Mitochondrial Alternative Oxidase Research Articles

Alternative Oxidase Alleviates Mitochondrial Oxidative Stress during Limited Nitrate Reduction in Arabidopsis thaliana.

The conversion of nitrate to ammonium, i.e., nitrate reduction, is a major consumer of reductants in plants. Previous studies have reported that the mitochondrial alternative oxidase (AOX) is upregulated under limited nitrate reduction conditions, including no/low nitrate or when ammonium is the sole nitrogen (N) source. Electron transfer from ubiquinone to AOX bypasses the proton-pumping complexes III and IV, thereby consuming reductants efficiently. Thus, upregulated AOX under limited nitrate reduction may dissipate excessive reductants and thereby attenuate oxidative stress. Nevertheless, so far there is no firm evidence for this hypothesis due to the lack of experimental systems to analyze the direct relationship between nitrate reduction and AOX. We therefore developed a novel culturing system for A. thaliana that manipulates shoot activities of nitrate reduction and AOX separately without causing N starvation, ammonium toxicity, or lack of nitrate signal. Using shoots processed with this system, we examined genome-wide gene expression and growth to better understand the relationship between AOX and nitrate reduction. The results showed that, only when nitrate reduction was limited, AOX deficiency significantly upregulated genes involved in mitochondrial oxidative stress, reductant shuttles, and non-phosphorylating bypasses of the respiratory chain, and inhibited growth. Thus, we conclude that AOX alleviates mitochondrial oxidative stress and sustains plant growth under limited nitrate reduction.

Roles of mitochondrial alternative oxidase in photosynthetic electron transport in illuminated leaves of Arabidopsis thaliana at low temperature

Roles of mitochondrial alternative oxidase in photosynthetic electron transport in illuminated leaves of Arabidopsis thaliana at low temperature

Mitochondrial alternative oxidase pathway helps in nitro-oxidative stress tolerance in germinating chickpea

Mitochondrial alternative oxidase pathway helps in nitro-oxidative stress tolerance in germinating chickpea

Mitochondrial alternative oxidase pathway accelerates non-motile cell germination by enhancing respiratory carbon metabolism and maintaining redox poise in Haematococcus pluvialis

Low efficiency of the cultivation process is a major obstacle in the commercial production of Haematococcus pluvialis. Germination of red, non-motile cells is an efficient strategy for rapid acquisition of zoospores. However, the regulatory mechanisms associated with germination remain unexplored. In the present study, it was confirmed that the mitochondrial alternative oxidase (AOX) pathway accelerates H. pluvialis cell germination, and the regulatory mechanisms were clarified. When the AOX pathway was inhibited, the transcriptomic and metabonomic data revealed a downregulation in respiratory carbon metabolism and nucleotide synthesis due to NADH accumulation. This observation suggested that AOX promoted the rapid consumption of NADH, which accelerated carbohydrate and lipid catabolism, thereby producing carbon skeletons for DNA replication through respiratory metabolism. Moreover, AOX could potentially enhance germination by disturbing the abscisic acid signaling pathway. These findings provide novel insights for developing industrial cultivation models based on red-cell-germination for achieving rapid proliferation of H. pluvialis.

PlantFUNCO: Integrative Functional Genomics Database Reveals Clues into Duplicates Divergence Evolution.

Evolutionary epigenomics and, more generally, evolutionary functional genomics, are emerging fields that study how non-DNA-encoded alterations in gene expression regulation are an important form of plasticity and adaptation. Previous evidence analyzing plants' comparative functional genomics has mostly focused on comparing same assay-matched experiments, missing the power of heterogeneous datasets for conservation inference. To fill this gap, we developed PlantFUN(ctional)CO(nservation) database, which is constituted by several tools and two main resources: interspecies chromatin states and functional genomics conservation scores, presented and analyzed in this work for three well-established plant models (Arabidopsis thaliana, Oryza sativa, and Zea mays). Overall, PlantFUNCO elucidated evolutionary information in terms of cross-species functional agreement. Therefore, providing a new complementary comparative-genomics source for assessing evolutionary studies. To illustrate the potential applications of this database, we replicated two previously published models predicting genetic redundancy in A. thaliana and found that chromatin states are a determinant of paralogs degree of functional divergence. These predictions were validated based on the phenotypes of mitochondrial alternative oxidase knockout mutants under two different stressors. Taking all the above into account, PlantFUNCO aim to leverage data diversity and extrapolate molecular mechanisms findings from different model organisms to determine the extent of functional conservation, thus, deepening our understanding of how plants epigenome and functional noncoding genome have evolved. PlantFUNCO is available at https://rocesv.github.io/PlantFUNCO.

