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Abiotic stress and plant responses from the whole vine to the genes
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G.R. CRAMER Department of Biochemistry and Molecular Biology, University of Nevada, Reno, NV 89557, USA Corresponding author: Dr Grant R. Cramer, fax +1 775 784 1650, email cramer@unr.edu Abstract Drought, salinity and extreme temperatures signicantly limit the distribution of grapes around the world. In this review, the literature of grape responses to abiotic stress with particular reference to whole plant and molecular responses observed in recent studies is discussed. A number of short-term and long-term studies on grapevine shoots and berries have been conducted using a systems biology approach. Transcripts, proteins and metabolites were proled. Water decit, salinity and chilling altered the steady-state abundance of a large number of transcripts. Common responses to these stresses included changes in hormone metabolism, particularly abscisic acid (ABA), photosynthesis, growth, transcription, protein synthesis, signalling and cellular defences. Some of the transcriptional changes induced by stress were conrmed by proteomic and metabolomic analyses. More than 2000 genes were identied whose transcript abundance was altered by both water decit and ABA. Different gene sets were used to map molecular pathways regulated by ABA, water decit, salinity and chilling in grapevine. This work supports the hypothesis that ABA is a central regulator of abiotic stress tolerance mechanisms. ABA affects signalling pathways that trigger important molecular activities involving metabolism, transcription, protein synthesis, and cellular defence and also regulates important physiological responses such as stomatal conductance, photoprotection and growth. Systems biology approaches are providing more comprehensive understanding of the complex plant responses to abiotic stress. The molecular sets generated from mapping the ABA-inducible stress responses provide numerous targets for genetic and cultural manipulation for improved plant protection and grape quality. Keywords: abscisic acid, salinity, water decit

Introduction Levitt (1980) dened stress as an environmental factor that is potentially unfavourable to an organism and resistance as the ability of the organism to survive such an environmental factor. Survival is an important issue, but when one considers crops, one is also concerned about production of the harvestable portion of the plant. So there are at least two variables that one must differentiate when one is discussing stress resistance or tolerance. The mechanisms for these types of resistances can be quite different. Common environmental factors or abiotic stresses around the world include, drought (water decit), salinity, temperature, and acid soils (Tester and Bacic 2005). Rarely is there a single abiotic stress affecting a plant; almost always there are interacting factors. For example, plants exposed to the direct sun in the eld will most likely be exposed to levels of light that can cause photoinhibition. If the plant is sufciently stressed that its photosynthesis is reduced, then the plant will almost certainly be experiencing the secondary effects of photoinhibition as well as the effects of the primary stress (e.g. drought, salinity, high or low temperatures). Thus, while we may study such stresses in the lab in an articial
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environment and elucidate important mechanisms of resistance, one must consider the whole environment that a plant grows in to fully comprehend the stress resistance mechanisms needed in the eld. A common response to many stresses (e.g. water decit, salinity, high and low temperatures, herbivory, and pathogens) is an osmotic response (Tester and Bacic 2005, Lopez et al. 2008). Either the whole plant experiences osmotic stress or specic cells under attack experience the osmotic stress. The osmotic stress is caused by water loss, which is a daily problem for any land plant that is photosynthesising and transpiring. There are many abiotic stresses that signicantly limit the distribution of grapes around the world. These stresses reduce crop yields, but only water decit has been used in a positive way to enhance avour and quality characteristics of the berries (Roby et al. 2004, Chapman et al. 2005). In part, this effect is because of reduced shoot vigour and competition for carbon resources (a change in source to sink relationship). Berry size can also be reduced, concentrating avours and colour by increasing the skin surface: berry mass ratio (the skin being a signicant tissue for producing avours, tannins and colour). In addition, there are fundamental biochemical

