Kock Susy Invertases

Sucrose metabolism: regulatory mechanisms and pivotal rolesin sugar sensing and plant developmentKaren Koch

Sucrose cleavage is vital to multicellular plants, not only for the

allocation of crucial carbon resources but also for the initiation of

hexose-based sugar signals in importing structures. Only the

invertase and reversible sucrose synthase reactions catalyze

known paths of sucrose breakdown in vivo. The regulation of

these reactions and its consequences has therefore become

a central issue in plant carbon metabolism. Primary mechanisms

for this regulation involve the capacity of invertases to alter

sugar signals by producing glucose rather than UDPglucose,

and thus also two-fold more hexoses than are produced by

sucrose synthase. In addition, vacuolar sites of cleavage

by invertases could allow temporal control via

compartmentalization. In addition, members of the gene families

encoding either invertases or sucrose synthases respond at

transcriptional and posttranscriptional levels to diverse

environmental signals, including endogenous changes that

reflect their own action (e.g. hexoses and hexose-responsive

hormone systems such as abscisic acid [ABA] signaling). At the

enzyme level, sucrose synthases can be regulated by rapid

changes in sub-cellular localization, phosphorylation, and

carefully modulated protein turnover. In addition to

transcriptional control, invertase action can also be regulated

at the enzyme level by highly localized inhibitor proteins and

by a system that has the potential to initiate and terminate

invertase activity in vacuoles. The extent, path, and site of

sucrose metabolism are thus highly responsive to both

internal and external environmental signals and can, in turn,

dramatically alter development and stress acclimation.

AddressesPlant Molecular and Cellular Biology Program, Horticultural Sciences

Department, University of Florida, Gainesville, Florida 32611, USA

e-mail: [email protected]

Current Opinion in Plant Biology 2004, 7:235246

This review comes from a themed issue on

Physiology and metabolismEdited by Christoph Benning and Mark Stitt

1369-5266/$ see front matter

2004 Elsevier Ltd. All rights reserved.

DOI 10.1016/j.pbi.2004.03.014

AbbreviationsABA abscisic acidCIN cytoplasmic invertaseCWIN cell wall invertaseENOD early nodulationPPi inorganic pyrophosphatePPV precursor protease vesicleSnRK SNF-related kinaseSUS sucrose synthase

UDP uridine di-phosphateUDPG UDPglucoseVIN vacuolar invertaseVPEc vacuolar processing enzyme g

IntroductionThe only known enzymatic paths of sucrose cleavage inplants are catalyzed by invertases (sucrose H2O !glucose fructose) and sucrose synthases (sucrose UDP ! fructose UDPglucose) (Figure 1). Both ofthese paths typically degrade sucrose in vivo but theproducts of their reactions differ in important ways[1,2]. Invertases produce glucose instead of UDPglucose(UDPG), and thus also form twice as many hexoses.Either of these features could give invertases a greatercapacity to stimulate specific sugar sensors [39]. Theresulting signals can alter expression of diverse genes[3,10,11], so invertases can potentially be strong effectorsof widely varying processes. These include the biosynth-esis and perception of hormones such as abscisic acid(ABA). In addition, both sugar and hormone signals canalso affect the expression of the genes that encode sucrosesynthase and invertase [13,5,7,12,13,14,15,16,17]. Thisreview explores the hypothesis that sucrose metabolismlies at the heart of a sensitive, self-regulatory develop-mental system in plants. Its influence appears to bebalanced by the capacity for sensing sucrose itself,

Figure 1

Current Opinion in Plant Biology

InvertaseGlucoseFructose

Hexosesignals

?Fructose

Hexose signals

Sucrose signals

Sucrose synthaseUDP glucose

Sucrose

The sucrose-cleaving enzymes are pivotal to maintaining the balance

between both sugar signals and metabolic paths. Different sensing

mechanisms are activated by hexoses and by other downstream

metabolites, and also by sucrose itself. The invertase path of

sucrose cleavage generates free glucose and twice as much overallsubstrate for hexose-based sugar sensing as does the degradative

action of the reversible sucrose synthase reaction (i.e. two hexoses

as opposed to fructose UDPG). In addition, genes that encodesucrose-cleaving enzymes are responsive to the action of their own

enzyme products. They respond to sugar signals in addition to

affecting the generation of these signals.

www.sciencedirect.com Current Opinion in Plant Biology 2004, 7:235246

but different systems and responses are involved[18,19,20,21,22].

Additional factors in the link between sucrose metabo-lism and sugar signals lie in the physical path of sucroseimport and sites of sucrose cleavage (Figure 2). Sucrosecan move from phloem into the cytoplasm of sink cellswith or without crossing the plasmamembrane or the cellwall space. This is an important distinction becausepoints of membrane interface have been implicated inspecific mechanisms of both sucrose and hexose sensing[1,4,8,22,23]. The plasma membrane can be exposed toabundant sucrose and/or hexoses if plasmodesmatal con-nections between cells are absent [3,4] (or functionallylimited [24]). Cell wall invertase (CWIN) can markedlyincrease localized hexose levels in these instances, whichtypify developing seeds and grains [6,25,26] in whichthere is no symplastic (plasmodesmatal) continuitybetween maternal and seed tissues. Pathogens and otherspecific stimuli can also induce cell wall invertases, evenwhen plasmodesmatal paths are intact [5,17,27], and

these instances can favor sucrose import and amplificationof hexose signals if sucrose moves out into the extracel-lular space.

In contrast, plasmodesmatal continuity remains intactand provides the predominant transfer path for sucrosethroughout most maternal sinks (i.e. roots, shoots, fruitflesh and so on) ([35]; Figure 2). In these tissues, sucrosecan be imported with minimal effect on known signalingmechanisms [3,4]. Once inside the importing cells,sucrose metabolism again becomes important to sugarsensing, this time to endogenous hexose- and possiblysucrose-signaling systems [4,5,10,11,22,28]. The pro-ducts of sucrose synthase (SUS) action initiate the fewesthexose-based signals [3,6] (which is advantageous undermany conditions [8,17,2931]), and cytoplasmic inver-tases (CINs) are minimally active in most systems [2].However, cytoplasmic sucrose is frequently transportedinto vacuoles for cleavage.

Vacuolar invertases (VINs), like their cell wall counter-parts, generate abundant hexoses and hexose-based sugarsignals [5,9,27,32]. The VINs also mediate the primarypath of sucrose cleavage in expanding tissues [1,2], andthus contribute to considerable hexose flux across thetonoplast and to the entry of hexoses into cytoplasmicmetabolism. Temporal control of both processes is furtherfacilitated by vacuolar compartmentalization, whichcould integrate the import and signaling functions ofVIN with its osmotic role in expansion.

Sucrose-cleaving enzymes can alter plantdevelopment through sugar signalsIn addition to their production of metabolic substrates,each site and path of sucrose cleavage described abovecan initiate a distinctive profile of sugar signals, whichin turn can have profound developmental effects(Figure 3). In general, hexoses favor cell division andexpansion, whereas sucrose favors differentiation andmaturation [6,26,33,34]. These effects, together withinformation from analyses of numerous systems, has ledto an invertase/sucrose-synthase control hypothesis fortransitions through the prominent stages of develop-ment [6,26,33,34]. According to this hypothesis, inver-tases mediate the initiation and expansion of many newsink structures [5,9,35], often with vacuolar activity pre-ceding that in cell walls [17,36]. The action of cell wallinvertase coincides with the elevated expression of hex-ose transporters in some systems [23]. Later transition tostorage and maturation phases is facilitated by changes inthe hexose/sucrose ratio (i.e. the cellular sugar state[6,26,33,34]), and by shifts from invertase to sucrose-synthase paths of sucrose cleavage. In some highly loca-lized regions, elevated levels of cell wall invertase persistduring maturation [17,37]. Sucrose synthase, too, can beactive in localized sites during the early phases of devel-opment (e.g. in the phloem and at sites of rapid cell wall

