Centro de Estudios Fotosintéticos y Bioquímicos

The respiratory pathways of glycolysis, the tricarboxylic acid (TCA) cycle, and mitochondrial electron transport chain (miETC) are central features of carbon metabolism and bioenergetics in aerobic organisms. Respiration is essential for growth, maintenance, and carbon balance of all plant cells. Although the majority of respiratory enzymes are common to all organisms, plant respiration has evolved as a complex metabolic network endowed with a wide variety of unique characteristics. Plants have the option of employing alternative enzymes that bypass several of the conventional steps in cytosolic glycolysis, the TCA cycle, and miETC. The extent and conditions under which these bypasses operate is the subject of intensive research. The highly flexible nature of respiratory metabolism in plants has likely evolved in response to the crucial biosynthetic role played by respiration beyond its role in ATP generation; both functions must proceed if plants are to survive under varying and often stressful environmental and nutritional conditions. Additional complexity arises due to the existence of tissue- and/or developmental-specific isozymes of many plant respiratory enzymes, as well as the extensive interactions between photosynthesis and respiration, and plastidic, cytosolic, and mitochondrial metabolism in general. Recent progress in biochemistry, physiology, cell biology, genomics, transcriptomics, proteomics, metabolomics, and in vivo flux analyses have resulted in exciting new insights into many aspects of plant respiratory metabolism. Experiments on transgenic or mutant plants possessing significantly elevated or reduced levels of respiratory enzymes are augmenting our understanding of the functions, organization, and control of plant respiration. Metabolic engineering of plant respiration is of significant practical interest as it provides both an important approach to enhancing crop yields, as well as a potential mechanism for mitigating global climate change due to elevated atmospheric CO 2 levels.

The genus Candida includes about 200 different species, but only a few species are human opportunistic pathogens and cause infections when the host becomes debilitated or immunocompromised. Candida infections can be superficial or invasive. Superficial infections often affect the skin or mucous membranes and can be treated successfully with topical antifungal drugs. However, invasive fungal infections are often life-threatening, probably due to inefficient diagnostic methods and inappropriate initial antifungal therapies. Here, we briefly review our current knowledge of pathogenic species of the genus Candida and yeast infection causes and then focus on current antifungal drugs and resistance mechanisms. An overview of new therapeutic alternatives for the treatment of Candida infections is also provided.

Flavonoids are plant specialized metabolites that consist of one oxygenated and two aromatic rings. Different flavonoids are grouped according to the oxidation degree of the carbon rings; they can later be modified by glycosylations, hydroxylations, acylations, methylations, or prenylations. These modifications generate a wide collection of different molecules which have various functions in plants. All flavonoids absorb in the UV wavelengths, they mostly accumulate in the epidermis of plant cells and their biosynthesis is generally activated after UV exposure. Therefore, they have been assumed to protect plants against exposure to radiation in this range. Some flavonoids also absorb in other wavelengths, for example anthocyanins, which absorb light in the visible part of the solar spectrum. Besides, some flavonoids show antioxidant properties, that is, they act as scavengers of reactive oxygen species that could be produced after high fluence UV exposure. However, to date most reports were based on in vitro studies, and there is very little in vivo evidence of how their roles are carried out. In this review we first summarize the biosynthetic pathway of flavonoids and their characteristics, and we describe recent advances on the investigation of the role of three of the most abundant flavonoids: flavonols, flavones, and anthocyanins, protecting plants against UV exposure and high light exposure. We also present examples of how using UV-B supplementation to increase flavonoid content, is possible to improve plant nutritional and pharmaceutical values.

Fruit from rosaceous species collectively display a great variety of flavors and textures as well as a generally high content of nutritionally beneficial metabolites. However, relatively little analysis of metabolic networks in rosaceous fruit has been reported. Among rosaceous species, peach (Prunus persica) has stone fruits composed of a juicy mesocarp and lignified endocarp. Here, peach mesocarp metabolic networks were studied across development using metabolomics and analysis of key regulatory enzymes. Principal component analysis of peach metabolic composition revealed clear metabolic shifts from early through late development stages and subsequently during postharvest ripening. Early developmental stages were characterized by a substantial decrease in protein abundance and high levels of bioactive polyphenols and amino acids, which are substrates for the phenylpropanoid and lignin pathways during stone hardening. Sucrose levels showed a large increase during development, reflecting translocation from the leaf, while the importance of galactinol and raffinose is also inferred. Our study further suggests that posttranscriptional mechanisms are key for metabolic regulation at early stages. In contrast to early developmental stages, a decrease in amino acid levels is coupled to an induction of transcripts encoding amino acid and organic acid catabolic enzymes during ripening. These data are consistent with the mobilization of amino acids to support respiration. In addition, sucrose cycling, suggested by the parallel increase of transcripts encoding sucrose degradative and synthetic enzymes, appears to operate during postharvest ripening. When taken together, these data highlight singular metabolic programs for peach development and may allow the identification of key factors related to agronomic traits of this important crop species.

Peach (Prunus persica L. Batsch) is a climacteric fruit that ripens after harvest, prior to human consumption. Organic acids and soluble sugars contribute to the overall organoleptic quality of fresh peach; thus, the integrated study of the metabolic pathways controlling the levels of these compounds is of great relevance. Therefore, in this work, several metabolites and enzymes involved in carbon metabolism were analysed during the post-harvest ripening of peach fruit cv 'Dixiland'. Depending on the enzyme studied, activity, protein level by western blot, or transcript level by quantitative real time-PCR were analysed. Even though sorbitol did not accumulate at a high level in relation to sucrose at harvest, it was rapidly consumed once the fruit was separated from the tree. During the ripening process, sucrose degradation was accompanied by an increase of glucose and fructose. Specific transcripts encoding neutral invertases (NIs) were up-regulated or down-regulated, indicating differential functions for each putative NI isoform. Phosphoenolpyruvate carboxylase was markedly induced, and may participate as a glycolytic shunt, since the malate level did not increase during post-harvest ripening. The fermentative pathway was highly induced, with increases in both the acetaldehyde level and the enzymes involved in this process. In addition, proteins differentially expressed during the post-harvest ripening process were also analysed. Overall, the present study identified enzymes and pathways operating during the post-harvest ripening of peach fruit, which may contribute to further identification of varieties with altered levels of enzymes/metabolites or in the evaluation of post-harvest treatments to produce fruit of better organoleptic attributes.