Biomolecular interaction of purified recombinant Arabidopsis thaliana's alternative oxidase 1A with TCA cycle metabolites: Biophysical and molecular docking studies

In higher plants, the mitochondrial alternative oxidase (AOX) pathway plays an essential role in maintaining the TCA cycle/cellular carbon and energy balance under various physiological and stress conditions. Though the activation of AOX pathway upon exogenous addition of α-ketoacids/TCA cycle metabolites [pyruvate, α-ketoglutarate (α-KG), oxaloacetic acid (OAA), succinate and malic acid] to isolated mitochondria is known, the molecular mechanism of interaction of these metabolites with AOX protein is limited. The present study is designed to understand the biomolecular interaction of pure recombinant Arabidopsis thaliana AOX1A with TCA cycle metabolites under in vitro conditions using various biophysical and molecular docking studies. The binding of α-KG, fumaric acid and OAA to rAtAOX1A caused conformational change in the microenvironment of tryptophan residues as evidenced by red shift in the synchronous fluorescence spectra (∆λ = 60 nm). Besides, a decrease in conventional fluorescence emission spectra, tyrosine specific synchronous fluorescence spectra (∆λ = 15 nm) and α-helical content of CD spectra revealed the conformation changes in rAtAOX1A structure associated with binding of various TCA cycle metabolites. Further, surface plasmon resonance (SPR) and microscale thermophoresis (MST) studies revealed the binding affinity, while docking studies identified binding pocket residues, respectively, for these metabolites on rAtAOX1A.

Effect of high light conditions on the response of <i>Arabidopsis thaliana</i> plants with suppressed mitochondrial alternative oxidase

BACKGROUND: Plants as sessile organisms have developed biochemical pathways to protect themselves from the excess light energy. Mitochondrial alternative oxidase (AOX) participates in the oxidation of reductants exported from chloroplasts, thereby optimizing photosynthesis and protecting cells from photodamage.
 AIM: The effect of high light on respiration and the relative transcripts content of a number of genes in Arabidopsis thaliana plants of the T-DNA insertional line for AOX1a (aox1a) was studied and compared with the response of the antisense silencing of AOX1a line (AS-12) and wild type line Col-0.
 MATERIALS AND METHODS: Four-week-old A. thaliana plants of three lines grown at 90 µmol/m2 · s and then exposed to moderately high light conditions, 400 µmol/m2 · s, in a short-term experiment (8 h). Respiratory pathways activity, gene expression, and superoxide anion content were determined during experiment.
 RESULTS: Plants of the aox1a line in response to high light were characterized by the absence of the total and alternative respiration reaction and the absence of the AOX1 protein in spite of the increased mRNA level of AOX1c, in contrast to the Col-0 and AS-12 lines. Also, an increased content of transcripts of only SAPX and CHS were found, while in the other lines a compensatory increase in the expression of many “defense” genes was revealed.
 CONCLUSIONS: Thus, the aox1a line was characterized by a low compensatory effect at the level of defense systems activation. This is apparently caused by the absence of the AOX1 protein and, as a result, the weakening of the stress signal and stress response. The results obtained indicate the important role of AOX in the response of respiration to light stress; can be used to study the signaling pathways of regulation of AOX1a expression.

Alternative oxidase 2 influences Arabidopsis seed germination under salt stress by modulating ABA signalling and ROS homeostasis