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changes in berries under water decit that cause important metabolic changes that inuence berry avour and quality (Castellarin et al. 2007a, Deluc et al. 2009). Semi-arid regions where grapes are grown not only suffer from water decit but also are prone to salinisation. Extreme cold can limit distribution in the northern- and southern-most latitudes and at high elevation. In addition, frost and chilling damage occurs during the spring and can limit yields in more moderate climates. Heat and light stress often interact with water decit to accelerate water loss and plant strain. This paper reviews the literature of grape responses to abiotic stress with particular emphasis on whole plant and molecular responses observed in investigations performed in the authors research group. It will not be possible to discuss all abiotic stresses in this review. Our group has focused on the effects of water decit, salinity and chilling stress on grapes. Experiments were conducted at many levels: in the eld (water decit only), greenhouse and the laboratory, and at the whole plant (growth and photosynthetic processes) and molecular levels (transcripts, proteins and metabolites). A systems biology approach was used to integrate the data in an attempt to better understand the plants responses to abiotic stresses (Cramer et al. 2005). This review is divided into two sections: the rst part will discuss the effects of stress on growing shoot tips and a smaller second part will address briey the effect of water decit on berry metabolism. Shoot responses to water decit, salinity and chilling Vitis vinifera grapevines grow well in arid and semi-arid environments. V. vinifera has relatively high drought tolerance (McKersie and Leshem 1994, Grimplet et al. 2007a) and genetic variability within the species exists (Gaudillere et al. 2002). Once established in a deep soil with adequate water retention characteristics, grapevines produce root systems many meters deep enabling the vines to survive severe water decits. However, global warming is causing signicant climate change, making it necessary for future crop adaptation (Howden et al. 2007). Increased temperatures along with changing rainfall will require improved crop performance under water decits. Abscisic acid (ABA) is a hormone that is central to regulating the plant response to osmotic stress (Seki et al. 2007). It operates at the whole plant level regulating important processes such as shoot and root growth, thereby affecting the root to shoot ratio (Munns and Cramer 1996). It also regulates transpiration and water loss via stomatal closure. ABA also operates at the molecular level by regulating gene transcription, protein synthesis, signalling pathways, ion transport (and the transport of other organic molecules) and the production of important protectants against dehydration and photoinhibition (Yang et al. 2006, Seki et al. 2007). ABA in the xylem sap is a key root to shoot signal in drought-stressed plants (Schachtman and Goodger 2008). Other chemical signals that may inuence stomatal conductance during water decits include malate, protons,
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cytokinins, and ABA conjugates (Schachtman and Goodger 2008). Peptides and proteins may also act as signals in the xylem (Aki et al. 2008). Metabolites, peptides and proteins have been analysed in the xylem sap of maize (Alvarez et al. 2008) and while protein concentrations are affected by water decit, ABA was considered the most important signal in the xylem sap that regulates transpiration. Small RNAs (e.g. mRNAs) play signicant roles in abiotic stress resistance (Sunkar et al. 2007, Li et al. 2008), including drought resistance (Li et al. 2008). However, the role of peptides, proteins and mRNAs in the xylem sap remains unclear and requires further investigation (Schachtman and Goodger 2008). It is a very interesting coincidence that cells closely associated with the xylem (e.g. xylem parenchyma) are located where hydraulic conductance to the epidermis (expanding cells) is most restricted for water ow and growth (Tang and Boyer 2002), where the expression of nine-cis-epoxycarotenoid dioxygenase (NCED), the rate limiting enzyme for ABA biosynthesis, rst increases in response to water decit (Endo et al. 2008), and where the aquaporins VvPIP1;1 and VvPIP2;2 are most abundantly expressed in grapevine roots (Vandeleur et al. 2008). It is intriguing that VvPIP1;1 expression appears to be quite responsive to changes in water potentials inuencing cell and root hydraulic conductivity (Vandeleur et al. 2008). There is a wide range of water-use efciencies between grape cultivars (Bota et al. 2001). These differences are largely attributed to variation in stomatal conductance in response to water decits (Bota et al. 2001, Schultz 2003, Soar et al. 2006), but also can be related to differences in the change of root hydraulic conductance and aquaporin (water channel) expression in response to water decit (Vandeleur et al. 2008). Water decit increases ABA concentrations in the xylem sap and leaves of grapevine and changes in stomatal conductance are well correlated with ABA concentrations of the xylem sap (Okamoto et al. 2004, Soar et al. 2004, Pou et al. 2008). ABA also inuences hydraulic conductance (Hose et al. 2000), aquaporin gene expression (Tyerman et al. 2002, Kaldenhoff et al. 2008) and embolism repair (Lovisolo et al. 2008) in grapevines. Furthermore, there is signicant variation in ABA concentrations between rootstocks originating from different Vitis species and which have an inuence on scion (V. vinifera L. cv. Shiraz) photosynthesis and stomatal conductance (Soar et al. 2006). Thus, it appears that ABA plays a vital role in grapevine water relations during osmotic stress. In addition to reductions in stomatal conductance, water decit, salinity and low temperatures can reduce photosynthesis by photoinhibition of photosystem II (Liakopoulos et al. 2006, Cramer et al. 2007, Tattersall et al. 2007). In grapevines, the main protective mechanisms for photoinhibition appear to be related to nonphotochemical quenching involving xanthophylls and the D1 repair rate (Hendrickson et al. 2004). In addition, photorespiration was able to dissipate up to 20% of the energy ux contributing to electron transport. In Arabidopsis, ABA can protect the plant against photoinhibition