Drought is the most important crop yield-limiting factor, and detailed knowledge of its impact on plant growth regulation is crucial. The maize (Zea mays) leaf growth zone offers unique possibilities for studying the spatiotemporal regulation of developmental processes by transcriptional analyses and methods that require more material, such as metabolite and enzyme activity measurements. By means of a kinematic analysis, we show that drought inhibits maize leaf growth by inhibiting cell division in the meristem and cell expansion in the elongation zone. Through a microarray study, we observed the down-regulation of 32 of the 54 cell cycle genes, providing a basis for the inhibited cell division. We also found evidence for an up-regulation of the photosynthetic machinery and the antioxidant and redox systems. This was confirmed by increased chlorophyll content in mature cells and increased activity of antioxidant enzymes and metabolite levels across the growth zone, respectively. We demonstrate the functional significance of the identified transcriptional reprogramming by showing that increasing the antioxidant capacity in the proliferation zone, by overexpression of the Arabidopsis (Arabidopsis thaliana) iron-superoxide dismutase gene, increases leaf growth rate by stimulating cell division. We also show that the increased photosynthetic capacity leads to enhanced photosynthesis upon rewatering, facilitating the often-observed growth compensation.

NADP-malic enzyme (NADP-ME) is a widely distributed enzyme that catalyzes the oxidative decarboxylation of L-malate. Photosynthetic NADP-MEs are found in C4 bundle sheath chloroplasts and in the cytosol of CAM plants, while non-photosynthetic NADP-MEs are either plastidic or cytosolic in various plants. We propose a classification of plant NADP-MEs based on their physiological function and localization and we describe recent advances in the characterization of each isoform. Based on the alignment of amino acid sequences of plant NADP-MEs, we identify putative binding sites for the substrates and analyze the phylogenetic origin of each isoform, revealing several features of the molecular evolution of this ubiquitous enzyme.

A compensation for differences in bone material quality by bone geometric properties in femora from two different strains of rats was previously shown by us. A feedback mechanism controlling the mechanical properties of the integrated bones was then proposed, in accordance with Frost's mechanostat theory. Evidence of such a system is now offered by the finding of a negative correlation between the modeling-dependent cross-sectional architecture (moment of inertia) and the mineral-dependent stiffness (elastic modulus) of bone material in the femoral diaphyses of 45 normal Wistar rats of different sexes, ages, and sizes. The strength and stiffness of the integrated diaphyses were found to depend on both cross-sectional inertia and body weight, not on bone mineral density. These findings are interpreted as supporting the hypothesis that the architectural efficiency of diaphyseal cross-sectional design resulting from the spatial orientation of bone modeling during growth is optimized as a function of the body weight-dependent bone strain history, within the constraints imposed by bone stiffness. Results suggest a modulating role of biomass, related to the system set point determination, and explain the usually observed lack of a direct correlation between mineral density and strength or stiffness of long bones in studies of geometrically inhomogeneous populations.

The Arabidopsis (Arabidopsis thaliana) genome contains four genes encoding putative NADP-malic enzymes (MEs; AtNADP-ME1-ME4). NADP-ME4 is localized to plastids, whereas the other three isoforms do not possess any predicted organellar targeting sequence and are therefore expected to be cytosolic. The plant NADP-MEs can be classified into four groups: groups I and II comprising cytosolic and plastidic isoforms from dicots, respectively; group III containing isoforms from monocots; and group IV composed of both monocots and dicots, including AtNADP-ME1. AtNADP-MEs contained all conserved motifs common to plant NADP-MEs and the recombinant isozymes showed different kinetic and structural properties. NADP-ME2 exhibits the highest specific activity, while NADP-ME3 and NADP-ME4 present the highest catalytic efficiency for NADP and malate, respectively. NADP-ME4 exists in equilibrium of active dimers and tetramers, while the cytosolic counterparts are present as hexamers or octamers. Characterization of T-DNA insertion mutant and promoter activity studies indicates that NADP-ME2 is responsible for the major part of NADP-ME activity in mature tissues of Arabidopsis. Whereas NADP-ME2 and -ME4 are constitutively expressed, the expression of NADP-ME1 and NADP-ME3 is restricted by both developmental and cell-specific signals. These isoforms may play specific roles at particular developmental stages of the plant rather than being involved in primary metabolism.

Pericarp Color1 (P1) encodes an R2R3-MYB transcription factor responsible for the accumulation of insecticidal flavones in maize (Zea mays) silks and red phlobaphene pigments in pericarps and other floral tissues, which makes P1 an important visual marker. Using genome-wide expression analyses (RNA sequencing) in pericarps and silks of plants with contrasting P1 alleles combined with chromatin immunoprecipitation coupled with high-throughput sequencing, we show here that the regulatory functions of P1 are much broader than the activation of genes corresponding to enzymes in a branch of flavonoid biosynthesis. P1 modulates the expression of several thousand genes, and ∼1500 of them were identified as putative direct targets of P1. Among them, we identified F2H1, corresponding to a P450 enzyme that converts naringenin into 2-hydroxynaringenin, a key branch point in the P1-controlled pathway and the first step in the formation of insecticidal C-glycosyl flavones. Unexpectedly, the binding of P1 to gene regulatory regions can result in both gene activation and repression. Our results indicate that P1 is the major regulator for a set of genes involved in flavonoid biosynthesis and a minor modulator of the expression of a much larger gene set that includes genes involved in primary metabolism and production of other specialized compounds.

sae is a regulatory locus that activates the production of several exoproteins in Staphylococcus aureus. A 3.4-kb fragment of a S. aureus genomic library, screened with a probe adjacent to the transposon insertion of a sae::Tn551 mutant, was cloned into a bifunctional vector. This fragment was shown to carry the sae locus by restoration of exoprotein production in sae mutants. The sae locus was mapped to the SmaI-D fragment of the staphylococcal chromosome by pulse-field electrophoresis. Sequence analysis of the cloned fragment revealed the presence of two genes, designated saeR and saeS, encoding a response regulator and a histidine protein kinase, respectively, with high homology to other bacterial two-component regulatory systems.