Figure 2


Vacuole

G + F UDPG + F G + F

Sink cell cytoplasm

Sucrose

Phloem

Sucrose

Cell wallspace

Plasmo-desmata

G + F

CIN SUS VINCWIN

The physical path of sucrose movement and site of its cleavage are

central not only to mechanisms of import but also to the sugar signals

generated. The mechanisms that regulate sucrose movement and

cleavage also differ for each compartment. Depending on the path of

sucrose entry into an importing structure (i.e. via plasmodesmata or

across the cell wall space), sucrose can be cleaved by cell wall

invertase (CWIN), cytoplasmic invertase (CIN), sucrose synthase (SUS),

or vacuolar invertase (VIN). Cytoplasmic sucrose cleavage typically

produces limited hexoses or hexose-based sugar signals because

of the generally low activity of CIN and the production of UDPG instead

of glucose by SUS. In contrast, sucrose that enters sinks via the cell

wall space can generate significant amounts of glucose and otherhexoses if CWIN is active. Glucose and fructose can, in turn, initiate

hexose-based signals both at the membrane and during subsequent

metabolism in the cytoplasm. Similarly, abundant hexoses and

hexose-based signals can be generated in the vacuole by VIN; this

constitutes the primary site of sucrose cleavage during the expansion

phase of most sink tissues. F, fructose; G, glucose.

236 Physiology and metabolism

Current Opinion in Plant Biology 2004, 7:235246 www.sciencedirect.com

deposition [17,3840]). Overall, however, diverse evi-dence supports the contention that the balance betweeninvertase and sucrose synthase activity can alter plantdevelopment through differential effects on sugar signal-ing systems.

Invertase-derived hexose signals can also markedly alterthe expression of genes for both the biosynthesis andsensing of specific hormones (e.g. ABA). Genes for bothprocesses can be regulated at transcriptional and post-transcriptional levels by hexose signals [11,41,42]. Insome instances, effects on one hormone system mayindirectly influence others (e.g. antagonisms betweenABA and ethylene or auxin [11,4143]). Nonetheless,the influence of sugar-sensing systems extends fromcytokinins [11,42] to gibberellins [42,44], and from auxin[11] to ethylene [10,4143,45]. Some thorough and strik-ing studies of the sugarABA interface have been carriedout recently. Here, hexose-based signals (originating fromsucrose cleavage) are implicated in regulation of ABAbiosynthetic genes [45], in the early steps of ABA sensing[46], and in influencing downstream elements in thissignal transduction system [46,47].

The influence of sucrose cleavage products on hormonesystems combined with the regulation of sucrose meta-bolism by those hormone systems together comprise ameans of integrating individual cellular responses intothose of the multicellular organism.

Roles and regulation of sucrose synthasesEssential roles of sucrose synthases

Recent developments indicate that the role of sucrosesynthase in sucrose import may involve a dual capacity todirect carbon toward both polysaccharide biosynthesisand an adenylate-conserving path of respiration. A keyfunction of sucrose synthase in biosynthetic processes issupported by evidence of its contribution to cell wallformation, which is inhibited in maize sucrose synthaseknock-outs (K Koch et al., unpublished), maize mutants[2,48], anti-sense carrot plants [5], and anti-sense cottonseeds [40]. The work on cotton [40] showed that high-cellulose fibers do not form if sucrose synthase activity hasbeen significantly decreased; and where antisense effectsextend from epidermis to the seed interior, embryo andendosperm development are also inhibited. Additionalanalysis of mutant and antisense plants that had reduc-tions in sucrose synthase (compiled in [9]) also showedthat these plants had markedly less starch in storageorgans (e.g. carrot roots and potato tubers). The UDPGproduct of sucrose synthase has been implicated in theformation of the starch [49] and in the synthesis of callose[39,50] and diverse cell wall polysaccharides [51,52].

The centrality of sucrose synthase to sucrose import bymany structures may relate to the typically moderate, butwidespread, reductions of oxygen levels within certain

Figure 3


Rel

ative

co

ntri

butio

nto

suc

rose

met

abol

ism

Sink initiation and expansionMaximal generation of vacuolar hexoses- Expansion (linked to photosynthate availability)- Respiration- Hexose-based sugar sensing (cell division and initial differentiation)

Vacuolar invertases:

Cell wall invertases:Continued sink initiation and expansionMaximal generation of cell wall hexoses- Reduction of turgor-based transport inhibition, especially as volume increases to near maximum- Turgor reduction in zones of phloem unloading (localized action often in much of development)- Respiration- Hexose-based sugar sensing (cell division and related gene expression)

Sucrose synthases:Storage and maturationGeneration of UDPG instead of glucose- Direct shuttling of UDPG to biosynthesis- ATP-conserving respiratory path- Minimization of hexose-based sugar sensing (enhanced expression of genes for storage and maturation)

Vacuolarinvertases

Cell wallinvertases

Sucrosesynthases

Developmental progression

A common developmental profile of the contributions of sucrose-cleaving enzymes to sequential stages of sink initiation, expansion, and

storage/maturation. Both vacuolar and cell wall invertases maximize

hexose production, which in turn enhances respiration and the

generation of hexose-based signals. These hexose-based signals

upregulate diverse early development genes, including those

involved in cell division. Vacuolar invertases concurrently favor cell

expansion through their two-for-one enhancement of osmotic

solutes in the vacuoles, as well as by linking the initial volume

increase to photosynthate availability. As cell expansion slows, the role

of cell wall invertase becomes increasingly important in preventing the

turgor-based inhibition of symplastic transfer, and also in reducing

turgor in the zones of phloem unloading. As the cell division and

expansion phases shift to those of storage and maturation, sucrose

synthases become more important, and contribute in at least three

overall areas. These are their capacity to direct carbon to an ATP-

conserving respiratory path that is advantageous in many sink

tissues, to facilitate the shuttling of UDPG to biosynthetic products

or other special functions, and to minimize the production of hexoses

and thus hexose-based signals during specific stages of sinkdevelopment.

Sucrose metabolism Koch 237


structures. Recent data indicate that endogenous oxygenlevels are generally reduced to varying degrees in activesinks such as potato tubers [53] and developing seeds[54], and also in the phloem [55]. Sucrose synthase canoperate effectively under these conditions [7], and caneven reduce the extent of oxygen depletion [53], butinvertases typically do not operate well under theseconditions [7]. The activity of invertases can also exacer-bate the problem of oxygen depletion [53]. Evidencefrom transgenic potatoes shows that the normal develop-mental shift to sucrose cleavage by sucrose synthase inwildtype tubers improves not only their adenylate bal-ance, starch biosynthesis, and respiratory costs, but alsotheir endogenous oxygen levels relative to those of tubersthat overexpress alternate sucrose breakdown enzymes(i.e. invertase or sucrose phosphorylase) [53]. Sucrosesynthase is one of only a few genes to be upregulatedunder low-oxygen conditions [8,30]; its essentiality underthese conditions is indicated by the capacity of wildtypeplants but not sucrose-synthase-deficient mutants to sur-vive flooding [56,57]. Under pronounced low-oxygenconditions, sucrose synthase responds rapidly to earlyrises in cytosolic Ca2 [31,50] and can support consider-able biosynthesis (of cellulose and callose) [50,51]. Thefunctional significance of sucrose synthase, as opposed toinvertase, is likely to be particularly important in low-oxygen conditions where it can aid the respiratory con-servation of adenylates through the production of UDPGinstead of additional hexoses (which would each requirean ATP for entry into glycolysis).

Sucrose synthases may also be central to the efficacy ofat least some symbioses (e.g. nitrogen-fixing nodules[58,59]) and have been implicated in the maintenanceof others (e.g. mycorrhizae). A rug4-a (rugosus meaningwrinkled) mutation that reduces sucrose synthase levelsin pea seeds and nodules inhibits nitrogen-fixation [58];conversely, if soybean nodules harbor symbionts that areunable to fix nitrogen, then sucrose synthase is notinduced [59]. Sucrose synthase and VIN are also inducedspecifically in root cells that have mycorrhizal arbuscules[60] (and sometimes also in adjacent cells [61]), and thisupregulation occurs early in development [62].