Seed germination is a critical checkpoint for plant survival under stresses, which requires functional mitochondria for energy and signal supply. Mitochondrial ALTERNATIVE OXIDASE 2 (AOX2) is specifically and highly expressed at early seed germination; however, its regulatory mechanism remains unknown. Here, the mutant (aox2), over-expression (AOX2OE) and complementation (aox2Comp, proAOX2::gAOX2/aox2) lines were used to investigate AOX2 roles. AOX2 transcript was significantly up-regulated under ABA, NaCl, H2O2, PEG6000 or mannitol treatments during seed germination. Compared with WT and aox2Comp, seed germination and greening were delayed in aox2 under NaCl treatment but enhanced in AOX2OE lines. The transcripts of key genes related to ABA biosynthesis and signal transduction were up-regulated in aox2 compared with those in WT under salt stress. Consequently, the higher ABA level was found in aox2. Na2WO4, the inhibitor of ABA biosynthesis, significantly enhanced germination rate of aox2. Mutation of ABA-INSENSITIVE 3 (ABI3) or ABI4 can partially reverse the salt-sensitive phenotypes of aox2. Yeast one-hybrid, D-luciferin enzyme (LUC) assay and transient expression experiments further revealed ABI3 and ABI4 can directly interact with AOX2 promoter and enhance its expression. Reactive oxygen species (ROS), mainly produced from the root tip mitochondria of germinated seeds, were accumulated more in aox2 than those in WT. Exogenous ascorbic acid (AsA) decreased ROS level and fully rescued the salt-hypersensitive phenotypes of aox2. Taken together, AOX2 plays an important role in Arabidopsis seed germination through regulating ABA signal and ROS homeostasis under salt stress; in the process, ABI3/ABI4 is indispensable for salinity-induced AOX2 expression.

Mitochondrial alternative oxidase across the tree of life: Presence, absence, and putative cases of lateral gene transfer

The alternative oxidase (AOX) is a terminal oxidase in the electron transport system that plays a role in mitochondrial bioenergetics. The past 20 years of research shows AOX has a wide yet patchy distribution across the tree of life. AOX has been suggested to have a role in stress tolerance, growth, and development in plants, but less is known about its function in other groups, including animals. In this study, we analyzed the taxonomic distribution of AOX across >2800 species representatives from prokaryotes and eukaryotes and developed a standardized workflow for finding and verifying the authenticity of AOX sequences. We found that AOX is limited to proteobacteria among prokaryotes, but is widely distributed in eukaryotes, with the highest prevalence in plants, fungi, and protists. AOX is present in many invertebrates, but is absent in others including most arthropods, and is absent from vertebrates. We found aberrant AOX sequences associated with some animal groups. Some of these aberrant AOXs were contaminants, but we also found putative cases of lateral gene transfer of AOX from fungi and protists to nematodes, springtails, fungus gnats, and rotifers. Our findings provide a robust and detailed analysis of the distribution of AOX and a method for identifying and verifying putative AOX sequences, which will be useful as more sequence data becomes available on public repositories.

Effects of OsAOX1a Deficiency on Mitochondrial Metabolism at Critical Node of Seed Viability in Rice.

Mitochondrial alternative oxidase 1a (AOX1a) plays an extremely important role in the critical node of seed viability during storage. However, the regulatory mechanism is still poorly understood. The aim of this study was to identify the regulatory mechanisms by comparing OsAOX1a-RNAi and wild-type (WT) rice seed during artificial aging treatment. Weight gain and time for the seed germination percentage decreased to 50% (P50) in OsAOX1a-RNAi rice seed, indicating possible impairment in seed development and storability. Compared to WT seeds at 100%, 90%, 80%, and 70% germination, the NADH- and succinate-dependent O2 consumption, the activity of mitochondrial malate dehydrogenase, and ATP contents all decreased in the OsAOX1a-RNAi seeds, indicating that mitochondrial status in the OsAOX1a-RNAi seeds after imbibition was weaker than in the WT seeds. In addition, the reduction in the abundance of Complex I subunits showed that the capacity of the mitochondrial electron transfer chain was significantly inhibited in the OsAOX1a-RNAi seeds at the critical node of seed viability. The results indicate that ATP production was impaired in the OsAOX1a-RNAi seeds during aging. Therefore, we conclude that mitochondrial metabolism and alternative pathways were severely inhibited in the OsAOX1a-RNAi seeds at critical node of viability, which could accelerate the collapse of seed viability. The precise regulatory mechanism of the alternative pathway at the critical node of viability needs to be further analyzed. This finding might provide the basis for developing monitoring and warning indicators when seed viability declines to the critical node during storage.

