The Biochemistry of the Grape Berry

Berry water content during development and at harvest is a crucial parameter of the vintage quality by directly impacting the concentration of sugars and flavour compounds. In the last decade, there has been considerable progress in understanding how berry water relations are involved in the ripening process as a result of exploring and integrating developmental changes in berry hydraulic conductivity, mineral nutrient loading, phloem unloading, xylem development, functionality of xylem vessels, cell vitality, cell water relations, gene expression and the molecular biology of aquaporins (water channels). In this chapter, we review the most recent advances in our understanding of the berry water influx and efflux during development and their consequences on cell turgor and apoplast solute concentrations, taking account of the changes in cell vitality observed late in ripening in some varieties. The role of aquaporins in the regulation of berry water relations is also presented, with a particular emphasis on the varietal differences observed in aquaporin functioning. Finally, the impact of water status of the parent vine on berry ripening is discussed. From this review, it appears that the varietal differences observed at different regulatory levels of berry water status represent a great opportunity to gain a better understanding of the ripening process.

Both organic and inorganic compounds are vital for grapevine health and maintenance of vigour suitable for fruit production. In particular, the mineral soil composition has an essential influence on grape quality and on the organoleptic properties of wine. Grape berries are very rich in K; however, other mineral elements such as N, Ca, P, Mg, S and several micronutrients affect berry development and maturation in ways that have not always been acknowledged. Some of these compounds directly affect berry set and yield, whereas others act indirectly by modulating vine physiology. In addition, the complex interaction between mineral elements and organic molecules also renders different outcomes in vine and grape berry. The present chapter provides an overview on the dynamics of the major mineral compounds in the grapevine, especially in the berry, and on the contribution of each element to berry quality and yield. Special emphasis is given to K, since it is the main cation in must and wine and plays a decisive role in berry development and wine quality.

Phloem transport of assimilates provides the materials needed for the growth and development of reproductive structures, storage and developing organs, and has long been recognised as a major determinant in crop yield. Thus, the understanding of the mechanisms and regulations of sugar transport into sink tissues has an important basic and applied relevance. The Grapevine is a good example of a crop where sugar accumulation in the fruit has an important economic role. Massive sugar transport and compartmentation into the grape berry mesocarp cells (up to 1 M glucose and fructose) start at veraison and continues until the harvest. Sucrose transported in the phloem is cleaved into hexoses by invertases and stored in the vacuole. The Sugar content determines the sweetness of table grapes, wine alcohol content, and regulates gene expression, including, for example, several genes involved in the synthesis of secondary compounds which contribute to grape and wine quality. Many viticultural practices affect source/sink relationships, thus altering sugar concentration in the berry. For instance, the rootstock used, which is a potential sink, has a strong impact on source activity, by affecting the morphology and activity of the aerial part of the plant. Molecular approaches have also provided major advances in grapevine research. Monosaccharide and disaccharide transporter genes have been recently identified and their products studied in heterologous systems. The sequencing of the grapevine genome and the development of grape microarrays have made a valuable contribution to the study of the biochemistry of grape berry development and ripening, for example, low affinity glucose uniporters identified in the genome may also be involved in the sugar uptake. In the present chapter, the routes of sugar import and storage in the grape cells are updated and discussed and a model with the main transport steps and biochemical pathways is proposed.

Grape berries are commonly perceived to be composed principally of high concentrations of fermentable sugars, accompanied by a complex suite of polyphenolic compounds responsible for colour and ‘mouthfeel’ properties. The organic acid composition of the berry, which is principally a reflection of the metabolism of tartaric and malic acids during development and ripening, has several important consequences for the use of grapes in winemaking. Early research showed two unusual features of acid metabolism in grapes – the occurrence of significant concentrations of tartaric acid, and a marked decrease in the concentration of malic acid as berries enter the ripening stage. Despite a few ‘false starts’, a synthetic pathway that led to the formation of tartaric acid from ascorbic acid was identified, and in the past few years it has been proved possible to confirm this by biochemical and molecular biological approaches. Evidence for the synthetic route to malate formation in the berry proved simpler to identify, but the greater metabolic versatility of malate compared with tartaric acid, and the post-veraison breakdown of malate by a variety of pathways have ensured a continuing interest in this component of the berry’s acid complement. Modern ‘post-genomic’ approaches have been increasingly used to enable the measurement and analysis of berry metabolism. These approaches, and the advent of a readily transformable ‘model’ grapevine system, will undoubtedly continue to play a major role in the development of the understanding needed for the rational modification of berry acid composition in response to changing environmental and cultural practices.

