Control of plant development and gene expression by sugar signaling

Susan I Gibson
Coordination of development with the availability of nutrients, such as soluble sugars, may help ensure an adequate supply of building materials and energy with which to carry out specic developmental programs. For example, in-vivo and in-vitro experiments suggest that increasing sugar levels delay seed germination and stimulate the induction of owering and senescence in at least some plant species. Higher sugar concentrations can also increase the number of tubers formed by potatoes and can stimulate the formation of adventitious roots by Arabidopsis. New insights into the mechanisms by which sugar-response pathways interact with other response pathways have been provided by microarray experiments examining sugar-regulated gene expression under different light and nitrogen conditions.
Addresses Department of Plant Biology, University of Minnesota, 122 Cargill Building, 1500 Gortner Avenue, St. Paul, Minnesota 55108-1095, USA Corresponding author: Gibson, SI (gibso043@tc.umn.edu)

onset of senescence (reviewed in [2]). Besides affecting the timing with which developmental events occur, soluble sugar levels can also affect organ number and shape. Higher sugar levels may lead to the formation of plants that produce larger and thicker leaves, and increased numbers of tubers and adventitious roots. Some of the most well-characterized examples of developmental processes that are affected by soluble sugar levels are discussed in more detail below and are summarized in Table 1. The mechanisms by which sugars act to inuence plant development and gene expression are just beginning to be deciphered. Understanding of sugar response is complicated by the fact that plants have multiple sugarresponse pathways and that the molecules actually being sensed are not known in all cases. Although glucose and sucrose appear to be sensed directly, in some cases, different sugars or sugar metabolites might sometimes act as the actual signal molecules. For example, recent evidence has implicated trehalose-6-phosphate in the control of some sugar responses (reviewed in [3,4]). In addition, many sugar responses may actually be regulated by alterations in sugar ux [5] or in C:N ratios (reviewed in [6,7]) rather than by absolute sugar or sugar-metabolite levels. Sugar response pathways also interact or exhibit cross-talk with numerous other pathways, including those for phytohormone (reviewed in [810]) and light responses (reviewed in [2,11]). An additional factor that complicates our understanding of sugar responses is that sugars can act by affecting osmotic potentials as well as by functioning as signaling molecules. For example, the inhibitory effects of glucose and sucrose on Arabidopsis hypocotyl elongation in the dark can be mimicked by osmoticum such as sorbitol (L Sommerlad, SI Gibson, unpublished). By contrast, osmoticum cannot completely mimic the effects of sugars on the rate of germination of Arabidopsis seeds [12,13,14,15]. Consequently, appropriate osmotic controls are essential in distinguishing between events that are mediated by sugars acting as osmotica and events in which sugars act via other mechanisms. A better understanding of sugar-response pathways will require the identication and characterization of more of their components. Information regarding sugar-response pathways can be obtained from recent reviews [3,4,9,10,16,1721]. Although not a focus of this review, genetic loci that are believed to affect sugar response are summarized in Table 2.
Current Opinion in Plant Biology 2005, 8:93102

Current Opinion in Plant Biology 2005, 8:93102 This review comes from a themed issue on Growth and development Edited by Liam Dolan and Michael Freeling Available online 19th November 2004 1369-5266/$ see front matter # 2005 Elsevier Ltd. All rights reserved. DOI 10.1016/j.pbi.2004.11.003

Abbreviations ABA ABI2 ATHB13 pho3 SnRK abscisic acid ABA INSENSITIVE2 Arabidopsis thaliana B13 transcription factor phosphate3 SNF1-RELATED PROTEIN KINASE

Introduction
Plants, like other organisms, need to coordinate development with the availability of crucial nutrients, such as soluble sugars. For example, it may be benecial for plants to adjust the timing with which nutrient-intensive events occur to ensure an adequate supply of materials and energy for successful completion of those events. Levels of sugars, such as sucrose, have been postulated to affect the timing with which at least some plant species ower (reviewed in [1]). Soluble sugar levels have also been shown to affect other phase changes, such as the
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94 Growth and development

Table 1 Selected developmental processes postulated to be sugar regulated. Process Embryogenesis Seed germination Seedling development Hypocotyl elongation Leaf formation Nodule growth Pollen development Tuber formation Formation of adventitious root Juvenile-to-adult phase transition Flowering Sugar effects Glucose promotes cell division whereas sucrose promotes cell expansion and reserve deposition in legume cotyledons. Glucose retards the rate of Arabidopsis seed germination but allows seeds to germinate on otherwise inhibitory concentrations of ABA. High concentrations of glucose and sucrose, as well as low concentrations of mannose, inhibit cotyledon expansion, true leaf formation and root growth of young Arabidopsis seedlings. Glucose and sucrose inhibit hypocotyl elongation of dark-grown Arabidopsis seedlings, but the similar effects of sorbitol suggest that this is an osmotic response. Elevated CO2 levels can induce the formation of larger and thicker leaves. Sucrose alleviates the negative effects of nitrate on the growth rates of soybean nodules. Barley plants expressing SnRK1 in an anti-sense orientation abort pollen development. Elevated sucrose concentrations can lead to the formation of increased numbers of potato tubers. Sucrose, glucose and fructose, but not mannose or sorbitol, stimulate the formation of adventitious roots on the hypocotyls of dark-grown Arabidopsis seedlings. Exogenous glucose prolongs the juvenile growth phase of the moss Physcomitrella patens. Most studies suggest that increasing sucrose levels help to induce owering. Exogenous sucrose allows Arabidopsis to ower in complete darkness. Trehalose-6-phosphate synthase activity is required for the owering of Arabidopsis. Glucose stimulates senescence in combination with low concentrations of nitrogen.

Induction of senescence

Seed and embryo development

The levels of soluble sugars, such as glucose and sucrose, are known or postulated to regulate developmental processes spanning from embryo development to senescence. The role of soluble sugars in embryo development has predominantly been studied using large-seeded legumes as models (reviewed in [8,22]). In these plants, development has been found to proceed in a wave-like fashion across the cotyledons. Using bioluminescence and single-photon counting, glucose concentrations were shown to be greatest in nondifferentiated mitotically active cells, whereas sucrose concentrations increase as cells mature and begin to accumulate starch storage reserves. On the basis of these ndings, soluble sugars have been postulated to act as regulatory molecules that help to control seed and embryo development. One of the most fascinating aspects of these ndings is that they suggest that glucose and sucrose act in almost opposite fashions during legume seed development, with glucose promoting cell division and sucrose being associated with cell expansion and starch synthesis [23,24]. Whether the seeds of other species respond to sugar gradients in a similar manner remains unclear, in part due to difculties in measuring tissue-specic sucrose and hexose concentrations during the development of small seeds such as those produced by Brassica napus and Arabidopsis thaliana [25,26]. The mechanisms by which sugar gradients are generated, as well as how they act to help regulate seed development, are currently being investigated. Tissue-specic and temporally regulated expression of hexose transporters and invertases might play a role in establishing and maintaining sugar gradients [27]. Interestingly, Snf1related protein kinases (SnRKs) have been postulated
Current Opinion in Plant Biology 2005, 8:93102

to be induced by high sucrose concentrations or low glucose concentrations, suggesting that these proteins could act as part of a mechanism to allow differential responses to sucrose and glucose (reviewed in [28]). Cross-talk with other response pathways, particularly phytohormone response pathways, has also been shown to be important in regulating response to sugars (reviewed in [9,10,29]). The manner in which phytohormone and sugar signals are integrated appears to be complex, with the roles of some loci not apparent when examining plants that carry mutations in only a single locus but becoming evident when plants carrying mutations in multiple loci are examined [30]. Another type of complexity arises from the likelihood that some sugar responses could actually be regulated by uctuations in the levels of other metabolites that vary in concentration in parallel with changing sugar levels. For example, alterations in sugar levels are also expected to affect the levels of other metabolites, such as ATP. In fact, ATP levels have recently been shown to vary across developing barley seeds, with higher ATP levels being found in regions in which storage starch is beginning to form [31].