Photosystems under high light stress: throwing light on mechanism and adaptation

High light stress decreases the photosynthetic rate in plants due to photooxidative damage to photosynthetic apparatus, photoinhibition of PSII, and/or damage to PSI. The dissipation of excess energy by nonphotochemical quenching and degradation of the D1 protein of PSII and its repair cycle help against photooxidative damage. Light stress also activates stress-responsive nuclear genes through the accumulation of phosphonucleotide-3'-phosphoadenosine- 5'-phosphate, methylerythritol cyclodiphosphate, and reactive oxygen species which comprise the chloroplast retrograde signaling pathway. Additionally, hormones, such as abscisic acid, cytokinin, brassinosteroids, and gibberellins, play a role in acclimation to light fluctuations. Several alternate electron flow mechanisms, which offset the excess of electrons, include activation of plastid or plastoquinol terminal oxidase, cytochrome b6/f complex, cyclic electron flow through PSI, Mehler ascorbate peroxidase pathway or water-water cycle, mitochondrial alternative oxidase pathway, and photorespiration. In this review, we provided insights into high light stress-mediated damage to photosynthetic apparatus and strategies to mitigate the damage by decreasing antennae size, enhancing NPQ through the introduction of mutants, expression of algal proteins to improve photosynthetic rates and engineering ATP synthase.

Limiting etioplast gene expression induces apical hook twisting during skotomorphogenesis of Arabidopsis seedlings.

When covered by a layer of soil, seedling development follows a dark-specific program (skotomorphogenesis). In the dark, seedlings consist of small, non-green cotyledons, a long hypocotyl, and an apical hook to protect meristematic cells. We recently highlighted the role played by mitochondria in the high energy-consuming reprogramming of Arabidopsis skotomorphogenesis. Here, the role played by plastids, another energy-supplying organelle, in skotomorphogenesis is investigated. This study was conducted in dark conditions to exclude light signals so as to better focus on those produced by plastids. It was found that limitation of plastid gene expression (PGE) induced an exaggerated apical hook bending. Inhibition of PGE was obtained at the levels of transcription and translation using the antibiotics rifampicin (RIF) and spectinomycin, respectively, as well as plastid RPOTp RNA polymerase mutants. RIF-treated seedlings also showed expression induction of marker nuclear genes for mitochondrial stress, perturbation of mitochondrial metabolism, increased ROS levels, and an augmented capacity of oxygen consumption by mitochondrial alternative oxidases (AOXs). AOXs act to prevent overreduction of the mitochondrial electron transport chain. Previously, we reported that AOX1A, the main AOX isoform, is a key component in the developmental response to mitochondrial respiration deficiency. In this work, we suggest the involvement of AOX1A in the response to PGE dysfunction and propose the importance of signaling between plastids and mitochondria. Finally, it was found that seedling architecture reprogramming in response to RIF was independent of canonical organelle retrograde pathways and the ethylene signaling pathway.

Changes in flower volatiles and analysis of alternative oxidase and ultrastructure in Dendrobium huoshanense induced by snail (Bradybaena ravida) infestation

Flower volatiles, as a class of signals that plants communicate with the outside world, are of great significance to the biological control of pests and plant breeding. To explore the biological control methods of snails and accumulate data for the selection and breeding of snail-resistant Dendrobium huoshanense varieties, this study analyzed characteristic changes in D. huoshanense flower volatiles, mitochondrial alternative oxidase (AOX), and ultrastructure under snail (Bradybaena ravida) infestation stress. The results showed that the volatile components of flowers induced by B. ravida infestation changed significantly. Variation of volatile components was mainly caused by five compounds, dipropyl disulfide, 1,8-cineol, β-elemene, β-ocimene, and β-caryophyllene. These compounds had significant repellent or luring effects on snails. AOX activity and AOX protein content also increased significantly after B. ravida infestation. The transcript levels of the AOX1a and AOX1b genes were significantly higher than those of healthy flowers (more than 1.8 folds). Moreover, ultrastructural indicators such as the size of intercellular space of epidermal cells and depth of depression on the cell surface, stomata area, mitochondrial volume, and the number of mitochondria and osmiophilic globules increased significantly. Pearson correlation analysis showed that AOX and ultrastructure were highly correlated with the above five compounds. The study clarified the change characteristics of volatiles in D. Huoshanense flowers induced by B. ravida infestation and preliminarily revealed the relation between flower volatile emissions and changes in AOX and ultrastructure. In the future, we need to strengthen the research on the molecular mechanism of flower volatile signal transmission and transduction.