Further roles for sucrose synthase in development andsucrose import by diverse sinks have been implicated.The sucrose synthase gene is one of the first to showelevated expression as leaf primordia differentiate fromthe apical meristem [63]. The leaves and roots of carrotplants in which sucrose synthase has been reduced trans-genically are also ultimately smaller than those of wild-type plants [5], and similar tomato transgenics showedreduced fruit set from the earliest flowers [64]. A sucrosesynthase path of sucrose cleavage also typically predomi-nates over those involving invertases during the storageand maturation stages of organ development [65,66]. Atleast some of these developmental effects are likely to

stem from the capacity of sucrose synthase to minimizehexose-based sugar signals, particularly during periodswhen these signals could be detrimental to differentiationand/or maturation [6].

It is also worth noting that sucrose synthase may besignificant to human nutrition in a context not previouslyappreciated: the amino acid content of this protein,together with its abundance in mature grains, make itan important contributor to nutritionally limiting lysinelevels in maize kernels [67]. Approximately 75% of totalkernel lysine is contributed by sucrose synthase and twoother cytoskeleton proteins (UDPG starch glucosyl trans-ferase and fructose 1,6-bisphosphate aldolase).

Sucrose synthases role in phloem sieve elements

Recent work has immunolocalized sucrose synthase tosieve-tube elements as well as to companion cells [17],thus linking sucrose cleavage to sieve-tube functionmore directly than previously recognized (Figure 4).Earlier studies had localized sucrose synthase to adjacentcompanion cells [68], a site compatible with the results ofmetabolite analysis that indicated activity in or near thetransport path [69]. Later work also showed that inor-ganic pyrophosphate (PPi) is essential for phloem func-tion, and implicated a sucrose-synthase-based path ofrespiration, probably in companion cells [70]. However,recent results move sucrose synthase into close physicalproximity with the sieve-tube plasma membrane and aphloem-specific ATPase that is believed to aid sucrosetransport and compartmentalization [17]. This sieve-tube locale could also facilitate sucrose synthases prob-able role in directly supplying UDPG for the rapidwound-induced biosynthesis of callose plugs. Furtheradvantages of sucrose synthase in both sucrose transportand callose biosynthesis are its capacities to favor thecycling of PPi as opposed to ATP [29,55,69] and tofunction effectively under low oxygen conditions[30,56]. Both PPi cycling and low oxygen conditionsare typical of the phloem [55,69]. The actin and/ormembrane association of sucrose synthase [2,71,72,73]may also provide an important physical anchor for itsaction within the phloem transport stream.

Transcriptional control of sucrose synthases

Several advances have arisen from attempts to unravel thebasis of the differential expression of sucrose synthasegenes, particularly in response to oxygen and sugar avail-ability. Those sucrose synthase genes that are upreg-ulated by carbohydrate deprivation are also the moststrongly induced by low-oxygen, and show narrow pat-terns of expression that could confer sucrose import orsurvival priority for key tissues [3,8,30]. The starvation-inducible maize sucrose synthase (Shrunken1 [Sh1]) alsorespond rapidly to low O2, with marked increases in bothits mRNA levels and enzyme activity, especially undermodest oxygen depletion (i.e. 3% O2) [30]. Additional



work on sugar-inducible sucrose synthases in potato hasindicated that this upregulation requires SNF-relatedkinase (SnRK), a SNF1 ortholog, because the responsewas abolished at the mRNA level in SnRK-antisenseplants [7476]. Although data from analogs that cannotbe metabolized suggest little or no hexokinase involve-ment in the induction of rice Rsus1 [77], separate roles forboth hexokinase-dependent and -independent sugar-sensing mechanisms are implicated in the responses ofan Arabidopsis Sus1 [78].

One of the strongest known plant enhancers of geneexpression is the first intron of the Sh1 maize sucrosesynthase, which is particularly effective when introducedinto the 50 region of a gene or construct. Recently, a 145-bp portion of the 1028 bp Sh1 intron was found to inducegene expression by 2050-fold. Most of this induction wasmediated by the T-richness of a 35-bp motif rather thanits specific sequence [79].

Translational control of sucrose synthase

A key contributor to sustaining elevated levels of sucrosesynthase mRNA and protein under low oxygen appears tobe the preferential loading of sucrose synthase mRNAsonto large polysomes [80], where their translation is en-hanced by upregulation of both initiation and elongation[80]. Sucrose synthase activity increases rapidly but tovarying degrees under low oxygen [30], but a still-greatercapacity for activity increase after stress release is indicatedby the high levels of accumulated mRNA. Such disparitiesbetween regulation at transcriptional and posttranscrip-tional levels may have specific physiological significance.

Regulation of sucrose synthase by sub-cellular

localization

Recent developments in determining how the membranelocalization of sucrose synthase is controlled are presagedby earlier work on the enzymes apparent association withrosettes of the plasma membrane cellulose synthase com-plex (Figure 5). Such a relationship could allow UDPGfrom sucrose synthase to feed directly into cellulose for-mation, and essential UDP to be recycled readily [81].Evidence indicates that the involvement of sucrosesynthase in this rapidly changing structure [38,52] mightalso extend to the analogous formation of mixed-linkagepoly-glycans [82], and to the production of callose near thephragmoplast or in localized exoplasmic zones [39,52,83].

Recent work indicates a still-broader role for the transientassociation of sucrose synthase with membranes, andfurther, that the control of this localization might

Figure 4


Sucrose synthaseSieve-tube element

Companion cell

Openlumen

Cellwall

UDP

UDPG Fructose

UTP

G1P

UGPase

Callosesynthase

PPiPPi

F6P

Sieve-tube

lumen

Sieve-tube cytoplasm

Biosynthesis Respiration

Sucrose

Sucrose synthase

H+

Glycolysis

?

?

Sucrose

ATP

H+

ADP

ATPfor transporter

Callosefor plugging

(a)

(b)

Sucrose metabolism in phloem sieve-tube elements. (a) Sucrosesynthase localization extends to the sieve-tube element, where it

visually co-localizes with phloem-specific membrane ATPases [17]and lies in close proximity to sucrose transporters and sites of rapid

wound-induced callose formation. The sieve-tube lumen is open to

solute flow, but detectable levels of sucrose synthase do not move

within it [69]. The capacity of sucrose synthase to bind actin [72] and

membranes [2,52,73,84,85,90], and/or to localize in specific regionsof the cytoplasm [39] is compatible with its minimal mobility and highly

localized presence on or near the sieve-tube plasmamembrane [17].(b) A model for the regulation of sucrose use within sieve tubesintegrates recent information on the cellular and sub-cellular localization

of this sugar with the results of analyses of metabolic changes in the

phloem sap [66,70] and internal environment [55]. Two of the greatestdemands for sucrose use are likely to be callose biosynthesis, which is

massive during defensive plugging, and ATP production for the

maintenance of steep sucrose gradients across the plasma

membrane. Both processes could be readily supported by products of

the sucrose synthase reaction with minimal input from local

mitochondria, especially if PPi levels favor the activity of UDPG

pyrophosphorylase (UGPase). The resulting uridine tri-phosphate(UTP) can contribute either directly or indirectly to glycolysis and ATP

formation. The alternative demands made by callose biosynthesis

in sieve tubes could be closely associated with sucrose synthase

through both PPi cycling (from callose synthase back to UGPase)

and with the proximal localization of sucrose synthase, UGPase, and

callose synthase [39]. F6P, fructose-6-phosphate; G1P, glucose-1-

phosphate.



influence the balance between sucrose synthases role inrespiration and special functions in biosynthesis or com-partmentalization (Figure 5c). The enzyme is typicallysoluble in the cytoplasm and can contribute readily to anATP-conserving path of respiration (Figure 4). However,it can also move rapidly on and off membrane or cyto-skeletal locations, indicating that it might also have anumber of special functions. These include activities atplasma membrane and Golgi sites for cell wall biosynth-esis [39,51,8183], a possible role at the tonoplast that isrelated to the use and/or storage of vacuolar sucrose [84],and an activity at points on actin that could facilitatestarch formation through plastid proximity and/or othermetabolic shuttling via metabalons [2,71,72].