Flavonols are important compounds for conditional male fertility in maize (Zea mays) and other crops, and they also contribute to protecting plants from UV-B radiation. However, little continues to be known on how maize and other grasses synthesize flavonols, and how flavonol biosynthesis is regulated. By homology with an Arabidopsis flavonol synthase (AtFLS1), we cloned a maize gene encoding a protein (ZmFLS1) capable of converting the dihydrokaempferol (DHK) and dihydroquercetin (DHQ) dihydroflavonols to the corresponding flavonols, kaempferol (K) and quercetin (Q). Moreover, ZmFLS1 partially complements the flavonol deficiency of the Arabidopsis fls1 mutant, and restores anthocyanin accumulation to normal levels. We demonstrate that ZmFLS1 is under the control of the anthocyanin (C1/PL1 + R/B) and 3-deoxy flavonoid (P1) transcriptional regulators. Indeed, using chromatin immunoprecipitation (ChIP) experiments, we establish that ZmFLS1 is an immediate direct target of the P1 and C1/R regulatory complexes, revealing similar control as for earlier steps in the maize flavonoid pathway. Highlighting the importance of flavonols in UV-B protection, we also show that ZmFLS1 is induced in maize seedlings by UV-B, and that this induction is in part mediated by the increased expression of the P1, B and PL1 regulators. Together, our results identify a key flavonoid biosynthetic enzyme so far missed in maize and other monocots, and illustrate mechanisms by which flavonol accumulation is controlled in maize.

Because of their sessile lifestyle, plants are continuously exposed to solar UV-B radiation. Inhibition of leaf growth is one of the most consistent responses of plants upon exposure to UV-B radiation. In this work, we investigated the role of Growth-Regulating Factors (GRFs) and of microRNA miR396 in UV-B-mediated inhibition of leaf growth in Arabidopsis thaliana plants. We demonstrate that miRNA396 is upregulated by UV-B radiation in proliferating tissues and that this induction is correlated with a decrease in GRF1, GRF2, and GRF3 transcripts. Induction of miR396 results in inhibition of cell proliferation, and this outcome is independent of the UV-B photoreceptor UV resistance locus 8, as well as ATM AND RAD3-related and the mitogen-activated protein kinase MPK6, but is dependent on MPK3. Transgenic plants expressing an artificial target mimic directed against miR396 (MIM396) with a decrease in the endogenous microRNA activity or plants expressing miR396-resistant copies of several GRFs are less sensitive to this inhibition. Consequently, at intensities that can induce DNA damage in Arabidopsis plants, UV-B radiation limits leaf growth by inhibiting cell division in proliferating tissues, a process mediated by miR396 and GRFs.

Antarctica is the coldest, windiest, and driest continent on Earth. In this sense, microorganisms that inhabit Antarctica environments have to be adapted to harsh conditions. Fungal strains affiliated with Ascomycota and Basidiomycota phyla have been recovered from terrestrial and marine Antarctic samples. They have been used for the bioprospecting of molecules, such as enzymes. Many reports have shown that these microorganisms produce cold-adapted enzymes at low or mild temperatures, including hydrolases (e.g. α-amylase, cellulase, chitinase, glucosidase, invertase, lipase, pectinase, phytase, protease, subtilase, tannase, and xylanase) and oxidoreductases (laccase and superoxide dismutase). Most of these enzymes are extracellular and their production in the laboratory has been carried out mainly under submerged culture conditions. Several studies showed that the cold-adapted enzymes exhibit a wide range in optimal pH (1.0-9.0) and temperature (10.0-70.0 °C). A myriad of methods have been applied for cold-adapted enzyme purification, resulting in purification factors and yields ranging from 1.70 to 1568.00-fold and 0.60 to 86.20%, respectively. Additionally, some fungal cold-adapted enzymes have been cloned and expressed in host organisms. Considering the enzyme-producing ability of microorganisms and the properties of cold-adapted enzymes, fungi recovered from Antarctic environments could be a prolific genetic resource for biotechnological processes (industrial and environmental) carried out at low or mild temperatures.

Ribosomal protein L10 (RPL10) proteins are ubiquitous in the plant kingdom. Arabidopsis (Arabidopsis thaliana) has three RPL10 genes encoding RPL10A to RPL10C proteins, while two genes are present in the maize (Zea mays) genome (rpl10-1 and rpl10-2). Maize and Arabidopsis RPL10s are tissue-specific and developmentally regulated, showing high levels of expression in tissues with active cell division. Coimmunoprecipitation experiments indicate that RPL10s in Arabidopsis associate with translation proteins, demonstrating that it is a component of the 80S ribosome. Previously, ultraviolet-B (UV-B) exposure was shown to increase the expression of a number of maize ribosomal protein genes, including rpl10. In this work, we demonstrate that maize rpl10 genes are induced by UV-B while Arabidopsis RPL10s are differentially regulated by this radiation: RPL10A is not UV-B regulated, RPL10B is down-regulated, while RPL10C is up-regulated by UV-B in all organs studied. Characterization of Arabidopsis T-DNA insertional mutants indicates that RPL10 genes are not functionally equivalent. rpl10A and rpl10B mutant plants show different phenotypes: knockout rpl10A mutants are lethal, rpl10A heterozygous plants are deficient in translation under UV-B conditions, and knockdown homozygous rpl10B mutants show abnormal growth. Based on the results described here, RPL10 genes are not redundant and participate in development and translation under UV-B stress.

Although the nonphotosynthetic NAD-malic enzyme (NAD-ME) was assumed to play a central role in the metabolite flux through the tricarboxylic acid cycle, the knowledge on this enzyme is still limited. Here, we report on the identification and characterization of two genes encoding mitochondrial NAD-MEs from Arabidopsis (Arabidopsis thaliana), AtNAD-ME1 and AtNAD-ME2. The encoded proteins can be grouped into the two clades found in the plant NAD-ME phylogenetic tree. AtNAD-ME1 belongs to the clade that includes known alpha-subunits with molecular masses of approximately 65 kD, while AtNAD-ME2 clusters with the known beta-subunits with molecular masses of approximately 58 kD. The separated recombinant proteins showed NAD-ME activity, presented comparable kinetic properties, and are dimers in their active conformation. Native electrophoresis coupled to denaturing electrophoresis revealed that in vivo AtNAD-ME forms a dimer of nonidentical subunits in Arabidopsis. Further support for this conclusion was obtained by reconstitution of the active heterodimer in vitro. The characterization of loss-of-function mutants for both AtNAD-MEs indicated that both proteins also exhibit enzymatic activity in vivo. Neither the single nor the double mutants showed a growth or developmental phenotype, suggesting that NAD-ME activity is not essential for normal autotrophic development. Nevertheless, metabolic profiling of plants completely lacking NAD-ME activity revealed differential patterns of modifications in light and dark periods and indicates a major role for NAD-MEs during nocturnal metabolism.