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-Carotene BHASE Chloroplast C13-norisoprenoid & C14-dialdehyde

-Cryptoxanthin BHASE

CCD1 Zeaxanthin Xan nthophyll Cycle NPQ1 Antheraxanthin ABA1 NPQ1 Violaxanthin Xan nthophyll Cycle ABA1

High g Light

Low Light

9 cis neoxanthin 9-cis-neoxanthin

Dihydroxy phaseic acid

NCED Cytoplasm Xanthoxin ABA2 ABA-aldehyde AAO3

Figure 1. The ABA metabolic pathway. BHASE, beta-carotene hydroxylase; ABA1, zeaxanthin epoxidase; NPQ1, violaxanthin de-epoxidase; CCD1, (9,10[9,10]carotenoid cleavage dioxygenase); NCED, nine-cis-epoxycarotenoid dioxygenase; ABA2, xanthoxin dehydrogenase; ABAHASE, (+)-abscisic acid 8-hydroxylase; UGT, UDP-glucose glucosyltransferase; BGL1, b-glucosidase. Shaded boxes represent different cellular compartments and have a compartment name inside of them, except for the grey box around BGL1, which represents endomembranes just as for ABAHASE.

Phaseic acid

8-Hydroxy abscisic acid

Endomembranes ABAHASE

UGT Abscisic acid BGL1 ABA-GE

by affecting brillin concentrations (Yang et al. 2006). Fibrillins are lipid-binding proteins that enhance photosystem II tolerance to light stress. The ABA metabolic pathway is displayed in Figure 1. NCED (nine-cis-epoxycarotenoid dioxygenase) represents the rst committed step in ABA biosynthesis (Schwartz et al. 2003) and its transcript abundance is often correlated with ABA concentrations. In grapevine, stomatal conductance is negatively correlated with ABA concentrations in the xylem sap and ABA concentrations in the leaves are correlated with the transcript abundance of VvNCED1 (Soar et al. 2004). In one study (access to the original data can be found at http://www.plexdb.org in the experiment entitled VV2), the impacts of long-term water decit and salinity on grapevine shoot tips was investigated (Cramer et al. 2007). A unique stress experiment was performed with Cabernet Sauvignon comparing the gradual effects of a decreasing water decit with equivalent salinity. Transcript and metabolite proling provided consistent and corroborative results with growth assays. At equivalent water potentials, water decit had a more severe effect than salinity on growth, gene expression and specic metabolites.