Sugars and other signals can affect the localization ofsucrose synthase, providing a potential means of fine-tuning the balance between respiration and biosynthesis[2,71,85]. Although sucrose synthase has a non-specificaffinity for membranes [85], early work indicated that thephosphorylation status of the enzyme is involved inregulating this association [57,72]. This is further sup-ported by the sensitivity of candidate kinases to cellularsignals [49,75,8689]. The extent of sucrose synthasephosphorylation in different fractions has varied withstudies and/or systems, however, such that the free cyto-plasmic enzyme can be hypo- [72,73], hyper- [72,73], orequally phosphorylated [89] relative to membrane- oractin-bound forms. Emerging data indicate that develop-mental status can influence this relationship [85], possi-bly because of shifts in membrane type or composition.

Regulation of sucrose synthase protein turnover

Despite the extreme stability of sucrose synthase pro-tein under some conditions [77], it is becoming apparentthat several mechanisms contribute to tightly control itsturnover (Figure 6). First, the phosphorylation that acti-vates the enzyme (at the S15 site or its equivalent) [86,87]is also implicated in predisposing sucrose synthase tophosphorylation at a second site (S170 or its equivalent)[90]. This second phosphorylation, in turn, targets theprotein for ubiquitin-mediated degradation via the pro-teosome [90]. Phosphorylation at the first site can belinked to sugar availability and/or to other signals throughthe catalysis of this step by both Ca2-dependant proteinkinases (CDPKs) [87,90,91] and/or a sugar-responsiveCa2-independent SnRK [75,88,89,90]. The secondphosphorylation can be catalyzed by CDPKs but notSnRKs [90]. Second, this avenue of sucrose synthasebreakdown can be inhibited if the second phosphoryla-tion site (S170 or equivalent) is blocked by the binding ofENOD40 proteins (which were initially named for theirrole in early nodule development but which are nowknown to have diverse functions throughout the plant)[90,92,93]. The protective association of ENOD40swith sucrose synthase has been implicated in the controlof vascular function, phloem loading/unloading, and

Figure 5


(c)

(a)

Sucrose synthase

Cellulose synthase complex

Cellu

lose

Sucrosesynthase

Cellulosesynthase

Cellulos

e

Fructose

UDP

UDPGUDPG

UDP

Cell wall

Poresubunit

Plasma membrane

Sucrose

Sucrose

(b)

Plasmamembrane

Cell wall

Cytoplasm

Cytoplasm

Cell wallbiosynthesis

SUS

CytoplasmicMembrane-bound

(Respiration)

(Special functions)

Vacuole

Plasma membrane

Golgi

Amyloplasts

Actin

Tonoplast

Regulation of sucrose synthase through sub-cellular localization. (a) Arapidly growing body of evidence supports an association between

sucrose synthase and rosettes of the plasma membrane cellulose

synthase complex, much as originally proposed by Delmer and

coworkers [82]. (b) The biochemical features of this model facilitate thedirect transfer of UDPG from the sucrose synthase reaction into

cellulose biosynthesis, and the rapid recycling of UDP. (c) Recentwork has indicated a broad spectrum of apparently transient

associations of sucrose synthase with membranes and/or actin. The

mechanisms that regulate this change in locale have become a

central issue, because they could provide a means of controlling the

balance between the cytoplasmic respiratory functions of sucrose

synthase and special roles of this enzyme in other processes. Among

the latter processes are cell wall biosynthesis (in the plasmamembrane and Golgi), starch biosynthesis (in the plastids), metabolic

channeling via metabalons (an actin-related process), and possibly

compartmentalization and/or mobilization (involving the tonoplast or the

sieve-tube plasma membrane). Cellular sugar status, developmental

stage, and phosphorylation appear to influence the shift of sucrose

synthase from cytoplasmic to membrane-bound forms.



assimilate import [90,92,94]. ENOD40 mRNAs areelevated, for example, at sites of high sink activity andat points of rapid unloading in phloem.

Roles and regulation of invertasesRoles of vacuolar invertases

It is now becoming evident that vacuolar invertases con-tribute prominently to both sucrose import ([5,17,36];P Commuri et al., unpublished data) and sugar signals[9,27,32], particularly during the expansion growth ofdiverse sink structures [1,5,9,17,26,36,95]. These rolesfor vacuolar invertases are additional to previously recog-nized functions including turgor regulation and the con-trol of sugar balance in fruit tissues and mature tubers[5,9,96]. The sucrose import and potential signalinginfluences of vacuolar invertases arise from their role in

the primary path of cleavage for sucrose entering growing/expanding sink tissues. The transfer of sucrose tovacuoles for initial metabolism has a dual advantage in,first, giving a two-for-one return on the cost of transport-ing osmotically active solutes into the vacuole for expan-sion and, second, adjusting organ expansion relative tocarbohydrate supply. Such contributions by vacuolarinvertase depend, however, on continued capacity forcell wall expansion. If turgor exceeds this potential, thensucrose cleavage by these invertases could have a minimalor even a negative effect on sucrose import [24]. Duringsink initiation and the initial expansion growth of manysinks, however, vacuolar invertases appear to have a keyrole [5,9,35,36,95].

Vacuolar invertase expression is also sensitive to an arrayof signals, including sugars, hormones, and environmentalstimuli [3,13,14,17,96]. Hence, these enzymes are in aposition to modulate responses to diverse inputs. Amongthese are the influence of gravity and indole acetic acid(IAA) on bending stems [13], cytokinins and IAA on theformation and expansion of tumors [17], drought andABA on hexose levels in leaves [14], and cold on thesweetening of tubers [96]. The apparent contribution ofvacuolar invertase in the adjustment of reproductive loadunder stress and resource limitation, when the earlyabortion of some fruits or seeds can allow the survivalof others, is also significant [36].

Roles of cell wall invertases

Cell wall invertases are central to phloem unloading insome, but not all, sucrose-importing structures. Theirsignificance is most prominent in sinks in which anapoplastic (cell wall) step is involved because of a gapin plasmodesmatal connections between cells [9,25]. Thisoccurs in developing seeds and pollen, where an unload-ing role for cell wall invertases is consistent with resultsfrom Vicia faba [6], barley [26], and maize [37]. Stress toovaries during early development can also induce abor-tion, which is preceded by rapid changes in first vacuolarthen cell wall invertases ([36]; P Commuri et al., pers.comm.; J Mullin, pers. comm.). Cell wall invertase con-tributes predominantly to the development of pollen(another apoplastically isolated sink structure), and loca-lized antisense reductions in cell wall invertases can beused to manipulate male fertility [16]. Cell wall invertasescan also influence sinks in which plasmodesmatal con-nections remain intact, if at least some sucrose movesacross the cell wall space. There is evidence in support ofsuch a role for cell wall invertases in developing carrotroots [5], potato tubers [48], and in response to signalsfrom some biotic [17,27] and abiotic stresses [5].

The highly responsive transcriptional regulation of

invertases

Invertase genes respond to diverse signals, includingthose generated by direct and indirect effects of their

Figure 6


S15 S170

SUS

S15 S170

SUS

Stable SUS

S15 S170

SUS

ENODproteinsP P

Unstable SUS

Ubiquination anddegradation by proteosomes

S15 S170

SUS

P P

S15 S170P P

SUS

P

+ P by CDPKs

Activatedby +P

Current model for SUS regulation through protein turnover. (a) Stableforms of SUS include those activated by phosphorylation at an S15

site (or equivalent) [49,73,86,87], especially if a second phosphorylationsite is blocked by the binding of ENOD proteins [90]. (b) If the protectiveENOD protein is absent, however, the initial phosphorylation event

predisposes SUS to a degradative process that begins with a second

phosphorylation at the S170 site (or its equivalent). This targets SUS

for ubiquitin-mediated degradation by the proteosome [90]. Theprotective association of ENOD40s with sucrose synthase has been

implicated in control of vascular function, phloem loading/unloading,

and assimilate import [92,93,94]. P, phosphate; S, serine.



own expression [3,15,16]. Invertases are also repressedby low oxygen, strongly and rapidly enough to serve aspotential markers of endogenous oxygen availability[7]. In addition, invertases respond to a full spectrumof plant hormones, and to a wide range of environmentaland pathogenic signals [13,14,15,16]. Different familymembers can also show contrasting responses to sugars orplant hormones [3,14,36], and the expression of thesame gene can differ markedly with tissue and/or condi-tions. The maize Ivr2 vacuolar invertase gene, for exam-ple, is upregulated in the leaves of drought-stressedplants [14] but downregulated in ovaries and youngkernels under similar conditions [36]. The effects ofstress signals, including ABA ([14]; K Koch, unpub-lished), are modified by other developmental signals.Collectively, these signals facilitate the contributions ofinvertase to the survival and acclimation of leaves, yetrestrict reproductive investment to a limited number ofoffspring that will have a greater chance of support tomaturity.

