermediate evolutionary link between the cyanobacterial PSI reaction center and its green algal/higher plant counterpart. We show that the C. merolae PSI-LHCI supercomplex is characterized by robustness in various extreme conditions. By a combination of biochemical, spectroscopic, mass spectrometry, and electron microscopy/single particle analyses, we dissected three molecular mechanisms underlying the inherent robustness of the C. merolae PSI-LHCI supercomplex: (1) the accumulation of photoprotective zeaxanthin in the LHCI antenna and the PSI reaction center; (2) structural remodeling of the LHCI antenna and adjustment of the effective absorption cross section; and (3) dynamic readjustment of the stoichiometry of the two PSI-LHCI isomers and changes in the oligomeric state of the PSI-LHCI supercomplex, accompanied by dissociation of the PsaK core subunit. We show that the largest low light-treated C. merolae PSI-LHCI supercomplex can bind up to eight Lhcr antenna subunits, which are organized as two rows on the PsaF/PsaJ side of the core complex. Under our experimental conditions, we found no evidence of functional coupling of the phycobilisomes with the PSI-LHCI supercomplex purified from various light conditions, suggesting that the putative association of this antenna with the PSI supercomplex is absent or may be lost during the purification procedure.
- MeSH
- Adaptation, Biological MeSH
- Chlorophyll metabolism MeSH
- Circular Dichroism MeSH
- Spectrometry, Fluorescence MeSH
- Photosystem I Protein Complex chemistry metabolism MeSH
- Hydrogen-Ion Concentration MeSH
- Evolution, Molecular MeSH
- Rhodophyta chemistry physiology MeSH
- Cyanobacteria chemistry physiology MeSH
- Light MeSH
- Light-Harvesting Protein Complexes chemistry metabolism MeSH
- Temperature MeSH
- Zeaxanthins metabolism MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Cyclic electron transport (CET) around photosystem I (PSI) plays an important role in balancing the ATP/NADPH ratio and the photoprotection of plants. The NAD(P)H dehydrogenase complex (NDH) has a key function in one of the CET pathways. Current knowledge indicates that, in order to fulfill its role in CET, the NDH complex needs to be associated with PSI; however, until now there has been no direct structural information about such a supercomplex. Here we present structural data obtained for a plant PSI-NDH supercomplex. Electron microscopy analysis revealed that in this supercomplex two copies of PSI are attached to one NDH complex. A constructed pseudo-atomic model indicates asymmetric binding of two PSI complexes to NDH and suggests that the low-abundant Lhca5 and Lhca6 subunits mediate the binding of one of the PSI complexes to NDH. On the basis of our structural data, we propose a model of electron transport in the PSI-NDH supercomplex in which the association of PSI to NDH seems to be important for efficient trapping of reduced ferredoxin by NDH.
- MeSH
- Microscopy, Electron MeSH
- Ferredoxins metabolism MeSH
- Photosystem I Protein Complex chemistry isolation & purification metabolism MeSH
- Hordeum chemistry enzymology radiation effects MeSH
- Plant Leaves chemistry enzymology radiation effects MeSH
- Models, Molecular MeSH
- NAD metabolism MeSH
- NADPH Dehydrogenase chemistry isolation & purification metabolism MeSH
- Native Polyacrylamide Gel Electrophoresis MeSH
- Oxidation-Reduction MeSH
- Light MeSH
- Light-Harvesting Protein Complexes chemistry isolation & purification metabolism MeSH
- Electron Transport MeSH
- Thylakoids metabolism MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Chlorobaculum tepidum is a representative of green sulfur bacteria, a group of anoxygenic photoautotrophs that employ chlorosomes as the main light-harvesting structures. Chlorosomes are coupled to a ferredoxin-reducing reaction center by means of the Fenna-Matthews-Olson (FMO) protein. While the biochemical properties and physical functioning of all the individual components of this photosynthetic machinery are quite well understood, the native architecture of the photosynthetic supercomplexes is not. Here we report observations of membrane-bound FMO and the analysis of the respective FMO-reaction center complex. We propose the existence of a supercomplex formed by two reaction centers and four FMO trimers based on the single-particle analysis of the complexes attached to native membrane. Moreover, the structure of the photosynthetic unit comprising the chlorosome with the associated pool of RC-FMO supercomplexes is proposed.