Stress broadly affected many gene transcripts involved with metabolism, protein fate, transport, transcription, cellular defence and communication/signalling (Cramer et al. 2007). Gene expression of the ABA and ethylene pathways was particularly increased by stress compared with other hormone pathways and was negatively correlated with stem water potentials. Energy metabolism was strongly increased by stress. There were higher concentrations of glucose, malate and proline in water decittreated plants as compared with salinised plants. These differences were linked to differences in gene expression and other metabolites of the photosynthetic, gluconeogenic and photorespiratory pathways. Not only are these solutes likely to aid plants in osmotic adjustment, but also may help plants cope with reactive oxygen species detoxication and photoinhibition. The simultaneous monitoring of the abundance of both transcripts and metabolites has reinforced the observations made with each metric alone. These observations demonstrate the information synergy derived from the use of integrative functional genomic approaches. The changes in the individual transcript abundance of many genes were similar to changes in our short-term study
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(Tattersall et al. 2007); the original data can be found in the experiment entitled VV1 in http://www.plexdb.org), however, there were indications that a larger and more complex response in the acclimation process occurred with a gradual long-term stress. In another study (Vincent et al. 2007), proteomic analysis indicated that the decrease in growth and stem water potentials of the vine was correlated with decreased amounts of proteins involved in photosynthesis and protein synthesis. Common transcript responses in shoot tips to drought, salinity and chilling (VV1 and VV2) In the short-term stress experiment (Tattersall et al. 2007), we found that grapevine shoot tips had distinct differences in response to chilling as compared with water decit or salinity. Nevertheless, there were also some common attributes for all three stresses. Microarray data sets for both the long-term (water decit and salinity, VV2) and short-term (water decit, salinity and chilling, VV1) experiments were surveyed to determine the set of transcripts in shoot tips that had common responses to these stresses (Table 1). There were 15 transcripts that were positively up-regulated in all conditions. Two of them were related to the biosynthesis of sugars (rafnose and trehalose), which putatively contribute to improved tolerance to osmotic stress (Taji et al. 2002, Downie et al. 2003). About half (8 of 15) were transcription factors and the rest were involved in signalling or stress responses (ABA, Ca2+ or H2O2). The catalase was up-regulated only after severe stress over time. The 9-cis-epoxycarotenoid dioxygenase 1 is NCED1, the rate-limiting step in ABA biosynthesis. The proteins in Table 1 most likely operate together in a common stress response pathway in grapevine. Particularly interesting was the up-regulation of a serine/threonine protein kinase that has high homology

with ATSR1 in Arabidopsis (Table 1). ATSR1 inuences the TOR gene in yeast, the central regulating kinase that stimulates and integrates metabolism for growth (BaenaGonzalez and Sheen 2008). Two transcription factors are worth noting. ATHB-12 in Arabidopsis is a homeodomain leucine zipper class I protein that responds to ABA through ABI1, a protein phosphatase. One of the functions of ATHB-12 is to inhibit growth (Olsson et al. 2004, Moes et al. 2008). The bZIP (basic-leucine zipper) transcription factor listed in Table 1 is very similar to a family of ABF transcription factors in Arabidopsis (Choi et al. 2000). They respond to ABA and bind to the abscisic acid responsive element motif in the promoter region of ABAinducible genes. Many of the genes are involved in stress tolerance mechanisms. It is apparent from the data in Table 1 and the previous discussion of the literature that ABA plays a central role in the response of grapevine to osmotic stress. Data from two recent studies in Arabidopsis were utilised (Huang et al. 2007, Matsui et al. 2008) in order to gain a wider view of ABAs role during osmotic stress. Both studies determined the response of the Arabidopsis transcriptome to ABA treatment using two different technologies and approaches. The set of transcripts affected signicantly by ABA from each experiment were downloaded and their sequences BLASTed against the grape transcript sequences that were affected signicantly by water decit. Note that this set of genes is a subset of all the water decitresponsive genes, indicating that there are factors other than ABA that contribute to water decit responses as well. The assumption made here is that these transcripts in this subset are also affected by ABA (this hypothesis remains to be validated by grapevine experiments with ABA applications or ABA mutants). The corresponding grape gene sets were placed into functional categories and