Flavonoids accumulate in plant vacuoles usually as O-glycosylated derivatives, but several species can also synthesize flavonoid C-glycosides. Recently, we demonstrated that a flavanone 2-hydroxylase (ZmF2H1, CYP93G5) converts flavanones to the corresponding 2-hydroxy derivatives, which are expected to serve as substrates for C-glycosylation. Here, we isolated a cDNA encoding a UDP-dependent glycosyltransferase (UGT708A6), and its activity was characterized by in vitro and in vivo bioconversion assays. In vitro assays using 2-hydroxyflavanones as substrates and in vivo activity assays in yeast co-expressing ZmF2H1 and UGT708A6 show the formation of the flavones C-glycosides. UGT708A6 can also O-glycosylate flavanones in bioconversion assays in Escherichia coli as well as by in vitro assays with the purified recombinant protein. Thus, UGT708A6 is a bifunctional glycosyltransferase that can produce both C- and O-glycosidated flavonoids, a property not previously described for any other glycosyltransferase. Background: Plant UDP-glycosyltransferases add sugars to acceptors like flavonoids either via hydroxyls (O-linkage) or carbons (C-linkage). Results: A maize glycosyltransferase produces both flavonoid C-glycosides and O-glycosides. Conclusion: This is the first description of a bifunctional C-/O-glycosyltransferase with a dual role in nature. Significance: This enzyme might be involved both in the biosynthesis of the natural insecticide maysin and in the formation of O-glycosides.