Seed germination and early seedling development

The levels of glucose and other sugars have been shown to affect seed germination and early seedling development. The effects of sugars on these processes are proving to be complicated. Sugars appear to exert a positive effect in some assays but negative effects in other assays. In some cases, different concentrations of sugars may exert varying effects. In addition, sugars appear to affect these developmental processes via more than one pathway. With hindsight, this complexity of control is probably not surprising given the complex nature of the many
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Control of plant development and gene expression by sugar signaling Gibson 95

Table 2 Selected loci postulated to affect plant sugar responses. Loci/genes abi3 Selected sugar-response defects Some loss-of-function mutations confer resistance to the combined inhibitory effects of glucose and ABA on early seedling development; overexpression confers glucose-hypersensitive early seedling development. Loss-of-function mutations confer sugar-insensitivity, and overexpression confers sugar-hypersensitivity and early seedling development. Loss-of-function mutations confer slight sugar-insensitivity, and overexpression confers sugar-hypersensitivity and early seedling development. Seedling growth defect partially rescued by glucose but not by sucrose. Sensitive to sucrose-induced chlorosis. ATHB13 overexpression inhibits the expansion of cotyledon cells in sucrose-grown plants and leads to hypersensitive sucrose induction of b-amylase and vegetative storage protein expression. Glucose- and sucrose-insensitive early seedling development. Glucose-hypersensitive early seedling development. Sucrose-hypersensitive early seedling development. Glucose-hypersensitive early seedling development. Glucose- and sucrose-hypersensitive early seedling development. Glucose-hypersensitive early seedling development and chlorophyll accumulation. Glucose-insensitive early seedling development Glucose-insensitive early seedling development Glucose-insensitive early seedling development Glucose-insensitive early seedling development Glucose-insensitive early seedling development Anti-sense AtGLR1.1 expression prevents seed germination on sucrose, but not on glucose, sorbitol or mannitol Hypersensitive to sugar-inducible expression of b-amylase Hypersensitive to sugar-inducible expression of ADP-glucose pyrophosphorylase subunit ApL3; sugar-hypersensitive early seedling development Hypersensitive to sugar-inducible expression of ADP-glucose pyrophosphorylase subunit ApL3; sugar-hypersensitive early seedling development Hypersensitive to sugar-inducible expression of ADP-glucose pyrophosphorylase subunit ApL3; sugar-hypersensitive early seedling development Hypersensitive to sugar-inducible expression of ADP-glucose pyrophosphorylase subunit ApL3; sugar-hypersensitive early seedling development Glucose-hypersensitive early seedling development Anti-sense AtHXK1 expression confers glucose-insensitivity, and overexpression confers glucose-hypersensitivity and early seedling development Resistant to sugar-inducible expression of ADP-glucose pyrophosphorylase subunit ApL3 Resistant to sugar-inducible expression of ADP-glucose pyrophosphorylase subunit ApL3 Resistant to sugar-inducible expression of ADP-glucose pyrophosphorylase subunit ApL3; sucrose-insensitive early seedling development Resistant to sugar-inducible expression of ADP-glucose pyrophosphorylase subunit ApL3; sucrose insensitive early seedling development Resistant to sugar-inducible expression of b-amylase Predicted gene products Transcription factor Reference(s) [81,82]

abi4

Transcription factor

[36,38,81,83,84]

abi5

Transcription factor

[36,38,81,85]

abi8 acn3 ATHB13

Protein function unknown Protein function unknown Transcription factor

[44] [86] [45]

ctr1 ein2 ein4 eto1 etr1 ghs1 gin1 gin2 gin4 gin5 gin6 GLR1.1 hba1 hsr1

Protein kinase Membrane-anchored protein Histidine protein kinase, ethylene receptor Inhibitor of 1-aminocyclopropane1-carboxylic acid synthase Histidine protein kinase, ethylene receptor Plastid 30S ribosomal protein S21 Short chain dehydrogenase/ reductase (= ABA2) Hexokinase (= HXK1) Protein kinase (= CTR1) Molybdenum cofactor sulphurase (= ABA3) Transcription factor (= ABI4) Putative glutamate receptor Gene product not identied Gene product not identied (locus maps to chromosome 1) Gene product not identied (locus maps to chromosome 4) Gene product not identied (locus maps to chromosome 1) Gene product not identied (locus maps to chromosome 1) Putative nuclear-localized membrane protein Hexokinase

[40,41,76] [87] [41] [40] [40,41] [88] [40,87] [72] [76] [38,76] [38] [89] [90] [91]

hsr2

[91]

hsr3

[91]

hsr4

[91]

hys1 HXK1

[70] [37,72]

isi1 isi2 isi3

Gene product not identied (locus maps to chromosome 4) Gene product not identied (locus maps to chromosome 1) Transcription factor (= ABI4)

[84] [84] [84]

isi4

Short chain dehydrogenase/ reductase (= ABA2) Gene product not identied (locus maps to chromosome 5)

[84]

lba1

[92]

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96 Growth and development

Table 2 Continued Loci/genes lba2 MYB10 prl1 ram1 rgs1 rsr1 rsr4 sis1 sis4 sis5 SnRK1 sun1 sun6 sun7 Selected sugar-response defects Resistant to sugar-inducible expression of b-amylase CpMYB10 overexpression confers glucose-insensitive early seedling development Sugar-hypersensitive early seedling development Resistant to sugar-inducible b-amylase expression Overexpression confers glucose-hypersensitivity, and loss-of-function confers sugar-insensitivity and early seedling development Resistant to sugar-inducible patatin expression Resistant to sugar-inducible patatin expression Glucose- and sucrose-insensitive early seedling development Glucose- and sucrose-insensitive early seedling development Glucose- and sucrose-insensitive early seedling development Anti-sense PKIN1 expression confers sucrose-insensitive expression of sucrose synthase Resistant to sucrose repression of plastocyanin expression Resistant to sucrose repression of plastocyanin expression; sucrose-insensitive early seedling development Resistant to sucrose repression of plastocyanin expression Predicted gene products Gene product not identied Transcription factor WD-40 protein b-amylase G-protein coupled receptor Gene product not identied (locus maps to chromosome 1) Gene product not identied (locus maps to chromosome 5) Protein kinase (= CTR1) Short chain dehydrogenase/ reductase (= ABA2) Transcription factor (= ABI4) Protein kinase Gene product not identied Transcription factor (= ABI4) Gene product not identied Reference(s) [92] [93] [39,94,95] [96] [97] [98] [98] [41] [36] [36] [99] [100] [83,100] [100]