Table 1. Common responses for all stresses and both experiments (VV1 and VV2).
Affy probe Set ID 1608995_at 1617699_at 1609107_at 1609172_at 1610064_at 1617931_at 1618998_at 1619029_at 1620621_at 1614779_s_at 1620564_at 1608587_at 1610871_s_at 1608022_at 1614892_at Gene GSVIVG00025569001 GSVIVG00010921001 GSVIVG00014947001 GSVIVG00027622001 GSVIVG00023994001 GSVIVG00000517001 GSVIVG00030292001 GSVIVG00036604001 GSVIVG00015416001 GSVIVG00025566001 GSVIVG00023957001 GSVIVG00036780001 GSVIVG00002880001 GSVIVG00000988001 No hit Protein Rafnose synthase Trehalose phosphatase C2H2 zinc nger NAC domain protein WRKY DNA-binding protein ATHB-12 RAV transcription factor bZIP transcription factor NAC domain protein Remorin-like protein Serine/threonine protein kinase Calcium dependent protein kinase Catalase 2 9-cis-epoxycarotenoid dioxygenase 1 Protein phosphatase 2C ABI1 (PP2C) MIPS 2.1 functional category 01.05 C-metabolism 01.05 C-metabolism 11.02.03.04.01 transcription activation 11.02.03.04.01 transcription activation 11.02.03.04.01 transcription activation 11.02.03.04.01 transcription activation 11.02.03.04.01 transcription activation 11.02.03.04.01 transcription activation 11.02.03.04.01 transcription activation 16.03.01 DNA binding 30.01.05.01 protein kinase cascades 30.01.09.03 Ca2+ mediated signal transduction 32.07.07.01 catalase reaction 36.20.18.05 abscisic acid response 36.20.18.05 abscisic acid response

Gene names were obtained from a BLAST of the probe set sequence against the 8x assembly of the grape genome. These data were collected from the grape annotation page of http://www.plexdb.org. The probe set identication number for the Affymetrix grape genome array (version 1.0). 2009 Australian Society of Viticulture and Oenology Inc.

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Figure 2. Functional categorisation of a subset of grape transcripts responsive to water decit and also linked with Arabidopsis genes that are affected by ABA. This subset grape transcripts is referred to as WD-ABA transcripts. ABA responsive genes in Arabidopsis were identied using the Arabidopsis tiling array data set from Matsui et al. (2008) and the Arabidopsis genome array data set from Huang et al. (2007). The initial subset of genes in grapes responsive to water decit and which were also identied as ABA-responsive genes in Arabidopsis were functionally categorised (black bars) and then separated into those genes with increased (grey bars) or decreased (white bars) transcript abundance during water decit. Each subset is displayed separately with the subset of 2095 grape transcripts based on the tiling array data on the left and the subset of 644 grape transcripts based upon the genome array data on the right. Functional categories are organised along the y-axis from the highest to the lowest number of transcripts in a functional category (bottom to top). The % of transcript set refers to the % of total transcripts in that particular set of transcripts that were placed into that functional category. The tiling array data are from Matsui et al. (2008) and the genome array data are from Huang et al. (2007).

these grape gene sets separated into those that had an increased or decreased abundance during water decit. The results from this analysis (Figure 2) are quite intriguing. Although the data sets are represented by quite large differences in the number of transcripts (2095 vs 644) that are matched because of the different technologies used (tiling vs genome array, respectively), there are some interesting commonalities that stand out. For example, the transcript abundance was decreased for nearly all of the grapevine transcripts affected by water decit that are involved in protein synthesis. The differences in the % values between these two studies (5 vs 30% in Figure 2) was affected largely by the number of total transcripts involved in either set (2095 vs 644). The large decrease in protein synthesis transcripts was a rather striking result and is consistent with our earlier proteomic study (Vincent et al. 2007), in which a decrease in proteins involved in protein synthesis was highly correlated with the inhibition of growth. It is not clear whether the inhibition of protein synthesis by ABA is the cause of the growth reduction or protein synthesis is merely responding to feedback regulation in response to inhibited growth (less demand or need for proteins). This hypothesis requires further testing before any rm conclusions can be made. In both transcript sets from the two technologies (Figure 2), there was an increased proportion of transcripts with increased abundance that were involved in

