ReviewRegulation of malate metabolism in grape berry and other developing fruits
Graphical abstract
An assembly of new microarray data with published protein and enzyme activity patterns in fruits, to explore pathways of malate metabolism in response to developmental and climatic changes.
Introduction
Fruits contain a wide variety of compounds, including organic acids that dictate acidity throughout development. Malate is the predominant acid in many fruits, both climacteric, including plum (Singh and Singh, 2008), tomato (Kortstee et al., 2007), peach (Wu et al., 2005), apple (Beruter, 2004), kiwifruit (Walton and De Jong, 1990), mango (Selvaraj and Kumar, 1989), banana (Agravante et al., 1991), pear (Chen et al., 2007), fig. (Shiraishi et al., 1996) and peach (Moing et al., 1998) and non-climacteric, including pineapple (Saradhuldhat and Paull, 2007), lime, orange, lemon (Albertini et al., 2006), cherry (Usenik et al., 2008), strawberry (Moing et al., 2001) and grape (Kliewer et al., 1967). While organic acids are chiefly in the ionised form at cellular pH, for the purposes of this review the term “malate”, which technically describes the conjugate base, will be used to refer to all physiological forms of the compound. This nomenclature will be maintained for all organic acids mentioned in the text.
Malate can play a variety of roles in plants, from controlling stomatal aperture, improving plant nutrition, and increasing resistance to heavy metal toxicity (Fernie and Martinoia, 2009, Schulze et al., 2002), to other processes more intricately linked with metabolic pathways, some of which are represented in Fig. 1. Patterns of malate accumulation differ between plant species, and even cultivars (Kliewer et al., 1967, Selvaraj and Kumar, 1989). In fruit, patterns of malate accumulation and degradation cannot be explained by the classification of species as climacteric or non-climacteric, nor can they be attributed to changes in overall respiration rates. Some climacteric fruit such as plum and tomato appear to utilize malate during the respiratory burst (Goodenough et al., 1985, Kortstee et al., 2007, Singh and Singh, 2008), while others such as mango and banana continue to accumulate malate throughout ripening, even at the climacteric stage (Agravante et al., 1991, Selvaraj and Kumar, 1989). Non-climacteric fruits also display widely varying malate accumulation and degradation events (Albertini et al., 2006, Moing et al., 2001, Saradhuldhat and Paull, 2007). However, it should be noted that most of the studies listed above have depicted malate accumulation in fruits as various concentration units or percentages, rather than on a whole fruit basis. It is difficult to determine whether malate losses shown in these data are due to active metabolism, or dilution as the fruits become larger. Both situations will result in a decrease in malate concentration (but not total fruit content), as demonstrated by Walton and De Jong (1990) in kiwifruit and Famiani et al. (2005) in numerous other fruits. Fruit malate levels can also be dramatically affected by exposure of the plant to various environmental conditions (Lakso and Kliewer, 1978, Richardson et al., 2004, Ruffner et al., 1976).
In the non-climacteric fruit of Vitis vinifera (grape), malate metabolism has been a strong focus of research, as the balance of acids in winegrape must (juice) is central for supporting desirable growth (and preventing undesirable growth) of microorganisms responsible for wine fermentation. Malate concentration can also affect final wine characteristics through involvement in secondary processes such as carbonic maceration and malolactic fermentation, and can even alter the growth capabilities of malolactic bacteria (Kunkee, 1991). Unlike many other fruits, grapes do not contain large amounts of citrate, and the large quantity of tartrate present in the fruit is not used in primary metabolic pathways. Therefore malate is the only high-proportion organic acid that is actively metabolised throughout ripening of grapes. Combined with the ever-improving density of genetic information available for V. vinifera, grapes make an ideal system for studying the metabolism of malate during ripening of non-climacteric fruit. A large body of work has also been reported for tomato acid metabolism, which is used to exemplify a climacteric equivalent to grapes in this review.