Abbreviations: acn3, acetate non-utilization3; ein2, ethylene insensitive2; eto1, ethylene overproducer1; etr1, ethylene receptor1; ghs1, glucose hypersensitive1; gin1, glucose insensitive1; GLR1.1, GLUTAMATE RECEPTOR 1.1; hba1, high b-amylase1; hsr1, high sugar response1; hys1, hypersenescence1; HXK1, HEXOKINASE1; isi1, impaired sucrose induction1; lba1, low b-amylase1; prl1, pleiotropic regulatory locus1; ram1, reduced b-amylase1; rgs1, regulator of G-protein signaling1; rsr1, reduced sucrose responses1; sis1, sugar insensitive1; sun1, sucrose uncoupled1.

developmental and metabolic changes that take place during these developmental stages. Although high concentrations (e.g. 0.3 M) of exogenous glucose and sucrose do not prevent seed germination, these concentrations of sugars delay germination signicantly. By contrast, equi-molar concentrations of sorbitol do not exert similar effects, indicating that the effects of high sugar concentrations on seed germination rates are not solely due to osmotic stress [14,15]. More recently, much lower glucose concentrations (e.g. 3060 mM) have been shown to cause measurable delays in seed germination [12,13]. Interestingly, the glucose analog 3-Omethylglucose at concentrations of about 84 mM was also shown to delay seed germination [12]. As 3-O-methylglucose is a poor substrate for hexokinase, with a catalytic efciency rate that is ve orders of magnitude lower than that for glucose [32], this result has been interpreted as suggesting that the delay of seed germination by relatively low concentrations of exogenous glucose and 3-Omethylglucose may occur via a hexokinase-independent mechanism [12]. By contrast, earlier studies examining the effects of lower concentrations of glucose analogs found that mannose, which is a much better substrate than 3-O-methylglucose for hexokinase [32], is able to retard seed germination signicantly at concentrations of 510 mM, whereas 10 mM 3-O-methylglucose has no signicant effects on rates of seed germination. On the basis of these ndings, the authors concluded that inhibition of seed germination by mannose occurs via a hexokinase-dependent pathway [33]. Together, these studies
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suggest that different sugars may delay seed germination via different pathways, and that the concentration of sugar necessary to exert a noticeable effect varies between different sugar species. Interactions between sugar- and phytohormone-response pathways during seed germination are similarly complex. In earlier studies, glucose was found to allow wildtype seeds to germinate on otherwise inhibitory concentrations of abscisic acid (ABA) [34,35]. These ndings indicate that, in the presence of exogenous ABA, glucose is able either to stimulate seed germination or to alleviate the inhibitory effects of ABA on seed germination. These ndings are in contrast to the results described above, in which glucose and other sugars were found to exert negative effects on seed germination. Furthermore, more recent studies suggest that exogenous glucose retards the rate at which endogenous ABA is broken down in germinating seeds [13]. On the basis of these results, glucose might be expected to exacerbate rather than to alleviate the negative effects of ABA on seed germination. Characterization of the effects of exogenous glucose on the seed germination rates of different ABA-metabolic and ABA-response mutants has, however, suggested that glucose retards seed germination via a pathway that does not involve the ABA-response pathway components ABI2, ABI4 and ABI5 [12,13,30]. Resolution of some of these apparent discrepancies will require more information about the mechanisms by which sugars, phytohormones and other factors regulate seed germination. Given the number of dramatic developmental and metabolic shifts
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Control of plant development and gene expression by sugar signaling Gibson 97

that occur during germination, the possibility that sugars and phytohormones may affect different processes in different ways and via multiple mechanisms seems not unlikely. Although exogenous sugars do retard seed germination, given sufcient time the majority of wildtype seeds will germinate on even very high concentrations of sugars. For example, almost 100% of wildtype Arabidopsis seeds of the Columbia ecotype germinate in the presence of 300 mM glucose or sucrose [36]. However, the majority of seedlings formed on these media fail to form expanded cotyledons, true leaves or extensive root systems [3641]. In addition, these seedlings fail to develop chloroplasts [42] and do not mobilize the majority of their seed storage lipid [14,43]. Although equi-molar concentrations of mannitol and sorbitol do not exert the same effects, higher concentrations of sorbitol (0.40.5 M) do exert similar effects. These results suggest that some, but not all, of the inhibitory effects of exogenous glucose and sucrose on early seedling development are due to osmotic stress [36]. This glucose- and sucrose-mediated developmental arrest has been used to screen for sugar hypersensitive and insensitive mutants. Characterization of these mutants has revealed that many of them are also defective in phytohormone metabolism or response (reviewed in [9,10,18,29]). Most recently, mutations in the ABI8 gene of Arabidopsis were found to confer resistance to both ABA inhibition of seed germination and glucose inhibition of early seedling development [44].

leaf and cotyledon shape only in response to metabolizable sugars [45]. However, as the concentrations of mannose and 3-O-methylglucose used in these experiments were much lower than those of glucose and sucrose, interpretation of the results of these experiments is complicated. Sugar levels also inuence the normal growth and development of nodules and pollen. When nitrate is added to the media of hydroponically grown soybeans, the rate of nodule growth is signicantly reduced. The negative effects of nitrate on nodule growth rates can be reduced by the addition of sucrose to the media, suggesting that nodule growth rates are partially dependent on carbon availability:nitrogen availability [46]. Evidence that sugar-response pathways may play a role in pollen development is provided by experiments on transgenic barley expressing the protein kinase SnRK1 in an anti-sense orientation [47]. Pollen carrying the transgene cease development at the binucleate stage. Ovule development may also be affected by the presence of the SnRK1 antisense transgene, as none of the progeny of the transgenic plants inherit a copy of the transgene [47]. As SnRK1 is believed to function in metabolic response pathways (reviewed in [16]), these results suggest that pollen and ovule development are dependent on normal metabolic response. In addition to affecting the growth or development of organs that would normally form on a plant, sugar levels can cause the formation of extra organs, such as extra tubers and adventitious roots. Several lines of evidence have led to the hypothesis that increasing sucrose concentrations correlate with formation of greater numbers of tubers (reviewed in [9,48,49]). This effect has been shown relatively directly by in-vitro experiments in which the application of exogenous sucrose resulted in the formation of increased numbers of tubers [50,51]. Antisense expression of ADP-glucose pyrophosphorylase and overexpression of invertase have also been used to manipulate sugar levels, with concomitant alterations in tuber size and number [52,53]. However, increasing sucrose concentrations do not always correlate with the formation of greater numbers of tubers. Constitutive overexpression of a spinach sucrose transporter in transgenic potatoes led to elevated sugar levels in the tubers but no signicant alterations in tuber size or total yield [54]. The reason that these experiments, unlike the experiments described above, showed no correlation between sugar levels and tuber formation remains unclear. Interestingly, underand overexpression of hexokinases 1 and 2 also failed to alter tuber yield. These experiments suggest that hexokinases do not play a major role in the regulation of tuber formation [55,56]. Exogenous sugars also stimulate the formation of adventitious roots under certain conditions [57]. When
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Effects of sugars on the formation of adult organs and tissues