Regulation of targeting and turnover for vacuolar

invertase

The regulation of vacuolar invertase at the protein levelinvolves a previously unrecognized means of controllingboth compartmentalization and breakdown, the precursorprotease vesicle (PPV) and vacuolar processing enzyme(VPEg) system ([97]; Figure 6). Analysis of an Arabidop-sis vacuolar invertase (AtFRUCT4 [At1g12240]) showsthat this protein can be compartmentalized for extendedperiods in the PPVs, which are distinctive spindle-shapedendomembrane vesicles that lie between ribosomes and

vacuoles [97]. This storage of invertases can apparentlydelay delivery to vacuoles, thus imposing an additionallevel of control beyond that of mRNA-, protein-, or total-enzyme activity levels. In addition, these PPV compart-ments house an inactive form of VPEg protease(At4g32940) that can be released into vacuoles togetherwith the invertase, and that can subsequently target theinvertase for degradation [97]. The VPEg auto-activatesupon entry into the acidic vacuole, and includes at leastone vacuolar invertase among its specific substrates. ThePPV compartmentalization of a vacuolar invertase couldthus regulate both the time at which its activity invacuoles begins and its vulnerability to subsequent turn-over by the VPEg protease (Figure 7).

Regulation of invertases by inhibitor proteins

Inhibitor proteins are gaining increasing recognition as apotentially important means of in-vivo control for inver-tase activity. The presence of these relatively smallprotein inhibitors has been well documented in varioussystems, and they inhibit both vacuolar and cell wallinvertases when present in extracts. However, the sitesof action and functional significance of these proteins invivo have not been fully defined. Although most inhibitorproteins appear to be cell wall proteins, a putative vacuo-lar homolog of a cell wall form from tobacco has beenidentified and introduced into transgenic potatoes [96].The tubers of these plants had reduced capacity for thecold-induced sweetening that typically results from hex-ose accumulation in vacuoles. A probable effect withinthe vacuole was indicated for this transgenic system. Inaddition, the recent work of Bate et al. [98] demonstrated

Figure 7


Ribosomes

PPV

VINtranscription

VIN transfer and extendedprotective storage

VIN turnover by the VPEcysteine protease system

Vacuole

VPEauto-activated

VINdegraded

VINactiveVIN

active?

VPEinactive

Regulation of vacuolar invertase (VIN) at the protein level. The transfer, protective retention, and protein turnover of VIN are potentially modulated bythe precursor protease vesicle (PPV) and vacuolar processing enzyme (VPEg) system. At least some of the newly translated VIN enters the PPV,where it can be retained for extended periods. It is presumably protected from proteolytic degradation in this compartment, but its functional

role remains unclear until it enters the vacuole. The PPV also stores a VPEg protease, which remains inactive until it too moves into the vacuole.Once activated, VPEg targets specific substrates that include a VIN. This invertase may thus be targeted for degradation as soon as it enters thesite of its presumed action in vivo. The PPV and VPEg system therefore provides a potential mechanism for modifying both the timing and theduration of at least some VIN activity.



that a maize invertase inhibitor, ZmINVINH1, is export-ed to the apoplast, where it could interact with invertasesduring early kernel development (i.e. 47 days afterpollination). These researchers further defined the mostprominent site of ZmINVINH1 expression as the embryosurrounding region (ESR), which they suggest may helpto preserve crucial differences in the extracellular sugarenvironments of the embryo and endosperm during earlydevelopment. Greater hexose levels in the endospermapoplast may favor more rapid cell division and minimaldifferentiation relative to those in embryos [6].

ConclusionsSucrose-cleaving enzymes lie at the heart of mechanismsfor the distribution and use of sucrose within multicellularplants. They also occupy a pivotal position in the balancebetween the different sugar signals generated byimported sucrose. Their regulation has thus becomethe focus of considerable interest, and involves diverseand highly integrated mechanisms operating at transcrip-tional and posttranscriptional levels.

AcknowledgementsSupported by the US National Science Foundation (CellularBiochemistry) and by the Florida Agricultural Experiment Station(Journal Series Number R-10079).

References and recommended readingPapers of particular interest, published within the annual period ofreview, have been highlighted as:

of special interestof outstanding interest1. Tymowska-Lalanne Z, Kreis M: The plant invertases: physiology,

biochemistry, and molecular biology. Adv Bot Res 1998,28:71-117.

2. Winter H, Huber SC: Regulation of sucrose metabolism in higherplants: localization and regulation of activity of key enzymes.Crit Rev Biochem Mol Biol 2000, 35:253-289.

3. Koch K: Carbohydrate-modulated gene expression in plants.Ann Rev Plant Physiol Plant Mol Biol 1996, 47:509-540.

4. Lalonde S, Boles E, Hellmann H, Barker L, Patrick JW,Frommer WB, Ward JM: The dual function of sugar carriers:transport and sugar sensing. Plant Cell 1999, 11:707-726.

5. Sturm A, Tang G-Q: The sucrose-cleaving enzymes of plants arecrucial for development, growth and carbon partitioning.Trends Plant Sci 1999, 4:401-407.

6. Wobus U, Weber H: Sugars as signal molecules in plant seeddevelopment. Biol Chem 1999, 390:937-944.

7. Zeng Y, Wu Y, Avigne WT, Koch KE: Rapid repression of maizeinvertases by low oxygen. Invertase/sucrose synthasebalance, sugar signaling potential, and seedling survival.Plant Physiol 1999, 121:599-608.

8. Koch KE, Ying Z, Wu Y, Avigne WT: Multiple paths of sugar-sensing and a sugar/oxygen overlap for genes of sucrose andethanol metabolism. J Exp Bot 2000, 51:417-427.

9. Koch KE, Zeng Y: Molecular approaches to altered Cpartitioning: genes for sucrose metabolism. J Amer Soc Hort Sci2002, 127:474-483.

10. Smeekens S: Sugar-induced signal transduction in plants.Ann Rev Plant Physiol Plant Mol Biol 2000, 51:49-81.

11. Rolland F, Moore B, Sheen J: Sugar sensing and signaling inplants. Plant Cell 2002, 14:S185-S205.

12. Kim J, Jun SH, Kang HG, Lee J, An G: Molecular characterizationof a GA-inducible gene, Cvsus1, in developing watermelonseeds. Mol Cell 2002, 14:255-260.

13. Long JC, Shao W, Rashotte AM, Muday GK, Huber SC:Gravity-stimulated changes in auxin and invertase geneexpression in maize pulvinal cells. Plant Physiol 2002,128:591-602.

14.

Trouverie J, Thenenol C, Rocher J-P, Sotta B, Prioul J-L:The role of abscisic acid in the response of a specific vacuolarinvertase to water stress in the adult maize leaf. J Exp Bot 2003,54:2177-2188.

This study provides an interesting example of how the ABA-mediatedresponses of a vacuolar invertase differ between leaves and reproductivestructures (compare with [36]). The different responses of this enzymecould provide a means of maintaining growth and photosynthetic inputfrom turgid leaves while concurrently adjusting reproductive load.