Loss of grape berry malate is due to metabolic degradation during ripening, which occurs after an earlier period of accumulation. The switch from net accumulation to degradation of malate occurs just before veraison, or the inception of ripening (Ruffner and Hawker, 1977). Pre-veraison grapes accumulate malate mostly through the metabolism of sugars that have been translocated to the berry, but also potentially through fruit photosynthesis (Hale, 1962). In post-veraison fruit, malate is liberated from the vacuole and becomes available for catabolism through various avenues, including the TCA cycle and respiration, gluconeogenesis, amino acid interconversions, ethanol fermentation, and the production of complex secondary compounds such as anthocyanins and flavonols (Famiani et al., 2000, Farineau and Laval-Martin, 1977, Ruffner, 1982, Ruffner and Kliewer, 1975). With the accumulation of sugars and inhibition of glycolysis in ripening grapes (Ruffner and Hawker, 1977), malate is likely a vital source of carbon for these pathways.
Some fruits, including mango, kiwifruit and strawberry may rely less on malate, and more on the hydrolysis of accumulated starch as a source of carbon for biosynthesis and energy metabolism during ripening (Han and Kawabata, 2002, Moing et al., 2001, Selvaraj and Kumar, 1989). These particular species’ display no net loss of malate throughout ripening (Moing et al., 2001, Ruffner, 1982, Selvaraj and Kumar, 1989, Walton and De Jong, 1990). For this reason, the metabolism of malate will be further explored with a focus on grape and tomato fruits, in which the acid plays a more metabolically active role (Goodenough et al., 1985, Ruffner, 1982).
Section snippets
Pathways of malate synthesis in fruit
In pre-veraison grape berries, sucrose transported from the leaves is broken down to glucose and fructose, which can enter glycolysis for use in respiration. Therefore translocated sucrose acts as the major fuel for ATP synthesis, and also enables synthesis of malate in the fruit. Enzymes responsible for this synthesis are present and active in grapes (Hawker, 1969, Taureilles-Saurel et al., 1995a, Taureilles-Saurel et al., 1995b). Photosynthesis is another means of synthesising malate in the
Pathways of malate degradation in fruit
Once grapes reach veraison, sugar metabolism begins to favour hexose accumulation and synthesis rather than catabolism, through regulation of key enzymes of the glycolytic and gluconeogenic pathways (Ruffner and Hawker, 1977). Therefore at this stage sugars relinquish the role of major carbon source for energy metabolism and biosynthesis. Malate released from the vacuole during ripening has the potential to fulfill this function, and can do so through involvement in gluconeogenesis, respiration
Intracellular transport of malate in the grape berry
In a recent analysis using introgression lines of tomato, QTL mapping of metabolomic profiles against morphological traits of the fruit and whole plant were used to uncover relationships between genes, metabolites and plant phenotype (Schauer et al., 2006). In this study, lines containing altered levels of malate and other organic acids showed little or no co-localisation with metabolic enzymes, suggesting that membrane transport and other regulatory influences are important contributors to
Approaches to identify genes linked to high and low malate fruit
Patterns of malate and citrate accumulation in a low-acid peach cultivar have been used to investigate causes of variation in fruit acidity, by exploring differences in gene expression and enzyme activity patterns relative to a normal-acid cultivar. These studies have indicated that PEPC activity, while linked to the synthesis of malate in normal acid varieties, cannot explain the variation in malate and citrate levels seen in the low-acid fruit (Moing et al., 2000), nor can NAD-MDH and NADP-ME
Temperature regulation of fruit malate metabolism