In addition to mediating early developmental events, soluble sugars also effect the formation of more adult structures, such as leaves, nodules, pollen, tubers and roots. For example, growth at elevated CO2 concentrations, which presumably increases sugar production, sometimes leads to the formation of larger and thicker leaves (reviewed in [2]). The homeodomain leucine zipper transcription factor ATHB13 of Arabidopsis has been implicated in regulation of leaf shape in response to sugar levels [45]. Overexpression of this gene in transgenic Arabidopsis leads to the formation of narrow cotyledons and leaves when plants are grown in the presence of exogenous sugars such as glucose and sucrose. Further examination of these transgenic plants revealed that sucrose inhibits lateral expansion of cotyledonary cells in plants overexpressing ATHB13. Sorbitol and mannitol do not exert similar effects, indicating that the effects of glucose and sucrose are not due to alterations in osmotic potential. Similarly, no signicant differences in the leaves or cotyledons of wildtype and ATHB13-overexpressing plants were observed when the plants were grown in the presence of the non-metabolizable sugar analogs mannose and 3-O-methylglucose. These results were interpreted as indicating that ATHB13 modulates
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98 Growth and development

wildtype Arabidopsis seedlings are grown in the dark on media containing around 1560 mM sucrose, the formation of adventitious roots on the hypocotyls is stimulated. Glucose and fructose, but not mannose or sorbitol, also increase the formation of adventitious roots, indicating that the effect is not due to alterations in the osmotic potential of the media and suggesting that only metabolizable sugars are effective in inducing adventitious root formation. In addition, direct contact of the hypocotyls with the sugar-containing media is necessary for the stimulation of adventitious root formation. The effects of sugars on the formation of adventitious roots are concentration dependent; higher concentrations of sucrose (e.g. 150 mM) suppress rather than stimulate induction [57]. High concentrations (300 mM) of sucrose also inhibit the elongation of carrot radicles. In this case, however, glucose, fructose and maltose did not exert similar effects [58].

owering by raising sugar concentrations above optimal levels, the nding that 30 mM sucrose also inhibits owering is in contrast to previous reports and will require further experimentation to explain. The roles of other sugars and sugar metabolites in owering are also being investigated. Recently, Arabidopsis plants carrying mutations in the trehalose-6-phosphate synthase gene were shown to be unable to ower, indicating that this gene is essential for the transition to owering in Arabidopsis [66]. The effects of sugar levels on leaf senescence are complex. Endogenous leaf sugar levels tend to increase during senescence. Similarly, application of exogenous sugars stimulates the early stages of senescence, but can inhibit the expression of some senescence-associated genes in the later stages of senescence (reviewed in [2,67]). Senescence can also be stimulated by low nitrogen concentrations [68], suggesting that senescence may be regulated more by C:N ratios than by absolute sugar levels. For example, application of moderate (111 mM) concentrations of glucose stimulates the senescence of Arabidopsis, but only if nitrogen levels are low [69]. Genetic evidence for a link between sugar response and senescence is provided by ndings that Arabidopsis hypersenescence1 (hys1) mutants senesce early and are hypersensitive to the inhibitory effects of exogenous sugars on early seedling development [70]. Recent studies are beginning to provide clues as to some of the molecular mechanisms by which sugars and other factors regulate senescence. Hexokinase has been implicated in the regulation of senescence because transgenic tomato plants that overexpress Arabidopsis HEXOKINASE1 senesce earlier [71], and Arabidopsis mutants with decreased HEXOKINASE1 activity senesce later [72], than wildtype plants. As sugars and ABA have been shown to interact in the regulation of other developmental programs, the roles of several components of the ABAresponse pathway in sugar-mediated regulation of senescence have been investigated. These studies suggest that Arabidopsis ABI5, but not ABI4, acts in the sugarmediated regulation of senescence [73]. A role for sugar metabolism in the regulation of senescence by cytokinin has also been identied. An extracellular invertase is required for the cytokinin-mediated delay of senescence: cytokinins no longer delay senescence in the presence of an inhibitor of that invertase [74].

Effects of sugars on timing of developmental events

Sugars also help to regulate the timing of developmental phase changes, such as the progression from juvenile to adult phases, owering and senescence. When the RUBISCO small subunit was expressed in an antisense orientation in tobacco, leaf source strength decreased with a concomitant extension in the length of an early phase of shoot development [59]. More recently, knockouts of a cyclin D gene in the moss Physcomitrella patens have suggested that cyclin D helps to integrate metabolism and development in this organism. Whereas wildtype Physcomitrella patens exhibits a prolonged juvenile phase when grown in the presence of glucose, the knockout strain is insensitive to this glucose-mediated developmental delay [60]. Similarly, sugar levels have been postulated to affect the timing of owering in at least some plant species (reviewed in [1]). In studies on Arabidopsis, a species that owers earlier under long-day conditions, a correlation was found between greater export of carbohydrates from the leaves and increased ower induction [61]. Similarly, the C:N ratios of leaf exudates from Arabidopsis and Sinapis alba increase under owering-inductive conditions [62]. Sugar levels have also been shown to affect owering in autonomously owering species, such as tomato. In-vitro studies on tomato revealed that optimal sucrose and nitrogen levels are necessary to promote owering [63]. Application of sucrose to the apical part of the plant has also been shown to allow owering of Arabidopsis in complete darkness [64], lending further support to the hypothesis that sugars promote owering. In other studies, however, high (e.g. 150 mM) concentrations of exogenous sucrose signicantly inhibited, and low (e.g. 30 mM) concentrations of sucrose slightly inhibited, owering of wildtype Arabidopsis [65]. Although it is possible that 150 mM sucrose could inhibit
Current Opinion in Plant Biology 2005, 8:93102

Sugar-regulation of gene expression

Analyses of limited sets of genes suggested that signicant numbers of plant genes are regulated at the steadystate mRNA level in response to sugar levels. In addition, several lines of evidence have suggested the existence of hexokinase-dependent, hexokinase-independent and sucrose-specic signaling pathways (reviewed in [6,7,9, 10,18,20,21,75,76]). More recently, Affymetrix GeneChips have been used to investigate sugar-regulated gene
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Control of plant development and gene expression by sugar signaling Gibson 99