15. Roitsch T, Ehne SR, Goetz M, Hause B, Hofmann M, Sinha AK:Regulation and function of extracellular invertase from higherplants in relation to assimilate partitioning, stress responsesand sugar signalling. Aust J Plant Physiol 2000, 27:815-825.

16. Roitsch T, Balibrea ME, Hofmann M, Proels R, Sinha AK:Extracellular invertase: key metabolic enzyme and PR protein.J Exp Bot 2003, 54:513-524.

17.

Wachter R, Langhans M, Aloni R, Gotz S, Weilmunster A, Koops A,Temguia L, Mistrik I, Pavlovkin J, Rascher U et al.: Vascularization,high-volume solution flow, and localized roles for enzymes ofsucrose metabolism during tumorigenesis by Agrobacteriumtumefaciens. Plant Physiol 2003, 133:1024-1037.

This detailed analysis of the development of the Agrobacterium tumor innormal and mutant plants provides a unique view of sink initiation andmaturation in a system mediated by a small and carefully defined set ofsignals. In addition, an investigation of vascular function both the tumorand in unaffected host phloem showed that the localization of sucrosesynthase extended to sieve tubes. There, sucrose synthase was found inclose physical proximity to a phloem ATPase that is believed to aid themaintenance of steep sucrose concentration gradients.

18. Loreti E, Alpi A, Perata P: Glucose and disaccharide-sensingmechanisms modulate the expression of alpha-amylase inbarley embryos. Plant Physiol 2000, 123:939-948.

19.

Tiessen A, Hendricks JH, Stitt M, Branscheid A, Gibon Y, Farre EM,Geigenberger P: Starch synthesis in potato tubers is regulatedby post-translational redox modification of ADP-glucosepyrophosphorylase: a novel regulatory mechanism linkingstarch synthesis to the sucrose supply. Plant Cell 2002,14:2191-2213.

A range of different approaches indicate that the degree of dimerizationcould be a source of ADP-glucose modulation in vivo, and that this ADP-glucose modulation could be affected by cellular sucrose status.

20.

Hajirezaei MR, Bornke F, Peisker M, Takahata Y, Lerchl J,Kirakosyan A, Sonnewald U: Decreased sucrose content triggersstarch breakdown and respiration in stored potato tubers(Solanum tuberosum). J Exp Bot 2003, 54:477-488.

A comparative analysis of transgenic potatoes in which sucrose levelswere altered by different means indicated that respiratory regulationcould respond to sucrose depletion directly, and that sugar effects arenot necessarily mediated by hexoses alone.

21. Tiessen A, Prescha K, Branscheid A, Palacios N, McKibbin R,Halford NG, Geigenberger P: Evidence that SNF1-related kinaseand hexokinase are involved in separate sugar-signallingpathways modulating post-translational redox activation ofADP-glucose pyrophosphorylase in potato tubers. Plant J 2003,35:490-500.

22.

Vaughn MW, Harrington GN, Bush DR: Sucrose-mediatedtranscriptional regulation of sucrose symporter activity inphloem. Proc Natl Acad Sci USA 2002, 99:10876-10880.

A sucrose transporter is found to be sucrose responsive at a transcrip-tional level, and also to be localized on the membranes of companioncells. Together, these features support a potential role for this sucrosetransporter as a central contributor to a system-wide mechanism forsensing and distributing sucrose within a plant.

23. Sherson SM, Afford HL, Forbes SM, Wallace G, Smith SM: Roles ofcell-wall invertases and monosaccharide transporters in thegrowth and development of Arabidopsis. J Exp Bot 2003,54:525-531.



24. Tauberger E, Hoffman-Benning S, Fleischer-Notter H, Willmitzer L,Fisahn J: Impact of invertase overexpression on cell size, starchgranule formation and cell wall properties during tuberdevelopment in potatoes with modified carbon allocationpatterns. J Exp Bot 1999, 50:477-486.

25. Patrick JW, Offler CE: Compartmentation of transport andtransfer events in developing seeds. J Exp Bot 2001,52:551-564.

26.

Weschke W, Panitz R, Gubatz S, Wang Q, Radchuk R, Weber H,Wobus U: The role of invertases and hexose transporters incontrolling sugar ratios in maternal and filial tissues ofbarley caryopses during early development. Plant J 2003,33:395-411.

Developmental changes in the expression and localization of invertasesand hexose transporters are appraised relative to the potential forhexoses produced by invertases to affect sugar sensing and cellularsugar status at different locales.

27. Herbers K, Sonnewald U: Altered gene expression broughtabout by inter- and intracellularly formed hexoses and itspossible implications for plantpathogen interactions.J Plant Res 1998, 111:323-328.

28. Moore B, Zhou L, Rolland F, Hall Q, Cheng WH, Liu YX, Hwang I,Jones T, Sheen J: Role of the Arabidopsis glucose sensor HXK1in nutrient, light, and hormonal signaling. Science 2003,300:332-336.

29. Huber SC, Akazawa T: A novel synthase pathway for sucrosedegradation in cultured sycamore cells. Plant Physiol 1986,81:1008-1013.

30. Zeng Y, Wu Y, Avigne WT, Koch KE: Differential regulation ofsugar-sensitive sucrose synthases by hypoxia and anoxiaindicate complementary transcriptional andposttranscriptional responses. Plant Physiol 1998,116:1573-1583.

31. Subbaiah CC, Sachs MM: Molecular and cellular adaptations ofmaize to flooding stress. Ann Botany 2003, 91:119-127.

32. Sonnewald U, Brauer M, von Schaewen A, Stitt M, Willmitzer L:Transgenic tobacco plants expressing yeast-derived invertasein either the cytosol, vacuole or apoplast: a powerful tool forstudying sucrose metabolism and sink/source interactions.Plant J 1991, 1:95-106.

33. Borisjuk L, Rolletschek H, Wobus U, Weber H: Differentiationof legume cotyledons as related to metabolic gradientsand assimilate transport into seeds. J Exp Bot 2003,54:503-512.

34. Borisjuk L, Walenta S, Rolletschek H, Mueller-Klieser W, Wobus U,Weber H: Spatial analysis of plant metabolism: sucrose imagingwithin Vicia faba cotyledons reveals specific developmentalpatterns. Plant J 2002, 29:521-530.

The authors provide dramatic visual evidence of the association ofsucrose gradients across developing cotyledons and a shift from celldivision to starch storage within the cells of these organs. The data areconsistent with the authors suggestion that sucrose gradient has acausal role in this shift.

35. Sonnewald U, Hajirezaei MR, Kossmann J, Heyer A, Trethewey RN,Willmitzer L: Increased potato tuber size resulting fromapoplastic expression of a yeast invertase. Nature Biotechnol1997, 15:794-797.

36.

Andersen MN, Asch F, Wu Y, Jensen CR, Naested H,Mogensen VO, Koch KE: Soluble invertase expression is an earlytarget of drought stress during the critical, abortion-sensitivephase of young ovary development in maize. Plant Physiol 2002,130:591-604.

The expression and contributions of sucrose-cleaving enzymes wereappraised during early ovary development in field-grown plants. Vacuolarinvertases predominated during the earliest phases of developmentbefore the upregulation of other genes that are involved in sucrosemetabolism. During this early period, drought rapidly repressed vacuolarinvertases and aborted many kernels.

37. Cheng WH, Endo A, Zhou L, Penney J, Chen HC, Arroyo A, Leon P,Nambara E, Asami T, Seo M, Koshiba T, Sheen J: A unique short-chain dehydrogenase/reductase in Arabidopsis glucosesignaling and abscisic acid biosynthesis and functions.Plant Cell 2002, 14:2723-2743.

38. Gardiner JC, Taylor NG, Turner SR: Control of cellulose synthasecomplex localization in developing xylem. Plant Cell 2003,15:1740-1748.

39. Salnikov VV, Grimson MJ, Seagull RW, Haigler CH: Localization ofsucrose synthase and callose in freeze-substituted secondary-wall-stage cotton fibers. Protoplasma 2003, 221:175-184.