In some fruits, particularly grape, it is well established that exposure of the ripening fruit to warmer climatic conditions leads to lower levels of malate at harvest (Lakso and Kliewer, 1978, Ruffner et al., 1976). Malate levels were also seen to be reduced upon high exposure to sunlight (Pereira et al., 2006), although this is most likely due to the effect of exposure on berry temperature (Spayd et al., 2002). Synthesis of malate involves an exothermic reaction that may occur more favourably
Transgenic approaches to modifying malate metabolism
Few attempts at genetic modification aimed at altering malate metabolism have been attempted in fruits, due to the long period required for transformation of plants and subsequent production of fruits. Tomato is perhaps the fastest model fruit-bearing plant to transform, and has been used to over-express and under-express an ADH gene (Adh2) (Speirs et al., 1998). ADH-over-expressing fruit had higher levels of some alcohols, in particular hexanol and z-3-hexenol, which gave the fruit a more ripe
Conclusions
The information described in this review serves to summarise advances made in fruit malate metabolism research, particularly over the last 20–30 years, as an update to the comprehensive review based in grapes by Ruffner (1982). Levels of malate in harvested fruit may be largely determined by the rate of degradation during ripening (Beruter, 2004, Kortstee et al., 2007, Ruffner and Hawker, 1977, Walton and De Jong, 1990). Results presented from publicly available microarray data (Deluc et al.,
Role of the funding source
Funding Source: Grape and Wine Research Development Corporation. Grant awarded to Dr. Christopher M. Ford and Assoc. Prof. Kathleen L. Soole. The funding source had no involvement in the writing of, or the decision to publish this paper.
Crystal Sweetman is a Ph.D. candidate in the School of Biological Sciences at Flinders University of South Australia, in the laboratory of Associate Professor Kathleen Soole. She completed her Bachelor of Science at Flinders University in 2004, and Honours in 2005. Her Ph.D. research is part of a larger project concerning metabolism of organic acids in grapevine. Crystal is currently investigating the influences of malic acid metabolism on berry characteristics during ripening and in response
References (147)
- et al.
Vacuolar ion channels of higher plants
Adv. Bot. Res.
(1997) - et al.
Biochemical and molecular characterization of fumarase from plants: purification and characterization of the enzyme-cloning, sequencing, and expression of the gene
Arch. Biochem. Biophys.
(1997) Carbohydrate metabolism in two apple genotypes that differ in malate accumulation
J. Plant Physiol.
(2004)- et al.
Chemical compositional characterization of eight pear cultivars grown in China
Food Chem.
(2007) - et al.
NADP-malic enzyme from plants: a ubiquitous enzyme involved in different metabolic pathways
FEBS Lett.
(2001) - et al.
Malate. Jack of all trades or master of a few?
Phytochemistry
(2009) - et al.
Respiratory metabolism: glycolysis, the TCA cycle and mitochondrial electron transport
Curr. Opin. Plant. Biol.
(2004) - et al.
NADP-linked malic enzyme and malate metabolism in ageing tomato fruit
Phytochemistry
(1985) Changes in the activities of malic enzyme, malate dehydrogenase, phosphopyruvate carboxylase and pyruvate decarboxylase during the development of a non-climacteric fruit (the grape)
Phytochemistry
(1969)- et al.
The physiology, biochemistry and molecular biology of the plant vacuolar ATPase
Adv. Bot. Res. Incorp. Adv. Plant Pathol.
(1997)
Organic acid activation of the alternative oxidase of plant mitochondria
FEBS Lett.
Fermentative metabolism in grape berries: isolation and characterization of pyruvate decarboxylase cDNA and analysis of its expression throughout berry development
Plant Sci.
Isolation of mitochondrial malate dehydrogenase and phosphoenolpyruvate carboxylase cDNA clones from grape berries and analysis of their expression pattern throughout berry development
J. Plant Physiol.
Identification of a novel transporter for dicarboxylates and tricarboxylates in plant mitochondria
J. Biol. Chem.
The growing family of mitochondrial carriers in Arabidopsis
Trends Plant Sci.
The ubiquitin–proteasome pathway is involved in rapid degradation of phosphoenolpyruvate carboxylase kinase for C4 photosynthesis
Plant Cell Physiol.