The Arabidopsis thaliana locus At5g06580 encodes an ortholog to Saccharomyces cerevisiae d-lactate dehydrogenase (AtD-LDH). The recombinant protein is a homodimer of 59-kDa subunits with one FAD per monomer. A substrate screen indicated that AtD-LDH catalyzes the oxidation of d- and l-lactate, d-2-hydroxybutyrate, glycerate, and glycolate using cytochrome c as an electron acceptor. AtD-LDH shows a clear preference for d-lactate, with a catalytic efficiency 200- and 2000-fold higher than that for l-lactate and glycolate, respectively, and a Km value for d-lactate of ∼160 μm. Knock-out mutants showed impaired growth in the presence of d-lactate or methylglyoxal. Collectively, the data indicated that the protein is a d-LDH that participates in planta in the methylglyoxal pathway. Web-based bioinformatic tools revealed the existence of a paralogous protein encoded by locus At4g36400. The recombinant protein is a homodimer of 61-kDa subunits with one FAD per monomer. A substrate screening revealed highly specific d-2-hydroxyglutarate (d-2HG) conversion in the presence of an organic cofactor with a Km value of ∼580 μm. Thus, the enzyme was characterized as a d-2HG dehydrogenase (AtD-2HGDH). Analysis of knock-out mutants demonstrated that AtD-2HGDH is responsible for the total d-2HGDH activity present in A. thaliana. Gene coexpression analysis indicated that AtD-2HGDH is in the same network as several genes involved in β-oxidation and degradation of branched-chain amino acids and chlorophyll. It is proposed that AtD-2HGDH participates in the catabolism of d-2HG most probably during the mobilization of alternative substrates from proteolysis and/or lipid degradation. The Arabidopsis thaliana locus At5g06580 encodes an ortholog to Saccharomyces cerevisiae d-lactate dehydrogenase (AtD-LDH). The recombinant protein is a homodimer of 59-kDa subunits with one FAD per monomer. A substrate screen indicated that AtD-LDH catalyzes the oxidation of d- and l-lactate, d-2-hydroxybutyrate, glycerate, and glycolate using cytochrome c as an electron acceptor. AtD-LDH shows a clear preference for d-lactate, with a catalytic efficiency 200- and 2000-fold higher than that for l-lactate and glycolate, respectively, and a Km value for d-lactate of ∼160 μm. Knock-out mutants showed impaired growth in the presence of d-lactate or methylglyoxal. Collectively, the data indicated that the protein is a d-LDH that participates in planta in the methylglyoxal pathway. Web-based bioinformatic tools revealed the existence of a paralogous protein encoded by locus At4g36400. The recombinant protein is a homodimer of 61-kDa subunits with one FAD per monomer. A substrate screening revealed highly specific d-2-hydroxyglutarate (d-2HG) conversion in the presence of an organic cofactor with a Km value of ∼580 μm. Thus, the enzyme was characterized as a d-2HG dehydrogenase (AtD-2HGDH). Analysis of knock-out mutants demonstrated that AtD-2HGDH is responsible for the total d-2HGDH activity present in A. thaliana. Gene coexpression analysis indicated that AtD-2HGDH is in the same network as several genes involved in β-oxidation and degradation of branched-chain amino acids and chlorophyll. It is proposed that AtD-2HGDH participates in the catabolism of d-2HG most probably during the mobilization of alternative substrates from proteolysis and/or lipid degradation. l- and d-lactate dehydrogenases belong to evolutionarily unrelated enzyme families (1Cristescu M.E. Innes D.J. Stillman J.H. Crease T.J. BMC Evol. Biol. 2008; 8: 268Crossref PubMed Scopus (36) Google Scholar). l-Lactate is oxidized by l-lactate:NAD oxidoreductase (EC 1.1.1.27), which catalyzes the reaction l-lactate + NAD → pyruvate + NADH, and by l-lactate cytochrome c oxidoreductase (l-lactate cytochrome c oxidoreductase, EC 1.1.2.3), which catalyzes the reaction l-lactate + 2 cytochrome c (oxidized) → pyruvate + 2 cytochrome c (reduced). Both groups are found in eubacteria, archebacteria, and eukaryotes. All known plant sequences belong to the EC 1.1.1.27 group (1Cristescu M.E. Innes D.J. Stillman J.H. Crease T.J. BMC Evol. Biol. 2008; 8: 268Crossref PubMed Scopus (36) Google Scholar). On the other hand, d-lactate is oxidized by d-lactate:NAD oxidoreductase (d-lactate:NAD oxidoreductase, EC 1.1.1.28), which catalyzes the reaction d-lactate + NAD → pyruvate + NADH, and by d-lactate cytochrome c oxidoreductase (d-lactate cytochrome c oxidoreductase, EC 1.1.2.4), which catalyzes the reaction d-lactate + 2 cytochrome c (oxidized) → pyruvate + 2 cytochrome c (reduced). Although l-lactate dehydrogenase belongs to the most intensely studied enzyme families (2Lodi T. Guiard B. Mol. Cell. Biol. 1991; 11: 3762-3772Crossref PubMed Scopus (52) Google Scholar, 3Passarella S. de Bari L. Valenti D. Pizzuto R. Paventi G. Atlante A. FEBS Lett. 2008; 582: 3569-3576Crossref PubMed Scopus (125) Google Scholar), our knowledge about the structure, kinetics, and biological function of d-LDH 3The abbreviations used are:d-LDHd-lactate dehydrogenased-2HG,d-2-hydroxyglutarated-2HGDHd-2-hydroxyglutarate dehydrogenaseDCIPdichlorophenolindophenold-LDHd-lactate dehydrogenaseETFelectron transfer proteinETFQOETF-ubiquinone oxidoreductaseMGmethylglyoxalTricineN-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycineNi2+-NTANi2+-nitrilotriacetic acidPMSphenazine methosulfateDCIP2,6-dichlorophenolindophenolMSMurashige and Skoogd-2HBd-2-hydroxybutyrate. is limited. d-LDHs have mainly been identified in prokaryotes and fungi where they play an important role in anaerobic energy metabolism (4Mizushima S. Hiyama T. Kitahara K. J. Gen. Appl. Microbiol. 1964; 10: 33-44Crossref Scopus (7) Google Scholar, 5Garvie E.I. Microbiol. Rev. 1980; 44: 106-139Crossref PubMed Google Scholar, 6Ogata M. Arihara K. Yagi T. J. Biochem. 1981; 89: 1423-1431Crossref PubMed Scopus (37) Google Scholar, 7Brockman H.L. Wood W.A. J. Bacteriol. 1975; 124: 1454-1461Crossref PubMed Google Scholar, 8Lodi T. Ferrero I. Mol. Gen. Genet. 1993; 238: 315-324Crossref PubMed Scopus (69) Google Scholar, 9Reed D.W. Hartzell P.L. J. Bacteriol. 1999; 181: 7580-7587Crossref PubMed Google Scholar, 10Horikiri S. Aizawa Y. Kai T. Amachi S. Shinoyama H. Fujii T. Biosci. Biotechnol. Biochem. 2004; 68: 516-522Crossref PubMed Scopus (17) Google Scholar). In Saccharomyces cerevisiae and Kluyveromyces lactis, a mitochondrial flavoprotein d-lactate ferricytochrome c oxidoreductase (d-lactate cytochrome c oxidoreductase), catalyzing the oxidation of d-lactate to pyruvate, is required for the utilization of d-lactate (8Lodi T. Ferrero I. Mol. Gen. Genet. 1993; 238: 315-324Crossref PubMed Scopus (69) Google Scholar, 11Lodi T. O'Connor D. Goffrini P. Ferrero I. Mol. Gen. Genet. 1994; 244: 622-629Crossref PubMed Scopus (36) Google Scholar). In S. cerevisiae it was suggested that d-LDH is involved in the metabolism of methylglyoxal (MG) (12Labeyrie F. Slonimski P. Bull. Soc. Chim. Biol. 1964; 44: 1793-1828Google Scholar). d-lactate dehydrogenase d-2-hydroxyglutarate d-2-hydroxyglutarate dehydrogenase dichlorophenolindophenol d-lactate dehydrogenase electron transfer protein ETF-ubiquinone oxidoreductase methylglyoxal N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine Ni2+-nitrilotriacetic acid phenazine methosulfate 2,6-dichlorophenolindophenol Murashige and Skoog d-2-hydroxybutyrate. In eukaryotic cells, d-lactate results from the glyoxalase system (13Atlante A. de Bari L. Valenti D. Pizzuto R. Paventi G. Passarella S. Biochim. Biophys. Acta. 2005; 1708: 13-22Crossref PubMed Scopus (25) Google Scholar, 14Thornalley P.J. Biochem. J. 1990; 269: 1-11Crossref PubMed Scopus (690) Google Scholar). This system is the main MG catabolic pathway, comprising the enzymes glyoxalase I (lactoylglutathione lyase, EC 4.4.1.5) and glyoxalase II (hydroxyacylglutathione hydrolase, EC 3.1.2.6). MG (CH3-CO-CHO; in is a as a of from and Lett. 1999; PubMed Scopus Google Scholar), and in is as and Y. J. Biol. PubMed Scopus Google Scholar, M. Biochem. Biophys. 2005; PubMed Scopus Google Scholar). the of glyoxalase I or II was to to in and M. Biochem. Biophys. 2005; PubMed Scopus Google Scholar, A. 2008; PubMed Scopus Google Scholar). It is that the role of the MG pathway, from MG to d-lactate cytochrome c oxidoreductase in the is to in the of it function as an for the acid Lett. 1999; PubMed Scopus Google Scholar). I catalyzes the of from the from MG and glyoxalase II catalyzes the of to and I and II are present in of eukaryotic I is found in the glyoxalase II to the and (13Atlante A. de Bari L. Valenti D. Pizzuto R. Paventi G. Passarella S. Biochim. Biophys. Acta. 2005; 1708: 13-22Crossref PubMed Scopus (25) Google Scholar, S. Mol. Biol. PubMed Scopus Google Scholar, G. S. M. Biochem. 