Mitochondrial alternative oxidase enhanced ABA-mediated drought tolerance in Solanum lycopersicum

The phytohormone abscisic acid (ABA) plays essential roles in modulating drought stress responses. Mitochondrial alternative oxidase (AOX) is critical for reactive oxygen species (ROS) scavenging in drought stress responses. However, whether ABA signal in concert with AOX to moderate drought stress response remains largely unclear. In our study, we uncover the positive role of AOX in ABA-mediated drought tolerance in tomato (Solanum lycopersicum). Here, we report that ABA participates in the regulation of alternative respiration, and the increased AOX was found to improve drought tolerance by reducing total ROS accumulation. We also found that transcription factor ABA response element-binding factor 1 (SlAREB1) can directly bind to the promoter of AOX1a to activate its transcription. Virus-induced gene silencing (VIGS) of SlAREB1 compromised the ABA-induced alternative respiratory pathway, disrupted redox homeostasis and decreased plant resistance to drought stress, while overexpression of AOX1a in TRV2-SlAREB1 plants partially rescued the severe drought phenotype. Taken together, our results indicated that AOX1a plays an essential role in ABA-mediated drought tolerance partially in a SlAREB1-dependent manner, providing new insights into how ABA modulates ROS levels to cope with drought stress by AOX.

Identification of Mitochondrial Alternative Oxidase Genes in Lupinus luteus and the Effect of Salicylic Acid on Their Expression

Identification of Mitochondrial Alternative Oxidase Genes in Lupinus luteus and the Effect of Salicylic Acid on Their Expression

The Mitochondrial Alternative Oxidase in Ustilago maydis Is Not Involved in Response to Oxidative Stress Induced by Paraquat.

It has been shown that the alternative oxidase in mitochondria of fungi and plants has important functions in the response against stress conditions, although their role in some organisms is still unknown. This is the case of Ustilago maydis. There is no evidence of the participation of the U. maydis Aox1 in stressful conditions such as desiccation, high or low temperature, and low pH, among others. Therefore, in this work, we studied the role of the U. maydis Aox1 in cells exposed to oxidative stress induced by methyl viologen (paraquat). To gain insights into the role of this enzyme, we took advantage of four strains: the FB2 wild-type, a strain without the alternative oxidase (FB2aox1Δ), other with the Aox1 fused to the Gfp under the control of the original promoter (FB2aox1-Gfp), and one expressing constitutively de Aox1-Gfp (FB2Potef:aox1-Gfp). Cells were incubated for various times in the presence of 1 mM paraquat and growth, replicative capacities, mitochondrial respiratory activity, Aox1 capacity, and the activities of several antioxidant enzymes (catalase, glutathione peroxidase, glutathione reductase, and superoxide dismutase) were assayed. The results show that (1) the response of U. maydis against oxidative stress was the same in the presence or absence of the Aox1; (2) the activities of the antioxidant enzymes remained constant despite the oxidative stress; and (3) there was a decrease in the GSH/GSSG ratio in U. maydis cells incubated with paraquat.

Suppression of mitochondrial alternative oxidase can result in upregulation of the ROS scavenging network: some possible mechanisms underlying the compensation effect.

Mitochondrial alternative oxidase is an important protein involved in maintaining cellular metabolic and energy balance, especially under stress conditions. AOX genes knockout is aimed at revealing the functions of AOX genes. Under unfavourable conditions, AOX-suppressed plants (mainly based on Arabidopsis AOX1a-knockout lines) usually experience strong oxidative stress. However, a compensation effect, which consists of the absence of AOX1a leading to an increase in defence response mechanisms, concomitant with a decrease in ROS content, has also been demonstrated. This review briefly describes the possible mechanisms underlying the compensation effect upon the suppression of AOX1a. Information about mitochondrial retrograde regulation of AOX is given. The importance of ROS and mitochondrial membrane potential in triggering the signal transmission from mitochondria in the absence of AOX or disturbance of mitochondrial electron transport chain functions is indicated. The few available data on the response of the cell to the absence of AOX at the level of changes in the hormonal balance and the reactions of chloroplasts are presented. The decrease in the relative amount of reduced ascorbate at stable ROS levels as a result of compensation in AOX1a-suppressed plants is proposed as a sign of stress development. Obtaining direct evidence on the mechanisms and signalling pathways involved in AOX modulation in the genome should facilitate a deeper understanding of the role of AOX in the integration of cellular signalling pathways.