transcription and cell rescue, defence and virulence. This is consistent with ABA playing a role in stress tolerance. The increased down-regulation of genes involved in biogenesis of the cell is consistent with the action of ABA and ATHB-12 described previously, however, these proportional differences are rather small. This data analysis serves only in the development of new hypotheses in order to stimulate new ideas and areas of research. Caution should be used in the interpretation of these results until additional experiments are performed. Other differences between other functional categories were not consistent between the studies and denitely require further testing. Transcript responding to osmotic stress only There are several transcripts that responded in all osmotic stress treatments, but not in chilling. They include NF-YA TF (1613912_at), a transcription factor responsive to ABA that affects approximately 130 Arabidopsis genes (Li et al. 2008). The transcript abundance of this transcription factor is regulated by a mRNA (miR169) that is affected by osmotic stress. Other transcription factors affected by osmotic stress include a member of the DREB family (1608315_at), which bind to a drought-responsive element in the promoter of drought-induced genes (Liu et al. 1998), ERF1 (1609559_at), a member of a family of ethylene response factors that bind to a cis-element in
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ethylene inducible genes (Hao et al. 1998), and KNAT3 (1615625_at), a homeodomain protein involved in regulating cell patterning and organ development (Truernit et al. 2006). In addition, osmotic stress affected CRY2 (1610235_at), a blue-light receptor that is a positive regulator of owering (Liu et al. 2008), thioredoxin (1615681_at), a chloroplast protein involved in redox regulation of photosynthesis and stress responses (Foyer et al. 2009), LTP (1616990_s_at), a lipid transfer protein that may be involved in signalling, and a avonol-oglucosyl transferase (1618155_at). Berry responses to water decit Using transcript and metabolite proling, our group showed that water decit had signicant impacts on the metabolism of grape berries (Deluc et al. 2009). Metabolic responses appear to be dependent on the cultivar and the colour of the grape. Water decit particularly affected ABA, carotenoid, amino acid, fatty acid and phenylpropanoid metabolism in two cultivars, Cabernet Sauvignon and Chardonnay, but in different ways. Water decit increased ABA concentrations in Cabernet Sauvignon berries, but not Chardonnay berries. ABA is known to enhance proline, sugar and anthocyanin accumulation in plants and the increased ABA concentration in Cabernet Sauvignon by water decit was consistent with this hypothesis resulting in increased accumulation of proline, sugar and anthocyanins relative to well-watered controls. In Chardonnay, water decit did not increase ABA concentration above that of well-watered berries. Likewise, sugar and proline concentration were not signicantly different from the well-watered controls. The response of anthocyanins to water decit was unaffected and is irrelevant because Chardonnay berries cannot produce anthocyanins. Water decit increased the transcript abundance of genes (lipoxygenase and hydroperoxide lyase) involved in fatty metabolism, a pathway known to affect berry and wine aromas. In Chardonnay, water decit activated parts of the phenylpropanoid, energy, carotenoid and isoprenoid metabolic pathways that contributed to increased concentrations of antheraxanthin and avonols. The effects of water decit on metabolism had important impacts on berry constituents that inuenced avour and quality characteristics in both grapes and wine and might contribute to increased antioxidants and human-health benets. Regulated-decit irrigation has been shown by several research groups to improve berry and wine quality (Roby et al. 2004, Chapman et al. 2005, Castellarin et al. 2007a,b, Deluc et al. 2009). Application of water decit early in the season before vraison resulted in greater concentrations of anthocyanins and phenolics (Matthews and Anderson 1988, Matthews et al. 1990). Colour differences were the result of increased anthocyanin synthesis caused by water decit applied either early or late in the season (Matthews and Anderson 1988, Castellarin et al. 2007a, Deluc et al. 2009). It was suggested that both ABA and sugar signalling might affect accelerated anthocyanin development. Additions of ABA and rhamnose to grape berry skins have induced anthocyanin biosynthesis