expression in Arabidopsis on a more global scale. The Arabidopsis phosphate3 (pho3) mutant was used to examine gene expression in adult plants in response to alterations in endogenous sugar levels. The pho3 mutant carries a mutation in SUC2, which encodes a sucrose transporter that functions in phloem loading. Mutant plants accumulate high concentrations of sucrose, glucose and fructose. Mutant and wildtype plants were grown in soil under long-day (16 h light) conditions until the rst owers began to open. Rosette leaves were then harvested and gene expression analyzed using Affymetrix ATH1 Arabidopsis GeneChips, which carry 22 500 probe sets. Small changes were detected in the expression of many genes, including some that are involved in carbon metabolism [77]. GeneChips have also been used to examine interactions between sugar- and light-regulated gene expression. Sixteen-day-old Arabidopsis plants were induced for eight hours by treatment with 0 mM or 30 mM sucrose in continuous light or the dark before harvesting whole seedlings. Gene expression was analyzed using Affymetrix AG GeneChips, which contain information from approximately 8000 Arabidopsis genes. These experiments indicate that many genes are regulated by interactions between light and sugar signaling, and that genes that are involved in metabolism are over-represented within this group [78]. Additional experiments have investigated interactions between sugar and nitrogen signaling. Six-day-old Arabidopsis seedlings were induced for three hours by treatment with glucose, nitrogen, glucose + nitrogen, 3-O-methylglucose or nothing in the presence or absence of the protein synthesis inhibitor cycloheximide. Gene expression was monitored in whole seedlings using Affymetrix ATH1 GeneChips. These experiments indicate that glucose treatment has stronger effects on gene expression than nitrogen treatment [79]. However, the possibility that different concentrations of glucose or nitrogen might affect the relative magnitudes of the observed effects should be considered. These experiments also indicate that cycloheximide affects the expression of glucose-induced genes more than that of glucose-repressed genes, suggesting that de novo protein synthesis is more typically required for glucose induction than for repression [79]. Interactions between carbon and nitrogen signaling were also suggested by studies examining nitrate response. In these experiments, ten-day-old Arabidopsis seedlings were induced for 20 min with nitrate or a negative control. Shoot and root tissue were harvested and analyzed separately using Affymetrix ATH1 GeneChips. Many genes that are involved in glucose and trehalose metabolism were shown to be nitrate regulated, suggesting the presence of mechanisms to coordinate nitrogen and carbon metabolism [80].
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Conclusions
Alterations in the levels of soluble sugars, such as glucose and sucrose, have been shown to affect developmental programs ranging from embryogenesis to senescence. In some cases, sugar levels or ux appear to determine whether an event occurs (e.g. whether additional tubers or adventitious roots are formed) or the timing with which an event occurs (e.g. the timing of owering and senescence). As the role of sugar levels in many developmental processes has not yet been examined, the number of developmental processes postulated to be affected by sugar levels is expected to grow in the future. A potential mechanism for identifying additional sugar-regulated developmental events will be to scan lists of sugar-regulated genes that are identied by microarray experiments for genes known to be involved in different developmental events. Also of future interest will be more precise information regarding the natures of the signals that trigger different sugar responses. Questions that need to be answered include which sugars or sugar metabolites are actually being sensed, whether sugar levels or C:N ratios are being measured and whether absolute sugar levels or ux are being sensed. A better understanding of the nature of the signals that are associated with different sugar responses will be useful in characterizing sugarresponse pathways.

Acknowledgements
Research in the authors laboratory in this area is supported by the Energy Biosciences Program of the US Department of Energy.

References and recommended reading

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A very recent review that summarizes information on the role of metabolites in regulating seed and embryo development. The authors include a good description of metabolic imaging studies that have led to the hypothesis that glucose and sucrose form gradients that help to regulate the development of legume cotyledons. 9. Gibson SI: Sugar and phytohormone response pathways:  navigating a signaling network. J Exp Bot 2004, 55:253-264. This review discusses interactions between sugar and phytohormone response pathways in the regulation of a-amylase activity, embryogenesis, seed germination, early seedling development and tuberization. on P, Sheen J: Sugar and hormone connections. Trends Plant 10. Le  Sci 2003, 8:110-116. An informative review of the mechanisms by which sugars and phytohormones, particularly abscisic acid and ethylene, interact to regulate processes such as seedling development. 11. Ellis C, Turner JG, Devoto A: Protein complexes mediate signalling in plant responses to hormones, light, sucrose and pathogens. Plant Mol Biol 2002, 50:971-980. 12. Dekkers BJW, Schuurmans JAMJ, Smeekens SCM: Glucose  delays seed germination in Arabidopsis thaliana. Planta 2004, 218:579-588. The work described in this paper demonstrates that low, physiologically relevant, concentrations of glucose signicantly retard germination rates of wildtype Arabidopsis seeds. The authors present evidence that glucose acts in this process via a hexokinase-independent mechanism. 13. Price J, Li T-C, Kang SG, Na JK, Jang J-C: Mechanisms of  glucose signaling during germination of Arabidopsis. Plant Physiol 2003, 132:1424-1438. The authors demonstrate that glucose delays the germination of wildtype Arabidopsis seeds at low, physiologically relevant, concentrations. They present evidence that glucose might affect seed germination rates by retarding the degradation of endogenous abscisic acid. 14. To JPC, Reiter W-D, Gibson SI: Mobilization of seed storage lipid by Arabidopsis seedlings is retarded in the presence of exogenous sugars. BMC Plant Biol 2002, 2:4. 15. Ullah H, Chen J-G, Wang S, Jones AM: Role of a heterotrimeric G protein in regulation of Arabidopsis seed germination. Plant Physiol 2002, 129:897-907. 16. Halford NG, Hey S, Jhurreea D, Laurie S, McKibbin RS, Paul M,  Zhang Y: Metabolic signalling and carbon partitioning: role of Snf1-related (SnRK1) protein kinase. J Exp Bot 2003, 54:467-475. SnRK1s have been shown to play key roles in metabolic response in a variety of organisms. This review provides a useful overview of the possible roles of these proteins in regulating carbon partitioning and plant development. 17. Moore BD: Bifunctional and moonlighting enzymes: lighting the way to regulatory control. Trends Plant Sci 2004, 9:221-228. 18. Rook F, Bevan MW: Genetic approaches to understanding sugar-response pathways. J Exp Bot 2003, 54:495-501. 19. Koch K: Sucrose metabolism: regulatory mechanisms and pivotal roles in sugar sensing and plant development. Curr Opin Plant Biol 2004, 7:235-246. 20. Smeekens S: Sugar-induced signal transduction in plants. Annu Rev Plant Physiol Plant Mol Biol 2000, 51:49-81. 21. Pego JV, Kortstee AJ, Huijser G, Smeekens SCM: Photosynthesis, sugars and the regulation of gene expression. J Exp Bot 2000, 51:407-416. 22. Hills MJ: Control of storage-product synthesis in seeds. Curr Opin Plant Biol 2004, 7:302-308. 23. Borisjuk L, Walenta S, Rolletschek H, Mueller-Klieser W, Wobus U, Weber H: Spatial analysis of plant metabolism: sucrose imaging within Vicia faba cotyledons reveals specic developmental patterns. Plant J 2002, 29:521-530. 24. Borisjuk L, Rolletschek H, Wobus U, Weber H: Differentiation  of legume cotyledons as related to metabolic gradients and assimilate transport into seeds. J Exp Bot 2003, 54:503-512. Measurements of sugar concentrations across developing legume cotyledons revealed the existence of glucose and sucrose gradients. Higher Current Opinion in Plant Biology 2005, 8:93102