Sugars and organic acids in ethanol-treated and ethylene-treated banana fruits
J. Jpn. Soc. Food Sci. Technol.
Changes in organic acids and sugars during early stages of development of acidic and acidless citrus fruit
J. Agric. Food Chem.
Fruit-specific V-ATPase suppression in antisense-transgenic tomato reduces fruit growth and seed formation
Planta
Ripening-related occurrence of phosphoenolpyruvate carboxykinase in tomato fruit
Plant Mol. Biol.
Metabolism of malic and tartaric acids in grape berries
Biochemistry (Moscow)
Respiration of fruits
Encyc. Plant Physiol.
Fruit Photosynthesis
Plant Cell. Environ.
Carbon metabolism of peach fruit after harvest: changes in enzymes involved in organic acid and sugar level modifications
J. Exp. Bot.
Aerobic fermentation in tobacco pollen
Plant Mol. Biol.
A genetic map of candidate genes and QTLs involved in tomato fruit size and composition
J. Exp. Bot.
Structure and function of malic enzymes, a new class of oxidative decarboxylases
Biochemistry
Alcohol dehydrogenase 2 and pyruvate decarboxylase induction in ripening and hypoxic tomato fruit
Plant Physiol. Biochem.
Phosphoenolpyruvate carboxylase: a ubiquitous, highly regulated enzyme in plants
Annu. Rev. Plant Physiol. Plant Mol. Biol.
Distribution of solutes within the developing grape berry in relation to its morphology
Am. J. Enol. Vitic.
Adoption of a system for identifying grapevine growth stages
Am. J. Enol. Vitic.
Water and salinity stress in grapevines: early and late changes in transcript and metabolite profiles
Funct. Integr. Genomics
The fine control of cytosolic pH
Physiol. Plant.
Transcriptomic and metabolite analyses of Cabernet Sauvignon grape berry development
BMC Genom.
Phosphoenolpyruvate carboxylase during grape berry development: protein level, enzyme activity and regulation
Aust. J. Plant Physiol.
Malic enzyme activity in apple fruit
Nature
Contribution of malic enzyme, pyruvate kinase, phosphoenolpyruvate carboxylase, and the Krebs cycle to respiration and biosynthesis and to intracellular pH regulation during hypoxia in maize root tips observed by nuclear magnetic resonance imaging and gas chromatography mass spectrometry
Plant Physiol.
The plant homolog to the human sodium/dicarboxylic cotransporter is the vacuolar malate carrier
Proc. Natl. Acad. Sci. USA
A genomic library of Lycopersicon pennellii in L. esculentum: A tool for fine mapping of genes
Euphytica
Isolation and characterization of six peach cDNAs encoding key proteins in organic acid metabolism and solute accumulation: involvement in regulating peach fruit acidity
Physiol. Plant.
An immunohistochemical study of the compartmentation of metabolism during the development of grape (Vitis vinifera L.) berries
J. Exp. Bot.
Phosphoenolpyruvate carboxykinase and its potential role in the catabolism of organic acids in the flesh of soft fruit during ripening
J. Exp. Bot.
Light versus dark carbon metabolism in cherry tomato fruits. II. Relationship between malate metabolism and photosynthetic activity
Plant Physiol.
Reverse genetic characterization of cytosolic acetyl-CoA generation by ATP-citrate lyase in Arabidopsis
Plant Cell
Inhibition of malic enzyme from grape berries by sulfhydryl-reagents and oxalic-acid
Am. J. Enol. Vitic.
Cloning of a full-length cDNA for malic enzyme (EC.1.1.1.40) from grape berries
Plant Physiol.
Isolation of mitochondria from leaf tissue of Panicum miliaceum, a NAD-malic enzyme type C4 plant
Plant Physiol.
Two ldh genes from tomato and their expression in different organs, during fruit ripening and in response to stress
Plant Mol. Biol.