40.

Ruan YL, Llewellyn DJ, Furbank RT: Suppression of sucrosesynthase gene expression represses cotton fiber cell initiation,elongation and seed development. Plant Cell 2003, 15:952-964.

An interesting gradation of phenotypes indicates that antisense reductionsof sucrose synthase at the seed surface inhibit the development oftrichomes, and that internal reductions have still more pronounced effects.The data support an association between sucrose synthase and cell-wallbiosynthesis, as well as other developmental roles for sucrose synthase.

41. Yanaglsawa S, Yoo S-D, Sheen J: Differential regulation of EIN3stability by glucose in ethylene signaling in plants. Nature 2003,425:521-525.

42. Gibson SI: Sugar and phytohormone response pathways:navigating a signaling network. J Exp Bot 2004, 55:253-264.

43. Leon P, Sheen J: Sugar and hormone connections. Trends PlantSci 2003, 8:110-116.

44. Lu C-A, Ho T-HD, Ho S-L, Yu S-M: Three novel MYB proteins withone DNA binding repeat mediate sugar and hormone regulationof a-amylase gene expression. Plant Cell 2002, 13:1963-1980.

45. Gazzarrini S, McCourt P: Genetic interactions between ABA,ethylene and sugar signaling pathways. Curr Opin Plant Biol2001, 4:387-391.

46. Brocard-Gifford IM, Lynch TJ, Finkelstein RR: Regulatorynetworks in seeds integrating developmental, abscisic acid,sugar, and light signaling. Plant Physiol 2003, 131:78-92.

47. Niu X, Helentjaris T, Bate NJ: Maize ABI4 binds couplingelement1 in abscisic acid and sugar response genes.Plant Cell 2002, 14:2565-2575.

48. Cheng WH, Tallercio EW, Chourey PS: The miniature1 seed locusof maize encodes a cell wall invertase required for normaldevelopment of endosperm and maternal cells in the pedicel.Plant Cell 1996, 8:971-983.

49. Asano T, Kunieda N, Omura Y, Ibe H, Kawasaki T, Takano M,Sato M, Furuhashi H, Mujin T, Takaiwa F et al.: Rice SPK, acalmodulin-like domain protein kinase, is required for storageproduct accumulation during seed development:phosphorylation of sucrose synthase is a possible factor.Plant Cell 2002, 14:619-628.

50. Subbaiah CC, Sachs MM: Altered patterns of sucrose synthasephosphorylation and localization precede callose inductionand root tip death in anoxic maize seedlings. Plant Physiol 2001,125:585-594.

51. Albrecht G, Mustroph A: Localization of sucrose synthase inwheat roots: increased in situ activity of sucrose synthasecorrelates with cell wall thickening by cellulose depositionunder hypoxia. Planta 2003, 217:252-260.

52. Doblin MS, Kurek I, Jacob-Wilk D, Delmer DP: Cellulosebiosynthesis in plants: from genes to rosettes. Plant Cell Physiol2002, 43:1407-1420.

53.

Bologa KL, Fernie AR, Leisse A, Loureiro ME, Geigenberger P:A bypass of sucrose synthase leads to low internal oxygen andimpaired metabolic performance in growing potato tubers.Plant Physiol 2003, 132:2058-2072.

The authors provide evidence of altered metabolic states in developingsinks, and of a central role for sucrose synthase in minimizing respiratorycosts and oxygen use.

54.

Rolletschek H, Borisjuk L, Koschorreck M, Wobus U, Weber H:Legume embryos develop in a hypoxic environment.J Exp Bot 2002, 53:1099-1107.

The results of this study indicate that considerably less than ambientoxygen concentrations are typical of the environment inside developingseeds.

55.

van Dongen JT, Schurr U, Pfister M, Geigenberger P: Phloemmetabolism and function have to cope with low internaloxygen. Plant Physiol 2003, 131:1529-1543.



Direct probe analyzes indicate that low oxygen is common in the phloemenvironment and has direct implications for phloem function.

56. Ricard B, Toai VT, Chourey P, Saglio P: Evidence for the criticalrole of sucrose synthase for anoxic tolerance of maize rootsusing a double mutant. Plant Physiol 1998, 116:1323-1331.

57. Fukao T, Kennedy RA, Yamasue Y, Rumpho ME: Genetic andbiochemical analysis of anaerobically induced enzymes duringseed germination of Echinochloa crus-galli varieties tolerantand intolerant of anoxia. J Exp Bot 2003, 54:1421-1429.

58. Gordon AJ, Minchin FR, James CL, Komina O: Sucrose synthasein legume nodules is essential for nitrogen fixation.Plant Physiol 1999, 120:867-878.

59. Xie ZP, Staehelin C, Broughton WJ, Wiemken A, Boller T, Muller J:Accumulation of soluble carbohydrates, trehalase and sucrosesynthase in effective (Fix(R)) and ineffective (Fix(S)) nodulesof soybean cultivars that differentially nodulate withBradyrhizobium japonicum. Funct Plant Biol 2003, 30:965-971.

60. Blee KA, Anderson AJ: Transcripts for genes encoding solubleacid invertase and sucrose synthase accumulate in root tipand cortical cells containing mycorrhizal arbuscules. Plant MolBiol 2002, 50:197-211.

61. Hohnjec N, Perlick AM, Puhler A, Kuster H: The Medicagotruncatula sucrose synthase gene MtSucS1 is activated both inthe infected region of root nodules and in the cortex of rootscolonized by arbuscular mycorrhizal fungi. Mol Plant MicrobeInteract 2003, 16:903-915.

62. Ravnskov S, Wu Y, Graham JH: Arbuscular mycorrhizalfungi differentially affect expression of genes coding forsucrose synthases in maize roots. New Phytol 2003,157:539-545.

63. Pien S, Wyrzykowska J, Fleming AJ: Novel marker genes for earlyleaf development indicate spatial regulation of carbohydratemetabolism within the apical meristem. Plant J 2001,25:663-674.

64. DAoust M-A, Yelle S, Nguyen-Quoc B: Antisense inhibition oftomato fruit sucrose synthase decreases fruit setting and thesucrose unloading capacity of young fruit. Plant Cell 1999,11:2407-2418.

65. King SP, Lunn JE, Furbank RT: Carbohydrate content andenzyme metabolism in developing canola siliques.Plant Physiol 1997, 114:153-160.

66. Fernie AR, Willmitzer L, Trethewey RN: Sucrose to starch: atransition in molecular plant physiology. Trends Plant Sci 2002,7:35-41.

67. Azama K, Abe S, Sugimoto H, Davies E: Lysine-containingproteins in maize endosperm: a major contribution fromcytoskeleton-associated carbohydrate-metabolizingenzymes. Planta 2003, 217:628-638.

68. Nolte KD, Koch KE: Companion-cell specific localization ofsucrose synthase in zones of phloem loading and unloading.Plant Physiol 1993, 101:899-905.

69. Geigenberger P, Langenberger S, Wilke I, Heineke D, Helt HW,Stitt M: Sucrose is metabolized by sucrose synthase andglycolysis within the phloem complex of Ricinus communisL. seedlings. Planta 1993, 190:446-453.

70. Lerchl J, Geigenberger P, Stitt M, Sonnewald U: Impairedphotoassimilate partitioning caused by phloem-specificremoval of pyrophosphate can be complemented by aphloem-specific cytosolic yeast-derived invertase intransgenic plants. Plant Cell 1995, 7:259-270.

71. Winter H, Huber JL, Huber SC: Identification of sucrose synthaseas an actin-binding protein. FEBS Lett 1998, 430:205-208.

72. Winter H, Huber JL, Huber SC: Membrane association of sucrosesynthase: changes during the graviresponse and possiblecontrol by protein phosphorylation. FEBS Lett 1997,420:151-155.

73.

Komina O, Zhou Y, Sarath G, Chollet R: In vivo and in vitrophosphorylation of membrane and soluble forms of soybeannodule sucrose synthase. Plant Physiol 2002, 129:1664-1673.