concentrations of glucose tend to correlate with non-differentiated cells, whereas sucrose levels become higher as cells begin to accumulate storage reserves. The nding that glucose and sucrose play very different roles in the regulation of legume cotyledon development is of particular interest, as glucose and sucrose are thought to exert the same effects on most other processes. 25. Baud S, Boutin J-P, Miquel M, Lepiniec L, Rochat C: An integrated overview of seed development in Arabidopsis thaliana ecotype WS. Plant Physiol Biochem 2002, 40:151-160. 26. Hill LM, Morley-Smith ER, Rawsthorne S: Metabolism of sugars in the endosperm of developing seeds of oilseed rape. Plant Physiol 2003, 131:228-236. 27. Weschke W, Panitz R, Gubatz S, Wang Q, Radchuk R, Weber H,  Wobus U: The role of invertases and hexose transporters in controlling sugar ratios in maternal and lial tissues of barley caryopses during early development. Plant J 2003, 33:395-411. The authors investigate the role of the tissue-specic and temporally regulated expression of hexose transporters and invertase in modulating the concentrations of different sugars during seed development. 28. Halford NG, Dickinson JR: Sugar sensing and cell cycle control: evidence of cross-talk between two ancient signalling pathways. In The Plant Cell Cycle and Its Interfaces. Edited by Francis D. Shefeld: Shefeld Academic Press; 2001:87-107. 29. Finkelstein RR, Gibson SI: ABA and sugar interactions regulating development: cross-talk or voices in a crowd? Curr Opin Plant Biol 2002, 5:26-32. 30. Brocard-Gifford IM, Lynch TJ, Finkelstein RR: Regulatory  networks in seeds integrating developmental, abscisic acid, sugar, and light signaling. Plant Physiol 2003, 131:78-92. The authors examine the effects of mutations in different combinations of ABA-response pathway components on ABA and sugar response. 31. Rolletschek H, Weschke W, Weber H, Wobus U, Borisjuk L: Energy state and its control on seed development: starch accumulation is associated with high ATP and steep oxygen gradients within barley grains. J Exp Bot 2004, 55:1351-1359. s S, Gromova M, Evrard A, Roby C, Heyraud A, Rolin DB, 32. Corte Raymond P, Brouquisse RM: In plants, 3-O-methylglucose is phosphorylated by hexokinase but not perceived as a sugar. Plant Physiol 2003, 131:824-837. 33. Pego JV, Weisbeek PJ, Smeekens SCM: Mannose inhibits Arabidopsis germination via a hexokinase-mediated step. Plant Physiol 1999, 119:1017-1023. 34. Finkelstein RR, Lynch TJ: Abscisic acid inhibition of radicle emergence but not seedling growth is suppressed by sugars. Plant Physiol 2000, 122:1179-1186. 35. Garciarrubio A, Legaria JP, Covarrubias AA: Abscisic acid inhibits germination of mature Arabidopsis seeds by limiting the availability of energy and nutrients. Planta 1997, 203:182-187. 36. Laby RJ, Kincaid MS, Kim D, Gibson SI: The Arabidopsis sugar-insensitive mutants sis4 and sis5 are defective in abscisic acid synthesis and response. Plant J 2000, 23:587-596. n P, Zhou L, Sheen J: Hexokinase as a sugar 37. Jang J-C, Leo sensor in higher plants. Plant Cell 1997, 9:5-19. n P: Analysis of 38. Arenas-Huertero F, Arroyo A, Zhou L, Sheen J, Leo Arabidopsis glucose insensitive mutants, gin5 and gin6, reveals a central role of the plant hormone ABA in the regulation of plant vegetative development by sugar. Genes Dev 2000, 14:2085-2096. meth K, Salchert K, Putnoky P, Bhalerao R, Koncz-Ka lma n Z, 39. Ne kre L, Mathur J, O sz L, Stabel S et Stankovic-Stangeland B, Bako al.: Pleiotropic control of glucose and hormone responses by PRL1, a nuclear WD protein, in Arabidopsis. Genes Dev 1998, 12:3059-3073. 40. Zhou L, Jang J-C, Jones TL, Sheen J: Glucose and ethylene signal transduction crosstalk revealed by an Arabidopsis glucose-insensitive mutant. Proc Natl Acad Sci USA 1998, 95:10294-10299. www.sciencedirect.com

Control of plant development and gene expression by sugar signaling Gibson 101

41. Gibson SI, Laby RJ, Kim D: The sugar-insensitive1 (sis1) mutant of Arabidopsis is allelic to ctr1. Biochem Biophys Res Commun 2001, 280:196-203. 42. To JPC, Reiter W-D, Gibson SI: Chloroplast biogenesis by  Arabidopsis seedlings is impaired in the presence of exogenous glucose. Physiol Plant 2003, 118:456-463. Exogenous glucose had previously been shown to inhibit the greening of tissues. The work reported in this manuscript indicates that, in fact, exogenous glucose can severely inhibit the formation of mature chloroplasts by young Arabidopsis seedlings. 43. Martin T, Oswald O, Graham IA: Arabidopsis seedling growth, storage lipid mobilization, and photosynthetic gene expression are regulated by carbon:nitrogen availability. Plant Physiol 2002, 128:472-481. 44. Brocard-Gifford I, Lynch TJ, Garcia ME, Malhotra B, Finkelstein RR: The Arabidopsis thaliana ABSCISIC ACID-INSENSITIVE8 locus encodes a novel protein mediating abscisic acid and sugar responses essential for growth. Plant Cell 2004, 16:406-421. m P: Sugar-dependent 45. Hanson J, Johannesson H, Engstro alterations in cotyledon and leaf development in transgenic plants expressing the HDZhdip gene ATHB13. Plant Mol Biol 2001, 45:247-262. 46. Fujikake H, Yamazaki A, Ohtake N, Sueyoshi K, Matsuhashi S,  Ito T, Mizuniwa C, Kume T, Hashimoto S, Ishioka N-S et al.: Quick and reversible inhibition of soybean root nodule growth by nitrate involves a decrease in sucrose supply to nodules. J Exp Bot 2003, 54:1379-1388. Nitrate was previously believed to inhibit nodule formation. Work described in this manuscript demonstrates that the addition of exogenous sucrose to the growth media can alleviate the negative effects of nitrate on soybean nodule formation. These results suggest that C:N ratios play an important role in the regulation of nodule formation. 47. Zhang Y, Shewry PR, Jones H, Barcelo P, Lazzeri PA, Halford NG: Expression of antisense SnRK1 protein kinase sequence causes abnormal pollen development and male sterility in transgenic barley. Plant J 2001, 28:431-441. 48. Jackson SD: Multiple signaling pathways control tuber induction in potato. Plant Physiol 1999, 119:1-8. 49. Fernie AR, Willmitzer L: Molecular and biochemical triggers of potato tuber development. Plant Physiol 2001, 127:1459-1465. 50. Xu X, van Lammeren AAM, Vermeer E, Vreugdenhil D: The role of gibberellin, abscisic acid, and sucrose in the regulation of potato tuber formation in vitro. Plant Physiol 1998, 117:575-584. 51. Simko I: Sucrose application causes hormonal changes associated with potato tuber induction. J Plant Growth Regul 1994, 13:73-77. 52. Sonnewald U, Hajirezaei M-R, Kossmann J, Heyer A, Trethewey RN, Willmitzer L: Increased potato tuber size resulting from apoplastic expression of a yeast invertase. Nat Biotechnol 1997, 15:794-797. ber B, Sonnewald U, Willmitzer L: Inhibition of the ADP ller-Ro 53. Mu glucose pyrophosphorylase in transgenic potatoes leads to sugar-storing tubers and inuences tuber formation and expression of tuber storage protein genes. EMBO J 1992, 11:1229-1238. 54. Leggewie G, Kolbe A, Lemoine R, Roessner U, Lytovchenko A, Zuther E, Kehr J, Frommer WB, Riesmeier JW, Willmitzer L et al.: Overexpression of the sucrose transporter SoSUT1 in potato results in alterations in leaf carbon partitioning and in tuber metabolism but has little impact on tuber morphology. Planta 2003, 217:158-167. 55. Veramendi J, Roessner U, Renz A, Willmitzer L, Trethewey RN: Antisense repression of hexokinase 1 leads to an overaccumulation of starch in leaves of transgenic potato plants but not to signicant changes in tuber carbohydrate metabolism. Plant Physiol 1999, 121:123-133. 56. Veramendi J, Fernie AR, Leisse A, Willmitzer L, Trethewey RN: Potato hexokinase 2 complements transgenic Arabidopsis plants decient in hexokinase 1 but does not play a key role in tuber carbohydrate metabolism. Plant Mol Biol 2002, 49:491-501. www.sciencedirect.com