Phenolic compounds are important components of the grape berry in the determination of wine style and quality. In the past decade, significant advances towards a better understanding of the genetics, biochemistry, and physiology governing the synthesis of this class of secondary metabolite have been made. This deeper knowledge of phenylpropanoid and flavonoid metabolism strengthens the foundation for practical applications in the vineyard; investigations involving cultural practices and manipulation of environmental effects can help viticulturists deliver grapes to winemakers that are better suited to particular enological objectives, as well as possibly enriching the product in health-promoting compounds.

The grape berry is the site of biosynthesis and accumulation of compounds that are likely to contribute in wines some of their aromatic characteristics. These compounds have the aromatic potential, and exist in part as volatile forms but mainly as non-volatile aroma precursors that can be released through chemical and biochemical reactions during vinification and ageing. The chemistry of aromas has gradually identified a number of key volatile compounds and their precursor forms. Among these compounds, we have considered methoxypyrazines, monoterpenes and C13-norisoprenoids, which are derivatives of carotenoids and sulfur compounds possessing a thiol group. They represent a large diversity of flavour nuances (herbaceous, fruity, floral, empyreumatic, etc.), often at trace concentrations (in the nanogram per litre range). The reactivity of these compounds in enological conditions, and the state of the knowledge on their biosynthesis in grapes, both in volatile form, possibly odorous, and in precursor form (glycosylated or conjugation of cysteine and glutathione) are described.

Polyamines have been correlated with numerous plant processes, and the elucidation of their physiological role(s) has been a state-of the-art research topic in the recent years. Both their anabolism and catabolism seem to possess a regulatory role in development, stressing the necessity of their fine-tuning. Amongst the most important is the control of fruit-set and development. In this Chapter, the role of polyamines in the different stages of the grape berry development is discussed. In addition, references to other plant species are also included to strengthen the pan-species’ specific role of polyamines.

Grape berry cells have a complex vacuolar membranous system highly specialised in solute storage. Vacuoles are the main reservoirs of sugars, organic acids, aromas, flavours, ions and water. After veraison, when growth occurs exclusively by cell enlargement, the vacuole volume greatly increases due to the massive sugar and water uptake. A large number of tonoplast proteins, including pumps, carriers, ion channels and receptors support the numerous functions of the plant vacuole. Some of them have been well characterised in several plant models at the biochemical and molecular level, including the most abundant ones, V-ATPase, V-PPase, and water channels (aquaporins). The present chapter provides an overview on the diversity and storage role of the vacuole of grape cells, and the molecular mechanism involved in solute transport across the tonoplast is updated and discussed.

The cell wall (CW) is the dynamic border of plant cells. In grape berries, the CW decisively accounts for the difference between the pulp and skin cells, with direct consequences on the grape characteristics, wine quality and wine-making methods. The softening of mature berries results from the depolymerisation and solubilisation of CW polymers. Modifications of grape pulp and skin CW provide the flexibility for cell expansion during fruit growth and to modulate the final texture. Wine making and berry processing methods are directly related with the absence, in white wines, or the presence, in red wines, of skin CW in the fermenting must. Anthocyanin extraction depends directly on skin yielding of the pigment upon CW degradation. During fruit growth and ripening, the cooperative action between different enzyme families is capital in CW metabolism. The sequencing and public availability of the Vitis genome allowed us to focus on individual pathways, to profile the expression pattern of isoforms associated with each tissue, developmental phase or stress response, anticipating the effects on berry (and wine) production and quality. Retrieving the sequences of genomic coding regions and the predicted enzymes that act on the Vitis, CW allows us for the first time to tackle the grape berry Cell Wallome.