1990; Google Scholar). Although glyoxalase I and II are the of Atlante (13Atlante A. de Bari L. Valenti D. Pizzuto R. Paventi G. Passarella S. Biochim. Biophys. Acta. 2005; 1708: 13-22Crossref PubMed Scopus (25) Google showed that d-lactate by and that was oxidized by a mitochondrial flavoprotein in The of Arabidopsis thaliana the to for genes with the d-LDH from S. cerevisiae an A. thaliana ortholog was In the and and of the recombinant d-LDH from A. thaliana and which was found to a d-2-hydroxyglutarate dehydrogenase is AtD-LDH a substrate and the substrates are d-lactate and d-2-hydroxybutyrate, AtD-2HGDH showed activity with coexpression analysis and analysis of knock-out the of mitochondrial in plant metabolism is was from of using the of total was using the and used for the specific the of the to a the and the of the The used for the as and and and by using the and the for by of for for for 2 and a for The and the and was using the system The with and a The as and the The and with an the of the recombinant The with the in and in the presence of and the an of the of AtD-LDH and was to the and the to for The by for in of and for to The was used for protein using Ni2+-nitrilotriacetic The was and with and The protein was using of and The of the than was about 2 The enzymes used for the using and with for 2 to the encoded by the amino acid The using a to the The used for The of the AtD-LDH and AtD-2HGDH was using a reaction and cytochrome c phenazine methosulfate and dichlorophenolindophenol in a of In the of of glycolate or d-lactate and for AtD-2HGDH (d-2HG) used as by the of cytochrome c with cytochrome the was by 2 2 of cytochrome c are for of substrate A was by the reaction for the of enzyme the substrate The substrate of the enzymes was using substrate and cytochrome c or and as electron as The Km and for the substrates with cytochrome c or and and the of the substrate the of the other All with enzyme and to of enzyme activity is as the that of This to the of 2 of cytochrome c or of The oxidation of glycolate, d- and l-lactate, and d-2HG by NAD or as as the of pyruvate, and by or was as The activity glycolate using was as H. 2008; PubMed Scopus Google Scholar). The of the recombinant by a system using a The was with and using The and the in a of a of The of AtD-LDH and AtD-2HGDH with a in 2 The from the recombinant by for and the protein was by The 2 2 and FAD 2 and in the with A of in with of The same a of an and an The was in using as and as and of and from the Arabidopsis by of The was using the and for or and for or and for or and for The was with the in with or using the and In the of the was using the and A. thaliana and the in and in a growth a and and a of in Murashige and Skoog and in the and of in the presence of of l-lactate, and glycolate was in a growth as the of genes in the total was from of using the was using the in a of using of the and The of used and for and or and for and as of for for and for 2 by the was by using the and from and from from and the in in the presence of of and The by for and to and of the as by P. P. 2005; 124: Scopus Google Scholar). of the mitochondrial was by using the dehydrogenase from which with the and with the mitochondrial one by of was to the of Biochem. PubMed Scopus Google Scholar). was using to PubMed Scopus Google Scholar). with or a for the was with using to the was an was and for activity by in the with and in a of that are with At5g06580 and identified using the data T. S. M. H. K. PubMed Scopus Google and in The which is is used to the of The is as the of the of A to and of to A. The Gene and genes d-LDH the A. thaliana was with the S. cerevisiae d-LDH or protein using the The was a of amino acids encoded by locus This protein a mitochondrial R. A. K. M. R. PubMed Scopus Google Scholar). A in the A. thaliana using the At5g06580 a protein with This acid protein was to to R. A. K. M. R. PubMed Scopus Google and is encoded by locus At4g36400. A using a of sequences of At5g06580 and showed which At5g06580 or and fungi to one of the groups and the analysis shows that and fungi group and The of enzymes in higher that have in The characterized to S. cerevisiae and K. d-LDH activity (8Lodi T. Ferrero I. Mol. Gen. Genet. 1993; 238: 315-324Crossref PubMed Scopus (69) Google Scholar, 10Horikiri S. Aizawa Y. Kai T. Amachi S. Shinoyama H. Fujii T. Biosci. Biotechnol. Biochem. 2004; 68: 516-522Crossref PubMed Scopus (17) Google Scholar, Biochim. Biophys. Acta. PubMed Google Scholar), and At5g06580 was as d-LDHs the of the At5g06580 the sequences are D.W. Hartzell P.L. J. Bacteriol. 1999; 181: 7580-7587Crossref PubMed Google Scholar, Biochim. Biophys. Acta. PubMed Scopus Google Scholar). On the other hand, to have been The enzymes from and d-2-hydroxyglutarate dehydrogenase activity Y. G. D. M. Biochem. J. 2004; PubMed Scopus Google Scholar), the from S. cerevisiae and have d-LDH activity A. Y. D. 1999; PubMed Scopus Google Scholar). and have been with and that a substrate for was as Analysis of the of AtD-LDH and AtD-2HGDH the existence of mitochondrial sequences of and amino acid in R. A. K. M. R. PubMed Scopus Google Scholar). the of the catalytic of the amino acid and respectively, A and the AtD-LDH and AtD-2HGDH in the and in of and and to AtD-LDH and AtD-2HGDH protein respectively, a encoded by the the present in the of in from A and to by A and with of of protein per of The by using the indicated with the same as by and revealed with the of The enzymes and 2 of in a in a of The AtD-LDH and AtD-2HGDH that a cofactor was with the The revealed and which of the FAD and and are as for a the of the the enzymes and the by The the by and by using and as the of the enzyme of enzymes highly to of FAD of by and in a using the cofactor from of AtD-LDH and AtD-2HGDH in a of the was that the of FAD is and the of the recombinant AtD-LDH and AtD-2HGDH are and respectively, it that of enzyme of In indicated of and for AtD-LDH and respectively, that the enzymes The substrate of AtD-LDH was using a of as substrates and or cytochrome c as The activity was found with cytochrome for which Km of and using d- and l-lactate as a of cytochrome the catalytic with and d-lactate, with l-lactate, and glycolate and that with d-lactate, The for l-lactate and was with Km of and are and higher than that with d-lactate, The for d-lactate, and glycolate with Km of and In AtD-LDH a catalytic efficiency with d-lactate using cytochrome c as about 200- and 2000-fold higher preference for d-lactate with l-lactate and glycolate, A was using the in electron AtD-LDH showed the catalytic efficiency and activity with and activity was using NAD or as electron and function as the for glycolate catalytic with the recombinant enzyme of the encoded by the A of the showed that AtD-LDH was to the of of and pyruvate in the presence of or in the of the reaction with d-lactate and cytochrome of recombinant AtD-LDH using substrates and cytochrome c as electron in a of recombinant AtD-LDH and cytochrome in a The for d-lactate oxidation a The same results using l-lactate and glycolate as substrates The enzyme activity to it activity A substrate screening to that for AtD-LDH was with the AtD-2HGDH was highly specific and the oxidation of d-2HG using as electron with a Km of for the substrate The enzyme a activity d-lactate, and with was to the that using cytochrome c as electron the activity with d-2HG was of that using and the enzyme showed d-lactate, and as with that in AtD-2HGDH the to an electron other than cytochrome The enzyme glycolate as a substrate with of the AtD-2HGDH was to the oxidation of d-2HG using NAD or the of using or which was with d-2HG and was the of results with the recombinant enzyme of the The recombinant AtD-LDH and AtD-2HGDH by and for activity using substrates and as an electron acceptor. In with the in AtD-LDH showed an activity the with d-lactate and for the was for a was with l-lactate and glycolate as substrates On the other hand, AtD-2HGDH showed an with d-2HG activity was with d-lactate and In the activity with as by analysis A. thaliana and and of by and for activity using the same substrates as the activity of AtD-LDH by in the the recombinant enzyme activity it is that the protein activity during the of It is that to the of of AtD-LDH of growth D. B. H. R. PubMed Scopus Google Scholar), the specific activity of the is the of the A with of that by the recombinant AtD-2HGDH was in and and of using d-2HG as a substrate of in the of the a of activity with d-lactate and as substrates Thus, results demonstrated that the recombinant enzyme the same substrate and catalytic as the enzyme present in the plant It is that d-2HGDH activity was in the mitochondrial in with the results with the that the mitochondrial with that and dehydrogenases to a mitochondrial of the and the substrate in with found in and for for by and and using to the used to the in the AtD-LDH in the and in and On the other hand, AtD-2HGDH in the and in the from and and mitochondrial of the knock-out by showed with d-2HG d-lactate, or The same results with of the knock-out that locus encodes a which is responsible for the total d-2HGDH activity in A. thaliana. growth or the knock-out showed in or with the the of d-lactate and MG the growth of A. thaliana and the knock-out mutants with of the the in the presence of d-lactate to or MG to was in and higher d-lactate or MG and to the and It is that in the presence of MG and d-lactate and than in that MG and d-lactate a which plant results that in AtD-LDH is to MG a higher in impaired the of d-LDH activity in The of A. thaliana locus At5g06580 encodes a of that a FAD group as by and with the protein and a The enzyme is as a in to the from which of subunits of S. Aizawa Y. Kai T. Amachi S. Shinoyama H. Fujii T. Biosci. Biotechnol. Biochem. 