57. Takahashi F, Sato-Nara K, Kobayashi K, Suzuki M, Suzuki H:  Sugar-induced adventitious roots in Arabidopsis seedlings. J Plant Res 2003, 116:83-91. Low-to-moderate concentrations of exogenous sucrose are shown to induce the formation of adventitious roots from the hypocotyls of darkgrown Arabidopsis seedlings. These ndings indicate that, directly or indirectly, sucrose levels can profoundly affect the development of entire organs. 58. Yang Z, Zhang L, Diao F, Huang M, Wu N: Sucrose regulates elongation of carrot somatic embryo radicles as a signal molecule. Plant Mol Biol 2004, 54:441-459. 59. Tsai C-H, Miller A, Spalding M, Rodermel S: Source strength regulates an early phase transition of tobacco shoot morphogenesis. Plant Physiol 1997, 115:907-914. 60. Lorenz S, Tintelnot S, Reski R, Decker EL: Cyclin D-knockout  uncouples developmental progression from sugar availability. Plant Mol Biol 2003, 53:227-236. Cyclin D was previously shown to function in the regulation of progression through the cell cycle. In this study, cyclin D knockout mutants of the moss Physcomitrella patens were found to be insensitive to the normal glucose-induced prolongation of the juvenile phase of development. These results suggest that, at least in this moss, cyclin D plays an important role in integrating metabolism and developmental progression. 61. Corbesier L, Lejeune P, Bernier G: The role of carbohydrates in the induction of owering in Arabidopsis thaliana: comparison between the wild type and a starchless mutant. Planta 1998, 206:131-137. rilleux C: C:N ratio increases in the 62. Corbesier L, Bernier G, Pe phloem sap during oral transition of the long-day plants Sinapis alba and Arabidopsis thaliana. Plant Cell Physiol 2002, 43:684-688. 63. Dielen V, Lecouvet V, Dupont S, Kinet J-M: In vitro control of oral transition in tomato (Lycopersicon esculentum Mill.), the model for autonomously owering plants, using the late owering uniora mutant. J Exp Bot 2001, 52:715-723. n M, Go mez-Mena C, Ruiz-Garc a L, Salinas J, 64. Rolda nez-Zapater JM: Sucrose availability on the aerial part Mart of the plant promotes morphogenesis and owering of Arabidopsis in the dark. Plant J 1999, 20:581-590. 65. Ohto M-a, Onai K, Furukawa Y, Aoki E, Araki T, Nakamura K: Effects of sugar on vegetative development and oral transition in Arabidopsis. Plant Physiol 2001, 127:252-261. 66. van Dijken AJH, Schluepmann H, Smeekens SCM: Arabidopsis  trehalose-6-phosphate synthase 1 is essential for normal vegetative growth and transition to owering. Plant Physiol 2004, 135:969-977. The authors generated a line of Arabidopsis that is homozygous for a mutation in the gene encoding trehalose-6-P synthase (TPS), and that carries a copy of the wildtype TPS1 gene under the control of an inducible promoter. Using this plant line, they demonstrate that TPS1 activity is required for owering. 67. Quirino BF, Noh Y-S, Himelblau E, Amasino RM: Molecular aspects of leaf senescence. Trends Plant Sci 2000, 5:278-282. 68. Stitt M, Krapp A: The interaction between elevated carbon dioxide and nitrogen nutrition: the physiological and molecular background. Plant Cell Environ 1999, 22:583-621. ` s M, Pourtau N: Spatial patterns and metabolic 69. Wingler A, Mare  regulation of photosynthetic parameters during leaf senescence. New Phytol 2004, 161:781-789. Showed that moderate (111 mM) concentrations of exogenous glucose stimulate the senescence of Arabidopsis, but only if nitrogen levels are low. These results indicate the important role played by sugar and nitrogen levels in controlling the timing of senescence. 70. Yoshida S, Ito M, Nishida I, Watanabe A: Identication of a novel gene HYS1/CPR5 that has a repressive role in the induction of leaf senescence and pathogen-defence responses in Arabidopsis thaliana. Plant J 2002, 29:427-437. 71. Dai N, Schaffer A, Petreikov M, Shahak Y, Giller Y, Ratner K, Levine A, Granot D: Overexpression of Arabidopsis hexokinase in tomato plants inhibits growth, reduces photosynthesis, and induces rapid senescence. Plant Cell 1999, 11:1253-1266. Current Opinion in Plant Biology 2005, 8:93102