- MeSH
- Bacterial Proteins chemistry metabolism ultrastructure MeSH
- Chlorobi chemistry MeSH
- Cytoplasm chemistry MeSH
- Photosynthetic Reaction Center Complex Proteins chemistry metabolism MeSH
- Intracellular Membranes chemistry MeSH
- Light-Harvesting Protein Complexes chemistry metabolism ultrastructure MeSH
- Microscopy, Electron, Transmission MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
In the present paper, we report an improved method combining sucrose density gradient with ion-exchange chromatography for the isolation of pure chlorophyll a/c antenna proteins from the model cryptophytic alga Rhodomonas salina. Antennas were used for in vitro quenching experiments in the absence of xanthophylls, showing that protein aggregation is a plausible mechanism behind non-photochemical quenching in R. salina. From sucrose gradient, it was also possible to purify a functional photosystem I supercomplex, which was in turn characterized by steady-state and time-resolved fluorescence spectroscopy. R. salina photosystem I showed a remarkably fast photochemical trapping rate, similar to what recently reported for other red clade algae such as Chromera velia and Phaeodactylum tricornutum. The method reported therefore may also be suitable for other still partially unexplored algae, such as cryptophytes.
Mitochondria are double membrane organelles of endosymbiotic origin, best known for constituting the centre of energetics of a eukaryotic cell. They contain their own mitochondrial genome, which as a consequence of gradual reduction during evolution typically contains less than two dozens of genes. In this review, we highlight the extremely diverse architecture of mitochondrial genomes and mechanisms of gene expression between the three sister groups constituting the phylum Euglenozoa - Euglenida, Diplonemea and Kinetoplastea. The earliest diverging euglenids possess a simplified mitochondrial genome and a conventional gene expression, whereas both are highly complex in the two other groups. The expression of their mitochondrial-encoded proteins requires extensive post-transcriptional modifications guided by complex protein machineries and multiple small RNA molecules. Moreover, the least studied diplonemids, which have been recently discovered as a highly abundant component of the world ocean plankton, possess one of the most complicated mitochondrial genome organisations known to date.
- Publication type
- Journal Article MeSH
- Review MeSH
Photosystem I (PSI) is a pigment-protein complex required for the light-dependent reactions of photosynthesis and participates in light-harvesting and redox-driven chloroplast metabolism. Assembly of PSI into supercomplexes with light harvesting complex (LHC) II, cytochrome b6f (Cytb6f) or NAD(P)H dehydrogenase complex (NDH) has been proposed as a means for regulating photosynthesis. However, structural details about the binding positions in plant PSI are lacking. We analyzed large data sets of electron microscopy single particle projections of supercomplexes obtained from the stroma membrane of Arabidopsis thaliana. By single particle analysis, we established the binding position of Cytb6f at the antenna side of PSI. The rectangular-shaped Cytb6f dimer binds at the side where Lhca1 is located. The complex binds with its short side rather than its long side to PSI, which may explain why these supercomplexes are difficult to purify and easily disrupted. Refined analysis of the interaction between PSI and the NDH complex indicates that in total up to 6 copies of PSI can arrange with one NDH complex. Most PSI-NDH supercomplexes appeared to have 1-3 PSI copies associated. Finally, the PSI-LHCII supercomplex was found to bind an additional LHCII trimer at two positions on the LHCI side in Arabidopsis. The organization of PSI, either in a complex with NDH or with Cytb6f, may improve regulation of electron transport by the control of binding partners and distances in small domains.
- MeSH
- Arabidopsis metabolism MeSH
- Chlorophyll metabolism MeSH
- Chloroplasts metabolism MeSH
- Photosynthesis physiology MeSH
- Photosystem I Protein Complex metabolism MeSH
- Cytochrome b6f Complex metabolism MeSH
- NADH Dehydrogenase metabolism MeSH
- Oxidation-Reduction MeSH
- Light MeSH
- Light-Harvesting Protein Complexes metabolism MeSH
- Electron Transport physiology MeSH
- Thylakoids metabolism MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
In nature, plants are continuously exposed to varying environmental conditions. They have developed a wide range of adaptive mechanisms, which ensure their survival and maintenance of stable photosynthetic performance. Photosynthesis is delicately regulated at the level of the thylakoid membrane of chloroplasts and the regulatory mechanisms include a reversible formation of a large variety of specific protein-protein complexes, supercomplexes or even larger assemblies known as megacomplexes. Revealing their structures is crucial for better understanding of their function and relevance in photosynthesis. Here we focus our attention on the isolation and a structural characterization of various large protein supercomplexes and megacomplexes, which involve Photosystem II and Photosystem I, the key constituents of photosynthetic apparatus. The photosystems are often attached to other protein complexes in thylakoid membranes such as light harvesting complexes, cytochrome b 6 f complex, and NAD(P)H dehydrogenase. Structural models of individual supercomplexes and megacomplexes provide essential details of their architecture, which allow us to discuss their function as well as physiological significance.