2004; 68: 516-522Crossref PubMed Scopus (17) Google Scholar). The data that the A. thaliana protein as a d-LDH The enzyme l-lactate, and glycolate, with catalytic efficiency to a l-lactate and and an catalytic glycolate the AtD-LDH showed the activity with and the one with glycolate activity d-lactate and and or activity with l-lactate and glycolate, of characterized d-LDHs from S. R. and H.L. Wood W.A. J. Bacteriol. 1975; 124: 1454-1461Crossref PubMed Google Scholar, 10Horikiri S. Aizawa Y. Kai T. Amachi S. Shinoyama H. Fujii T. Biosci. Biotechnol. Biochem. 2004; 68: 516-522Crossref PubMed Scopus (17) Google Scholar, Biochim. Biophys. Acta. PubMed Google Scholar). In is in as a during catabolism J. PubMed Scopus Google or J. S. M. L. M. H. H. R. J. and and and is a in K. Lett. Appl. Microbiol. PubMed Scopus (7) Google Scholar). our are about the biological function of in plant In the activity of AtD-LDH substrate a of a substrate and a reaction of AtD-LDH showed an catalytic glycolate, it that it is involved in glycolate metabolism in as the of in the of in mutants of the subunits of glycolate the of glycolate oxidation R. R. R. T. J. 2004; PubMed Scopus Google Scholar), that the enzyme to glycolate, of substrate are present as in the same was in and the in with glycolate to that AtD-LDH involved in the of glycolate in The screening of as substrates to that AtD-LDH is of acids the results that At5g06580 the EC group cytochrome The enzyme was the EC group R. R. R. T. J. 2004; PubMed Scopus Google Scholar). It is that group with or to our knowledge the have been In and d-lactate is from MG by the of glyoxalase I and glyoxalase In A. glyoxalase I and glyoxalase II are I is found in the P.J. Biochem. J. 1990; 269: 1-11Crossref PubMed Scopus (690) Google Scholar), and glyoxalase II is to the and (13Atlante A. de Bari L. Valenti D. Pizzuto R. Paventi G. Passarella S. Biochim. Biophys. Acta. 2005; 1708: 13-22Crossref PubMed Scopus (25) Google Scholar, S. Mol. Biol. PubMed Scopus Google Scholar), and AtD-LDH been to R. R. R. T. J. 2004; PubMed Scopus Google Scholar). In d-lactate the of AtD-LDH from the In H. to d-lactate by of a or a (13Atlante A. de Bari L. Valenti D. Pizzuto R. Paventi G. Passarella S. Biochim. Biophys. Acta. 2005; 1708: 13-22Crossref PubMed Scopus (25) Google Scholar, Bari L. Valenti D. Pizzuto R. Paventi G. Atlante A. Passarella S. Biochem. Biophys. 2005; PubMed Scopus Google Scholar). The of AtD-LDH D. B. H. R. PubMed Scopus Google Scholar), mitochondrial R. A. K. M. R. PubMed Scopus Google Scholar, R. R. R. T. J. 2004; PubMed Scopus Google Scholar), and clear preference for d-lactate that enzyme is a for the oxidation of d-lactate in or the glyoxalase system in A. thaliana growth in with MG or d-lactate was impaired in knock-out with in the d-LDH that in planta enzyme to the of MG to In with analysis T. S. M. H. K. PubMed Scopus Google showed that AtD-LDH is with glyoxalase I in the coexpression The in data with the recombinant AtD-LDH showed that cytochrome c is a for the from d-lactate oxidation to for S. cerevisiae and K. lactis, where the oxidation of d-lactate a mitochondrial d-LDH to cytochrome c (8Lodi T. Ferrero I. Mol. Gen. Genet. 1993; 238: 315-324Crossref PubMed Scopus (69) Google Scholar, 11Lodi T. O'Connor D. Goffrini P. Ferrero I. Mol. Gen. Genet. 1994; 244: 622-629Crossref PubMed Scopus (36) Google Scholar), and for R. where the are to cytochrome S. Aizawa Y. Kai T. Amachi S. Shinoyama H. Fujii T. Biosci. Biotechnol. Biochem. 2004; 68: 516-522Crossref PubMed Scopus (17) Google Scholar). Atlante (13Atlante A. de Bari L. Valenti D. Pizzuto R. Paventi G. Passarella S. Biochim. Biophys. Acta. 2005; 1708: 13-22Crossref PubMed Scopus (25) Google showed that a enzyme d-lactate with electron to in H. Thus, it is that the oxidation of d-lactate by d-LDH is to the mitochondrial most by to cytochrome c in A. thaliana from d-lactate to the the glyoxalase system is highly the electron is The of A. thaliana locus encodes a of that FAD as group in the of the protein and a and the enzyme is as a Analysis of the substrate revealed that the recombinant of A. thaliana locus catalyzes the oxidation of d-2HG with a preference other substrates and it is a results by to activity of and and mitochondrial of A. thaliana and the knock-out mutants which demonstrated that AtD-2HGDH is responsible for the total d-2HGDH activity in the AtD-2HGDH is to mitochondrial enzymes from other that are of a of acids Y. G. D. M. Biochem. J. 2004; PubMed Scopus Google Scholar, Biochem. J. PubMed Scopus Google Scholar, R. Biochem. J. PubMed Scopus Google Scholar). enzymes d-2HG showed activity and d-lactate, and In to the plant enzyme to have as it showed to specific for On the other hand, AtD-2HGDH cytochrome c as an electron and is to transfer to NAD or The of the in electron to Although the substrate of is a known in and is known about the biological function of in plant In d-2HG is as a degradation of G. G. S. Biochem. Biophys. PubMed Scopus Google and it was to the from a in a reaction in which is to H. FEBS Lett. 2004; PubMed Scopus Google Scholar). ortholog of enzyme been found in d-2HG in during d-2HG J. PubMed Scopus Google Scholar). This is by in d-2HGDH Y. S. J. Genet. 2005; PubMed Scopus Google Scholar), the electron transfer protein D. The and of Scholar), or the ETF-ubiquinone oxidoreductase D. The and of Scholar). and to transfer by mitochondrial from to the to D. The and of Scholar, A. Biochem. J. PubMed Scopus Google Scholar). In the system a role in β-oxidation of acids and catabolism of amino acids and K. J. PubMed Scopus Google Scholar, K. Cell. 2005; PubMed Scopus Google Scholar). which the mitochondrial with from substrates other than K. J. PubMed Scopus Google Scholar, K. Cell. 2005; PubMed Scopus Google Scholar). The analysis of A. thaliana knock-out mutants in genes demonstrated that they are involved in the catabolism of and which are during and and are for in K. J. PubMed Scopus Google Scholar, K. Cell. 2005; PubMed Scopus Google Scholar). The as to which the of d-2HG in The of d-2HG the of the of and the of a (EC in The existence of activity was in and J. Bacteriol. PubMed Google Scholar, J. Bacteriol. PubMed Google Scholar). is in the of and catabolism is a of metabolism of the from the degradation of the of J. Biol. PubMed Scopus Google Scholar). is a in other it to to which the acid in the existence of catabolic is clear as to or found J. Biol. PubMed Scopus Google Scholar). it was that by a β-oxidation J. Biol. PubMed Scopus Google Scholar). The existence of a d-2HGDH activity to an alternative for degradation in which mitochondrial d-2HGDH and a d-2HG in to transfer from to the during of substrates from proteolysis and/or lipid degradation In d-2HG is which is the acid of in which is by and is to the to P. A. M. B. A. Biochem. Scholar). In with the of in analysis of T. S. M. H. K. PubMed Scopus Google showed that AtD-2HGDH is with the enzymes of enzymes of the β-oxidation of and the acids for degradation to and and required for acid S. Y. Biol. 10: PubMed Scopus Google Scholar, I. Cell. 2005; PubMed Scopus Google Scholar). several enzymes involved in and degradation are in the same and β-oxidation in the degradation of is by the of a M. B. B. J. Biol. PubMed Scopus Google Scholar), and the of the in that is a of from and/or in the Y. H. Y. Y. T. M. Y. I. PubMed Scopus Google Scholar). In several enzymes involved in the degradation and the acid oxidation are found in the same coexpression network and Although the of or degradation in is S. Rev. Biol. PubMed Scopus Google Scholar), it was that is oxidized to the which to as was for the degradation of in G. Mol. Genet. 2004; PubMed Scopus Google Scholar). A. thaliana a and it was that A. thaliana is involved in degradation K. Cell. 2005; PubMed Scopus Google Scholar). and are in the same This and the that AtD-2HGDH cytochrome c as electron that most probably AtD-2HGDH to the electron the of the data P. M. L. 2004; PubMed Scopus Google that the of AtD-2HGDH the of of genes enzymes of β-oxidation in a of Thus, AtD-2HGDH required for the of from and protein In analysis of the and mitochondrial analysis that AtD-2HGDH is to R. A. K. M. R. PubMed Scopus Google Scholar, H. Mol. Cell. 2008; PubMed Scopus Google Scholar), as it is from and Y. G. D. M. Biochem. J. 2004; PubMed Scopus Google Scholar, H. FEBS Lett. 2004; PubMed Scopus Google Scholar). In the mitochondrial of AtD-2HGDH the proposed are to the of AtD-2HGDH to plant with