Evolving concepts in plant glycolysis: two centuries of progress
Biol. Rev. Cambr. Philos. Soc.
Mitochondria/nuclear signaling of alternative oxidase gene expression occurs through distinct pathways involving organic acids and reactive oxygen species
Plant Cell Rep.
Cited by (348)
Optimization of apple fruit flavor by MdVHP1-2 via modulation of soluble sugar and organic acid accumulation
2024, Plant Physiology and Biochemistry
Crystal Sweetman is a Ph.D. candidate in the School of Biological Sciences at Flinders University of South Australia, in the laboratory of Associate Professor Kathleen Soole. She completed her Bachelor of Science at Flinders University in 2004, and Honours in 2005. Her Ph.D. research is part of a larger project concerning metabolism of organic acids in grapevine. Crystal is currently investigating the influences of malic acid metabolism on berry characteristics during ripening and in response to high temperatures, using a biochemical and molecular biology approach. This work is co-supervised by Dr. Chris Ford from the University of Adelaide.
Laurent Deluc graduated in 1996 in Forestry at the University of Nancy I, France. Later he obtained his Ph.D in 2004 at the University of Bordeaux I under the supervision of Prof. Said Hamdi investigating the transcriptional control of the flavonoid pathway in grape berry development. From 2005 to 2006, he worked as Post-Doctoral associate with Prof. John Cushman at the University of Nevada, Reno USA working on transcriptomic approaches of berry development and the effect of water deficit in berry quality. In 2007, he worked with Prof. Grant Cramer in the same university on proteomic and metabolomic aspects of the grape bud endodormancy in strong collaboration with Prof. Anne Fennell from South Dakota State University, USA. Since beginning 2009, he joined the Department of Horticulture at Oregon State University, Corvallis USA as Assistant Professor. He is mainly interested in different aspects of the berry quality (metabolomic and hormonal control throughout berry development), effects of abiotic stress on berry development and Rootstock-Scion interactions on berry quality.
Grant R. Cramer graduated from UC Davis with a Ph.D. in Plant Physiology in 1985. He started as an Assistant Professor at the University of Nevada, Reno in 1988 and became a Professor of Biochemistry and Molecular Biology in 2003. Dr. Cramer has studied the two components responsible for plant growth inhibition by salinity: osmotic and ionic effects for more than 20 years. In the last 10 years, he has focused on abiotic stress effects (water deficit, salinity and chilling) on grapevines. He has developed an integrated systems biology approach to study the effects of abiotic stress on mRNA, protein expression and metabolites in vegetative and fruit tissues of Vitis vinifera. Comprehensive analyses have characterized berry development, cultivar and tissue differences, and indicate wide-ranging effects of stress on many processes and metabolic pathways in both shoots and fruit.
Chris Ford has been lecturing in the area of Oenology since 2001; before which he held a number of post-doctoral research positions within Australia and the UK. His first degree was obtained in 1984 from Hatfield Polytechnic, and in 1988 Chris received a DPhil. from the University of Sussex for work in the biochemistry of nitrogen fixation. In 1990 Chris moved to The University of Adelaide. His research falls into three broad areas: developing an understanding of organic acid metabolism in plants using a range of biochemical, molecular and metabolic approaches; characterisation of glucosyltransferase enzymes (GTases) involved in secondary metabolism, and unravelling the outcomes of extended maceration (EM) in the production of red wines using a combination of chemical and sensory analyses.
Kathleen Soole holds an Associate Professor position in the School of Biological Sciences at Flinders University, Adelaide, Australia. She graduated with a Ph.D. in plant biochemistry in 1990 from the University of Adelaide, Australia. Her research interests lie in mitochondrial function in plant development and stress adaptation as well as a general interest in plant metabolism and its regulation.
- 1
Present address: Department of Horticulture, Oregon State University, Corvallis, OR 97331, USA.