102 Growth and development

72. Moore B, Zhou L, Rolland F, Hall Q, Cheng W-H, Liu Y-X,  Hwang I, Jones T, Sheen J: Role of the Arabidopsis glucose sensor HXK1 in nutrient, light, and hormonal signaling. Science 2003, 300:332-336. The authors of this manuscript report the best evidence to date that hexokinases can have dual functions in sugar response and metabolism in plants. s M, Purdy S, Quentin N, Rue l A, Wingler A: 73. Pourtau N, Mare Interactions of abscisic acid and sugar signalling in the regulation of leaf senescence. Planta 2004, 219:765-772. 74. Balibrea Lara ME, Gonzalez Garcia M-C, Fatima T, Ehne R,  Lee TK, Proels R, Tanner W, Roitsch T: Extracellular invertase is an essential component of cytokinin-mediated delay of senescence. Plant Cell 2004, 16:1276-1287. Cytokinins were previously known to delay senescence. This report shows that an extracellular invertase is required for the cytokininmediated delay of senescence, as cytokinin no longer inhibits senescence in the presence of an inhibitor of this invertase. This nding represents signicant progress in understanding the mechanism through which cytokinins and sugars help to regulate senescence. 75. Koch KE: Carbohydrate-modulated gene expression in plants. Annu Rev Plant Physiol Plant Mol Biol 1996, 47:509-540. 76. Rolland F, Moore B, Sheen J: Sugar sensing and signaling in plants. Plant Cell 2002, 14:S185-S205. 77. Lloyd JC, Zakhleniuk OV: Responses of primary and secondary  metabolism to sugar accumulation revealed by microarray expression analysis of the Arabidopsis mutant, pho3. J Exp Bot 2004, 55:1221-1230. This study is interesting because it characterizes the effects of alterations in endogenous sugar concentrations, rather than the effects of supplying plants with exogenous sugars, on sugar-regulated gene expression. The pho3 mutant has elevated glucose, sucrose and fructose levels due to a mutation in a sucrose transporter. Affymetrix GeneChips carrying 22 500 probe sets were used to characterize gene expression in rosette leaves of adult mutant and wildtype plants. Small changes were found in the expression of many genes that are involved in primary carbon metabolism. 78. Thum KE, Shin MJ, Palenchar PM, Kouranov A, Coruzzi GM:  Genome-wide investigation of light and carbon signaling interactions in Arabidopsis. Genome Biol 2004, 5:R10. Affymetrix GeneChips carrying information on approximately 8000 Arabidopsis genes were used to examine gene expression in whole tissues of approximately 16-day-old seedlings grown on 0% or 1% sucrose under dark or white-light conditions. The authors determined that genes that are involved in metabolism are over-represented among sugar- and lightregulated genes. 79. Price J, Laxmi A, St Martin SK, Jang J-C: Global transcription  proling reveals multiple sugar signal transduction mechanisms in Arabidopsis. Plant Cell 2004, 16:2128-2150. Affymetrix GeneChips carrying approximately 22 500 probe sets were used to examine the effects of different glucose and nitrogen combinations on gene expression in whole tissues of 6-day-old Arabidopsis seedlings. The authors also examined the effects of cycloheximide on glucose- and nitrogen-regulated gene expression. They determined that cycloheximide affects the expression of glucose-induced genes more frequently than that of glucose-repressed genes, suggesting that de novo protein synthesis is more commonly required for glucose induction than for repression. 80. Wang R, Okamoto M, Xing X, Crawford NM: Microarray analysis of the nitrate response in Arabidopsis roots and shoots reveals over 1,000 rapidly responding genes and new linkages to glucose, trehalose-6-phosphate, iron, and sulfate metabolism. Plant Physiol 2003, 132:556-567. 81. Finkelstein RR, Gampala SSL, Rock CD: Abscisic acid signaling in seeds and seedlings. Plant Cell 2002, 14:S15-S45. 82. Nambara E, Suzuki M, Abrams S, McCarty DR, Kamiya Y, McCourt P: A screen for genes that function in abscisic acid signaling in Arabidopsis thaliana. Genetics 2002, 161:1247-1255. 83. Huijser C, Kortstee A, Pego J, Weisbeek P, Wisman E, Smeekens S: The Arabidopsis SUCROSE UNCOUPLED-6 gene is identical to ABSCISIC ACID INSENSITIVE-4: involvement of abscisic acid in sugar responses. Plant J 2000, 23:577-586.

84. Rook F, Corke F, Card R, Munz G, Smith C, Bevan MW: Impaired sucrose-induction mutants reveal the modulation of sugarinduced starch biosynthetic gene expression by abscisic acid signaling. Plant J 2001, 26:421-433. 85. Brocard IM, Lynch TJ, Finkelstein RR: Regulation and role of the Arabidopsis abscisic acid-insensitive 5 gene in abscisic acid, sugar, stress response. Plant Physiol 2002, 129:1533-1543. 86. Hooks MA, Turner JE, Murphy EC, Graham IA: Acetate nonutilizing mutants of Arabidopsis: evidence that organic acids inuence carbohydrate perception in germinating seedlings. Mol Genet Genomics 2004, 271:249-256. 87. Cheng W-H, Endo A, Zhou L, Penney J, Chen H-C, Arroyo A, Leon P, Nambara E, Asami T, Seo M et al.: A unique short-chain dehydrogenase/reductase in Arabidopsis glucose signaling and abscisic acid biosynthesis and functions. Plant Cell 2002, 14:2723-2743. 88. Morita-Yamamuro C, Tsutsui T, Tanaka A, Yamaguchi J: Knock-out of the plastid ribosomal protein S21 causes impaired photosynthesis and sugar-response during germination and seedling development in Arabidopsis thaliana. Plant Cell Physiol 2004, 45:781-788. 89. Kang J, Turano FJ: The putative glutamate receptor 1.1 (AtGLR1.1) functions as a regulator of carbon and nitrogen metabolism in Arabidopsis thaliana. Proc Natl Acad Sci USA 2003, 100:6872-6877. 90. Mita S, Hirano H, Nakamura K: Negative regulation in the expression of a sugar-inducible gene in Arabidopsis thaliana. A recessive mutation causing enhanced expression of a gene for b-amylase. Plant Physiol 1997, 114:575-582. 91. Baier M, Hemmann G, Holman R, Corke F, Card R, Smith C, Rook F, Bevan MW: Characterization of mutants in Arabidopsis showing increased sugar-specic gene expression, growth, and developmental responses. Plant Physiol 2004, 134:81-91. 92. Mita S, Murano N, Akaike M, Nakamura K: Mutants of Arabidopsis thaliana with pleiotropic effects on the expression of the gene for b-amylase and on the accumulation of anthocyanin that are inducible by sugars. Plant J 1997, 11:841-851. 93. Villalobos MA, Bartels D, Iturriaga G: Stress tolerance and glucose insensitive phenotypes in Arabidopsis overexpressing the CpMYB10 transcription factor gene. Plant Physiol 2004, 135:309-324. lma n Z, Koncz C: Control of cell 94. Salchert K, Bhalerao R, Koncz-Ka elongation and stress responses by steroid hormones and carbon catabolic repression in plants. Philos Trans R Soc Lond B Biol Sci 1998, 353:1517-1520. kre L, O sz L, Szabados L, 95. Bhalerao RP, Salchert K, Bako Muranaka T, Machida Y, Schell J, Koncz C: Regulatory interaction of PRL1 WD protein with Arabidopsis SNF1-like protein kinases. Proc Natl Acad Sci USA 1999, 96:5322-5327. 96. Laby RJ, Kim D, Gibson SI: The ram1 mutant of Arabidopsis exhibits severely decreased b-amylase activity. Plant Physiol 2001, 127:1798-1807. 97. Chen J-G, Jones AM: AtRGS1 function in Arabidopsis thaliana. Methods Enzymol 2004, 389:338-350. 98. Martin T, Hellmann H, Schmidt R, Willmitzer L, Frommer WB: Identication of mutants in metabolically regulated gene expression. Plant J 1997, 11:53-62. 99. Purcell PC, Smith AM, Halford NG: Antisense expression of a sucrose non-fermenting-1-related protein kinase sequence in potato results in decreased expression of sucrose synthase in tubers and loss of sucrose-inducibility of sucrose synthase transcripts in leaves. Plant J 1998, 14:195-202. 100. Dijkwel PP, Huijser C, Weisbeek PJ, Chua N-H, Smeekens SCM: Sucrose control of phytochrome A signaling in Arabidopsis. Plant Cell 1997, 9:583-595.

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