AIMS: Expression of the HER2 oncogene in breast cancer is associated with resistance to treatment, and Her2 may regulate bioenergetics. Therefore, we investigated whether disruption of the electron transport chain (ETC) is a viable strategy to eliminate Her2(high) disease. RESULTS: We demonstrate that Her2(high) cells and tumors have increased assembly of respiratory supercomplexes (SCs) and increased complex I-driven respiration in vitro and in vivo. They are also highly sensitive to MitoTam, a novel mitochondrial-targeted derivative of tamoxifen. Unlike tamoxifen, MitoTam efficiently suppresses experimental Her2(high) tumors without systemic toxicity. Mechanistically, MitoTam inhibits complex I-driven respiration and disrupts respiratory SCs in Her2(high) background in vitro and in vivo, leading to elevated reactive oxygen species production and cell death. Intriguingly, higher sensitivity of Her2(high) cells to MitoTam is dependent on the mitochondrial fraction of Her2. INNOVATION: Oncogenes such as HER2 can restructure ETC, creating a previously unrecognized therapeutic vulnerability exploitable by SC-disrupting agents such as MitoTam. CONCLUSION: We propose that the ETC is a suitable therapeutic target in Her2(high) disease. Antioxid. Redox Signal. 26, 84-103.
- MeSH
- Biomarkers MeSH
- Cell Death drug effects MeSH
- Cell Respiration drug effects MeSH
- Molecular Targeted Therapy MeSH
- Electron Transport Chain Complex Proteins antagonists & inhibitors chemistry metabolism MeSH
- Inhibitory Concentration 50 MeSH
- Humans MeSH
- Membrane Potential, Mitochondrial drug effects MeSH
- Mitochondria drug effects metabolism MeSH
- Molecular Conformation MeSH
- Models, Molecular MeSH
- Cell Line, Tumor MeSH
- Breast Neoplasms drug therapy metabolism pathology MeSH
- Antineoplastic Agents chemistry pharmacology MeSH
- Reactive Oxygen Species metabolism MeSH
- Receptor, ErbB-2 antagonists & inhibitors metabolism MeSH
- Electron Transport Complex I antagonists & inhibitors chemistry metabolism MeSH
- Tamoxifen pharmacology MeSH
- Protein Binding MeSH
- Check Tag
- Humans MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
Tamoxifen resistance remains a clinical obstacle in the treatment of hormone sensitive breast cancer. It has been reported that tamoxifen is able to target respiratory complex I within mitochondria. Therefore, we established two tamoxifen-resistant cell lines, MCF7 Tam5R and T47D Tam5R resistant to 5 μM tamoxifen and investigated whether tamoxifen-resistant cells exhibit mitochondrial changes which could help them survive the treatment. The function of mitochondria in this experimental model was evaluated in detail by studying i) the composition and activity of mitochondrial respiratory complexes; ii) respiration and glycolytic status; iii) mitochondrial distribution, dynamics and reactive oxygen species production. We show that Tam5R cells exhibit a significant decrease in mitochondrial respiration, low abundance of assembled mitochondrial respiratory supercomplexes, a more fragmented mitochondrial network connected with DRP1 Ser637 phosphorylation, higher glycolysis and sensitivity to 2-deoxyglucose. Tam5R cells also produce significantly higher levels of mitochondrial superoxide but at the same time increase their antioxidant defense (CAT, SOD2) through upregulation of SIRT3 and show phosphorylation of AMPK at Ser 485/491. Importantly, MCF7 ρ0 cells lacking functional mitochondria exhibit a markedly higher resistance to tamoxifen, supporting the role of mitochondria in tamoxifen resistance. We propose that reduced mitochondrial function and higher level of reactive oxygen species within mitochondria in concert with metabolic adaptations contribute to the phenotype of tamoxifen resistance.
- MeSH
- Apoptosis MeSH
- Cell Cycle MeSH
- Drug Resistance, Neoplasm * MeSH
- Phenotype MeSH
- Glycolysis * MeSH
- Antineoplastic Agents, Hormonal pharmacology MeSH
- Humans MeSH
- Mitochondria metabolism pathology MeSH
- Mice, Nude MeSH
- Mice MeSH
- Tumor Cells, Cultured MeSH
- Breast Neoplasms drug therapy metabolism pathology MeSH
- Cell Movement MeSH
- Cell Proliferation MeSH
- Reactive Oxygen Species metabolism MeSH
- Electron Transport Complex I metabolism MeSH
- Superoxides metabolism MeSH
- Tamoxifen pharmacology MeSH
- Xenograft Model Antitumor Assays MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Mice MeSH
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