) leaf growth without causing any other visible stress symptoms, including the accumulation of DNA damage. We conducted kinematic analyses of cell division and expansion to understand the impact of UV-B radiation on these cellular processes. Our results demonstrate that the decrease in leaf growth in UV-B-irradiated leaves is a consequence of a reduction in cell production and a shortened growth zone (GZ). To determine the molecular pathways involved in UV-B inhibition of leaf growth, we performed RNA sequencing on isolated GZ tissues of control and UV-B-exposed plants. Our results show a link between the observed leaf growth inhibition and the expression of specific cell cycle and developmental genes, including growth-regulating factors (GRFs) and transcripts for proteins participating in different hormone pathways. Interestingly, the decrease in the GZ size correlates with a decrease in the concentration of GA19, the immediate precursor of the active gibberellin, GA1, by UV-B in this zone, which is regulated, at least in part, by the expression of GRF1 and possibly other transcription factors of the GRF family.

Shipping of peaches to distant markets and storage require low temperature; however, cold storage affects fruit quality causing physiological disorders collectively termed 'chilling injury' (CI). In order to ameliorate CI, different strategies have been applied before cold storage; among them heat treatment (HT) has been widely used. In this work, the effect of HT on peach fruit quality as well as on carbon metabolism was evaluated. When fruit were exposed to 39 degrees C for 3 d, ripening was delayed, with softening inhibition and slowing down of ethylene production. Several differences were observed between fruit ripening at ambient temperature versus fruit that had been heat treated. However, the major effects of HT on carbon metabolism and organoleptic characteristics were reversible, since normal fruit ripening was restored after transferring heated peaches to ambient temperature. Positive quality features such as an increment in the fructose content, largely responsible for the sweetness, and reddish coloration were observed. Nevertheless, high amounts of acetaldehyde and low organic acid content were also detected. The differential proteome of heated fruit was characterized, revealing that heat-induced CI tolerance may be acquired by the activation of different molecular mechanisms. Induction of related stress proteins in the heat-exposed fruits such as heat shock proteins, cysteine proteases, and dehydrin, and repression of a polyphenol oxidase provide molecular evidence of candidate proteins that may prevent some of the CI symptoms. This study contributes to a deeper understanding of the cellular events in peach under HT in view of a possible technological use aimed to improve organoleptic and shelf-life features.