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in a synergistic manner (Hiratsuka et al. 2001) and ABA applications to Crimson Seedless grapes increased their red colour and the transcript abundance of a UDPglucose: avonoid 3-O-glucosyltransferase (UFGT) gene, the rate limiting step in anthocyanin biosynthesis (Peppi et al. 2008). The increase of anthocyanin concentration is a common response in stressed plants and probably functions to provide greater photoprotection (Merzlyak et al. 2008a,b). In red wines, of course, it serves to provide deeper and darker colour, which is often perceived as an indicator of quality by the consumer. A berry tissue analysis using global gene expression techniques indicated that water decit affected the mRNA abundance of 13% of genes at grape maturity within the three tissues of the berry (skin, pulp and seeds), with the greatest changes located in the pulp and skin (Grimplet et al. 2007b). While the function of many of the genes differentially expressed within the seed and pulp remain to be elucidated, other genes over-represented in the skin were clearly associated with phenylpropanoid metabolism, ethylene, pathogenesis-related responses, energy metabolism and stress responses. Proteomic and metabolomic analyses of the berry tissues indicated signicant changes because of water decit (Grimplet et al. 2009). Water decit altered the abundance of approximately 7% of proteins in the pericarp, but had little effect on the expression of proteins in the seeds. Comparison of the protein and transcript expression proles indicated that 32% of the pericarp and 69% of the seed proteins had similar quantitative expression patterns. This indicated that proteins might be inuenced by post-transcriptional processes. There were increases in the relative abundance of proteolytic enzymes in the skin, and the enzyme, cytosolic ascorbate peroxidase. There were increases in isoavone reductase, glutamate decarboxylase and an endochitinase in the pulp. About half of the 32 metabolites measured had differences in abundance between tissues with water decit, including some sugars, amino acids and organic acids. Conclusion Osmotic stress is a common feature of many abiotic stresses that afict grapevines. A systems biology approach has enabled a broad picture of grapevine responses to abiotic stress to be drawn. By comparing and contrasting responses of grapevine to different stress treatments, common and distinct stress-response pathways in grapevine were elucidated. Many of the grapevine responses to osmotic stress appear to be transcriptionally regulated, but proteomic studies indicate that there are post-translational controls as well. ABA acts as a central regulator of many osmotic responses in grapevines. NCED1 appears to be the rate-limiting step for the bulk of ABA biosynthesis, because it is highly correlated with ABA concentrations in both shoots and berries. ABA acts to reduce water loss and increase stress tolerance in grapevines. It reduces stomatal conductance and limits leaf area expansion. It affects the abundance of many transcripts and proteins involved in many different plant processes including osmo- and photoprotection, photosynthesis,

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and growth. One of the most striking features from these analyses is the potential inhibitory effect of ABA on the transcript abundance of nearly all genes involved in protein synthesis that respond to water decit. Modern biology is going through a revolution and grapevine research is no exception. It is expected that we will make much more rapid progress in the development of stress tolerance genotypes in grapevines in the near future, because there are large genetic resources for grapevine and there are a large number of highthroughput genomic tools available to conduct functional genomic analyses. Acknowledgements I would like to thank all my grapevine colleagues who have contributed to such an interesting eld of science. I would especially like to thank my lab personnel and colleagues at UNR for their valuable help in the work cited in this review. I would also like to acknowledge that much of the research from my lab that was cited in this review was supported by a grant (DBI-0217653) from the National Science Foundation (NSF) Plant Genome program.
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Manuscript received: 3 February 2009 Revised manuscript received: 20 April 2009 Accepted: 17 May 2009