This chapter describes the role of plant hormones during the development of the nonclimacteric grape berry. Advances in analytical methods have enabled a better understanding of hormone accumulation although the role of some hormones, for example ethylene, during berry development, remains unclear. Even though the biosynthesis pathways for a number of hormones have been elucidated in grapevine, our understanding of some, for example, the auxin biosynthesis pathway, is still embryonic in grapes. Recent work regarding the breakdown of ABA and auxins has demonstrated the importance of catabolism in determining the final hormone profile during development. Much attention has been focused on the role of hormones during ripening due to the importance of the ripening process to the relevant grape industries. Brassinosteroids have now been added to the list of hormones that can advance ripening. High throughput techniques for analysing changes in gene expression, such as microarray analysis and deep sequencing, are now being used to assess the role of hormones during berry development. These techniques are ideally suited to analyse the effects of exogenous hormones on gene transcript levels, and the information gained will provide a global view of hormonal control at the transcriptome level. It is much more difficult to obtain an overall picture of the changes in protein levels as a response to hormone treatment. However, proteomic studies have demonstrated changes in the levels of a number of proteins, and some of these changes reflect changes in gene transcript levels. This chapter is a review of the current state of knowledge regarding the hormonal control of berry development (excluding polyamines which are described in Chapter 7). A portion of this work has been referred to in a previous review [1].

The investigation of key molecular events underlying grape berry development can provide useful data for the improvement of mature berry quality traits, a goal in this commercially important fruit crop. Berry development has been investigated at the transcriptional level to study global expression profiles during formation and ripening, while a smaller number of large-scale metabolite studies have been carried out till now. Here we present a meta-analysis of the “transcriptional story” of grape berry development by analysing the data from all transcriptional studies published to date using different analysis methods in diverse cultivars.

The microbial community of grape berry is composed of an array of species exhibiting differential physiological characteristics and relevance to vine growing and winemaking. The most important phytopathogens responsible for grapevine diseases worldwide are the oomycete Plasmopara viticola (downy mildew) and the ascomycete Erysiphe necator (powdery mildew). The causal agent of grey rot is the saprophytic mould Botrytis cinerea. A wide diversity of yeast species are also common contaminants of berry surfaces, but the key agent of wine fermentation, Saccharomyces cerevisiae, is rarely recovered from the grapes. Bacterial groups include the spoiling acetic acid bacteria and lactic acid bacteria responsible for the malolactic fermentation. These microorganisms colonise grape surfaces from berry set to ripening, following a repeatedly cyclic pattern year after year. Highly complex interactions and chemical signalling take place among grapevines themselves and with the intervening biota, which also include insects, birds and mammals. The fundamental role played by the nonmicrobial biota in grape berry microbiota ranges from their role (especially in the case of insects) as microbial vectors to the damage directly inflicted on the grapes, which pave the way to the entrance of the saprophytes. The precise biota and the resulting interactions depend fundamentally on the berry development stage, on the intactness of the grape skin and on the prevailing environmental conditions, and exert a profound effect on the fruit quality. Given the great ecological, technological and economical importance of studying the grape microbiota, it is somewhat surprising to find scarce and fragmented information available on these topics.

Here we provide a balanced, highly multidisciplinary overview of the most relevant components of grape berry microbiota. Our proposal establishes four distinct groups of microorganisms - residents, adventitious, invaders and opportunists - which are defined on the basis of grape biochemical evolution, nutrient availability and ability to proliferate on berry surface. Their natural proliferation is particularly dependent on two main events: veraison and berry damage. The origin and the colonisation sequence on berry surface by those several groups of microorganisms will be tentatively settled.

The results of epidemiological cohort studies show the separate influence of a moderate consumption for different types of alcoholic beverages (beer, wine, spirits) and their effects on coronary heart diseases, with a superior protective effect for wine. Wine possesses specificity due to the phenolic antioxidants (flavonoids and non-flavonoids). Several ways for metabolism and polyphenol excretion of wine are eligible. Moderate wine consumption can lead to decreased platelet aggregation and vasodilatation. Physiological effects derived from the nutritional consumption of wine polyphenol extracts, ethanol or their possible synergistic combination may lead to a prevention of atherosclerosis, diabetes or hypertension. This synergy permits:

• In the case of atherosclerosis, a decrease in the fatty streak area and cholesterolemia, and an increased level in specific antioxidant enzymes active against free radicals,

• To restore the antioxidant capacity of plasma, thus allowing for an improvement of the defences against oxidative stress of diabetes and a favourable action on insulin secretion, to normalise the systolic pressure, and a high reduction of cardiac hypertrophy, as well as free radical generation to the thoracic aorta and heart tissues in the case of hypertension. Wine can play a role in preventive nutrition, when its regular and daily consumption with moderation is added to the diet.