Evidence that mitochondrial alternative oxidase respiration supports carbon balance in source leaves of Nicotiana tabacum

Alternative oxidase (AOX) represents a non-energy conserving pathway within the mitochondrial electron transport chain. One potential physiological role of AOX could be to manage leaf carbohydrate amounts by supporting respiratory carbon oxidation reactions. In this study, several approaches tested the hypothesis that AOX1a gene expression in Nicotiana tabacum leaf is enhanced in conditions expected to promote an increased leaf carbohydrate status. These approaches included supplying leaves with exogenous carbohydrates, comparing plants grown at different atmospheric CO2 concentrations, comparing sink leaves with source leaves, comparing plants with different ratios of source to sink activity, and examining gene expression over the diel cycle. In each case, the pattern of AOX1a gene expression was compared with that of other genes known to respond to carbohydrates and/or other factors related to source:sink activity. These included GPT1 and GPT3 (that encode chloroplast glucose 6-phosphate/phosphate translocators), SPS (that encodes sucrose phosphate synthase), SUT1 (that encodes a sucrose/H+ symporter involved in phloem loading) and UCP1 (that encodes a mitochondrial uncoupling protein). The AOX1a transcript amount was higher following the leaf sink-to-source transition, and in plants with higher source relative to sink activity due to increasing plant age. Further, these effects were amplified in plants grown at elevated CO2 to stimulate source activity, particularly at end-of-day time periods. The AOX1a transcript amount was also higher following treatment of leaves with carbohydrate, in particular sucrose. Overall, the results provide evidence that, while source leaf sucrose accumulation may signal for a down-regulation of sucrose synthesis and transport, it also signals for means to manage the excess cytosolic carbohydrate pools. This includes increased AOX respiration to support carbon oxidation pathways even if energy charge is high, in combination perhaps with some return flux of carbohydrate from cytosol to stroma through the GPT3 translocator. As discussed, these activities could contribute to maintaining plant source:sink balance, as well as photosynthetic and phloem loading capacity.

Genome-wide identification of AOX family genes in Moso bamboo and functional analysis of PeAOX1b_2 in drought and salinity stress tolerance.

Five PeAOX genes from Moso bamboo genome were identified. PeAOX1b_2-OE improved tolerance to drought and salinity stress in Arabidopsis, indicating it is involved in positive regulation of abiotic stress response. Mitochondrial alternative oxidase (AOX), the important respiratory terminal oxidase in organisms, catalyzes the energy wasteful cyanide (CN)-resistant respiration, which can improve abiotic stresses tolerance and is considered as one of the functional markers for plant resistance breeding. Here, a total of five putative AOX genes (PeAOXs) were identified and characterized in a monocotyledonous woody grass Moso bamboo (Phyllostachys edulis). Phylogenetic analysis revealed that PeAOXs belonged to AOX1 subfamily, and were named PeAOX1a_1, PeAOX1a_2, PeAOX1b_1, PeAOX1b_2 and PeAOX1c, respectively. Evolutionary and divergence patterns analysis revealed that the PeAOX, OsAOX, and BdAOX families experienced positive purifying selection and may have undergone a large-scale duplication event roughly 1.35-155.90 million years ago. Additionally, the organ-specific expression analysis showed that 80% of PeAOX members were mainly expressed in leaf. Promoter sequence analysis of PeAOXs revealed cis-acting regulatory elements (CAREs) responding to abiotic stress. Most PeAOX genes were significantly upregulated after methyl jasmonate (MeJA) and abscisic acid (ABA) treatment. Moreover, under salinity and drought stresses, the ectopic overexpression of PeAOX1b_2 in Arabidopsis enhanced seed germination and seedling establishment, increased the total respiratory rate and the proportion of AOX respiratory pathway in leaf, and enhanced antioxidant ability, suggesting that PeAOX1b_2 is crucial for abiotic stress resistance in Moso bamboo.

Upregulation of the mitochondrial alternative oxidase pathway improves PSII function and photosynthetic electron transport in tomato seedlings under chilling stress

The aim of this study was to explore how the mitochondrial alternative oxidase (AOX) pathway alleviates photoinhibition in chilled tomato (Solanum lycopersicum) seedlings. Chilling induced photoinhibition in tomato seedlings despite the increases in thermal energy dissipation and cyclic electron flow around PSI (CEF-PSI). Chilling inhibited the function of PSII and blocked electron transport at the PSII acceptor side, however, it did not affect the oxygen-evolving complex on the donor side of PSII. Upregulation of the AOX pathway protects against photoinhibition by improving PSII function and photosynthetic electron transport in tomato seedlings under chilling stress. The AOX pathway maintained the open state of PSII and the stability of the entire photosynthetic electron transport chain. Moreover, the protective role of the AOX pathway on PSII was more important than that on PSI. However, inhibition of the AOX pathway could be compensated by increasing CEF-PSI activity under chilling stress.