Soybean nodule sucrose synthases are shown to be tightly associatedwith membranes. Data indicate that this localization is closely controlledby phosphorylation.

74. Etxeberria E, Gonzalez P: Evidence for a tonoplast-associatedform of sucrose synthase and its potential involvement insucrose mobilization from the vacuole. J Exp Botany 2003,54:1407-1414.

75. Purcell PC, Smith AM, Halford NG: Antisense expression of asucrose non-fermenting-1-related protein kinase sequence inpotato results in decreased expression of sucrose synthase intubers and loss of sucrose-inducibility of sucrose synthasetranscripts in leaves. Plant J 1998, 14:195-202.

76. Halford NG, Hey S, Jhurreea D, Laurie S, McKibbin RS, Paul M,Zhang Y: Metabolic signalling and carbon partitioning: roleof Snf1-related (SnRK1) protein kinase. J Exp Bot 2003,54:467-475.

77. Halford NG, Hey S, Jhurreea D, Laurie S, McKibbin RS, Zhang Y,Paul MJ: Highly conserved protein kinases involved in theregulation of carbon and amino acid metabolism. J Exp Bot2004, 55:35-42.

78. Liao YC, Wang AY: Sugar-modulated gene expression ofsucrose synthase in suspension-cultured cells of rice.Physiol Plant 2003, 118:319-327.

79. Ciereszko I, Kleczkowski LA: Glucose and mannose regulate theexpression of a major sucrose synthase gene in Arabidopsis viahexokinase-dependent mechanisms. Plant Physiol Biochem2002, 40:907-911.

80. Clancy M, Hannah LC: Splicing of the maize Sh1 first intron isessential for enhancement of gene expression, and a T-richmotif increases expression without affecting splicing.Plant Physiol 2002, 130:918-929.

81. Fennoy SL, Nong T, Bailey-Serres J: Transcriptional andpost-transcriptional processes regulate gene expression inoxygen-deprived roots of maize. Plant J 1998, 15:727-735.

82. Amor Y, Haigler CH, Johnson S, Wainscott M, Delmer DP:A membrane-associated form of sucrose synthase and itspotential role in synthesis of cellulose and callose in plants.Proc Nat Acad Sci USA 1995, 92:9353-9357.

83. Buckeridge MS, Vergara CE, Carpita NC: Insight into multi-sitemechanisms of glycosyl transfer in (1!4)beta-D-glycansprovided by the cereal mixed-linkage (1!3),(1!4)beta-D-glucan synthase. Phytochemistry 2001, 57:1045-1053.

84. Hong Z, Zhang Z, Olson JM, Verma DP: A novel UDP-glucosetransferase is part of the callose synthase complex andinteracts with phragmoplastin at the forming cell plate.Plant Cell 2001, 13:769-779.

85.

Hardin SC, Winter H, Huber SC: Phosphorylation of theamino-terminus of maize sucrose synthase in relation tomembrane association and enzyme activity. Plant Physiol 2004,in press.

The authors present several lines of evidence showing that both thesucrose synthase phosphorylation state and its role in membrane asso-ciation are controlled developmentally.

86. Huber SC, Huber JL, Laio PC, Gage DA, McMichael RWJr,Chourey PS, Hannah LC, Koch KE: Phosphorylation of serine-15of maize leaf sucrose synthase. Plant Physiol 1996, 112:793-802.

87. Zhang XQ, Lund AA, Sarath G, Cerny RL, Roberts DM, Chollet R:Soybean nodule sucrose synthase (nodulin-100): furtheranalysis of its phosphorylation using recombinant andauthentic root-nodule enzymes. Arch Biochem Biophys 1999,371:70-82.

88. Chikano H, Ogawa M, Ikeda Y, Koizumi N, Kusano T, Sano H: Twonovel genes encoding SNF-1 related protein kinases fromArabidopsis thaliana: differential accumulation of AtSR1 andAtSR2 transcripts in response to cytokinins and sugars, andphosphorylation of sucrose synthase by AtSR2. Mol GenGenet 2001, 264:674-681.

89. Haigler CH, Ivanova-Datcheva M, Hogan PS, Salnikov VV,Hwang S, Martin K, Delmer DP: Carbon partitioning tocellulose synthesis. Plant Mol Biol 2001, 47:29-51.



90.

Hardin SC, Tang G-Q, Scholz A, Holtgraewe D, Winter H, Huber SC:Phosphorylation of sucrose synthase at serine 170: occurrenceand possible role as a signal for proteolysis. Plant J 2003,35:588-603.

The authors describe a previously unrecognized phosphorylation site ofsucrose synthase and its potential role in controlling the turnover of thesucrose synthase protein.

91. Cheng SH, Willmann MR, Chen HC, Sheen J: Calcium signalingthrough protein kinases. The Arabidopsis calcium-dependentprotein kinase gene family. Plant Physiol 2002, 129:469-485.

92. Kouchi H, Takane K, So RB, Ladha JK, Reddy PM: Rice ENOD40:isolation and expression analysis in rice and transgenicsoybean root nodules. Plant J 1999, 18:121-129.

93.

Rohrig H, Schmidt J, Miklashevichs E, Schell J, John M: SoybeanENOD40 encodes two peptides that bind to sucrose synthase.Proc Natl Acad Sci USA 2002, 99:1915-1920.

A combination of in-vitro translation and western blotting indicates thattwo small ENOD40 peptides are produced from overlapping regions atthe 50 end of the same mRNA. Further work demonstrates that thesepeptides bind specifically to sucrose synthase. This work, together withdata from other studies [90,94], implicates these ENOD40 peptides inenhancing the activity and longevity of sucrose synthase in diversesystems.

94.

Varkonyi-Gasic E, White DW: The white clover enod40 genefamily. Expression patterns of two types of genes indicate arole in vascular function. Plant Physiol 2002, 129:1107-1118.

In-situ localization of clover ENOD40 mRNAs showed them to be presentin vascular regions, especially at sites of extensive sucrose transfer.Together with data from [90], this work indicates a role for ENOD40 instabilizing functional sucrose synthase at these sites.

95. Klan EM, Hall B, Bennett AB: Antisense acid invertase (TINV1)gene alters soluble sugar composition and size in transgenictomato fruit. Plant Physiol 1996, 112:1321-1330.

96. Greiner S, Rausch T, Sonnewald U, Herbers K: Ectopic expressionof a tobacco invertase inhibitor homolog prevents cold-induced sweetening of potato tubers. Nat Biotechnol 1999,17:708-711.

97.

Rojo E, Zouhar J, Carter C, Kovaleva V, Raikhel NV: A uniquemechanism for protein processing and degradation inArabidopsis thaliana. Proc Natl Acad Sci USA 2003,100:7389-7394.

The authors describe a mechanism for the protective storage of vacuolarinvertase and a possible delay in its vacuolar action. A system for theinactivation of this enzyme after its release into the vacuole is alsodescribed (and involves proteolysis by a specific VPEg).

98.

Bate NJ, Niu X, Wang Y, Reimann KS, Helentjaris TG: An invertaseinhibitor from maize localizes to the embryo surroundingregion during early kernel development. Plant Physiol 2003,134:1-9.

Several lines of evidence indicate that a localized expression of a cell wallinvertase inhibitor from maize may have implications for early embryodevelopment.



Sucrose metabolism: regulatory mechanisms and pivotal roles in sugar sensing and plant developmentIntroductionSucrose-cleaving enzymes can alter plant development through sugar signalsRoles and regulation of sucrose synthasesEssential roles of sucrose synthasesSucrose synthase's role in phloem sieve elementsTranscriptional control of sucrose synthasesTranslational control of sucrose synthaseRegulation of sucrose synthase by sub-cellular localizationRegulation of sucrose synthase protein turnover

Roles and regulation of invertasesRoles of vacuolar invertasesRoles of cell wall invertasesThe highly responsive transcriptional regulation of invertasesRegulation of targeting and turnover for vacuolar invertaseRegulation of invertases by inhibitor proteins

ConclusionsAcknowledgementsReferences and recommended reading

Kock Susy Invertases

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