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TRENDS in Biochemical Sciences Vol.27 No.3 March 2002

http://tibs.trends.com 0968-0004/02/$ – see front matter © 2002 Elsevier Science Ltd. All rights reserved. PII: S0968-0004(01)02054-0

139ReviewReview

Conservation of amino acid transporters

in fungi, plants and animals

Daniel Wipf, Uwe Ludewig, Mechthild Tegeder, Doris Rentsch, Wolfgang Koch and

Wolf B. Frommer

In yeast, animals and plants, amino acids playfundamental roles in a multitude of processes,including protein synthesis, hormone metabolism,nerve transmission, cell growth, production ofmetabolic energy, nucleobase synthesis, nitrogenmetabolism and urea biosynthesis. In multicellularorganisms, many of the nitrogenous compounds aretransported between cells. Respective mediators; thatis, amino acid transporters, have been characterizedphysiologically in animals, plants and yeast. Thesestudies suggest the existence of multiple transporters,differing in substrate spectrum, transport mechanism(i.e. ions used in cotransport) and tissue specificity. Inanimal cells, >20 different transport systems weredefined on the basis of cotransport mechanism andsubstrate spectrum (see Box 1 for transportterminology). In recent years, much progress has beenmade in elucidating the molecular identity of theproteins underlying these physiological activities. Thefirst mammalian amino acid transporter wasidentified in human brain as a γ-aminobutyric acid(GABA) transporter [1]. The first plant transporterswere identified by suppressor cloning in yeast mutantsdefective in amino acid transport [2]. Subsequently,oocyte expression systems and suppression of yeastmutants [3] enabled identification of a wide spectrumof plasma membrane amino acid transporters fromdifferent eukaryotic kingdoms [4]. The knowntransporters fall into five superfamilies, and homologsof several have been found in many differentorganisms. In addition, several organellar familieshave been identified. Comparison of the transportersfrom three completely sequenced eukaryotic genomes– Saccharomyces cerevisiae, Arabidopsis thaliana andHomo sapiens – has enabled many physiologically

defined systems to be assigned to individual genes onthe basis of their transport properties (e.g. substratespectrum, transport mechanism and tissuespecificity). The genome-wide approach now allowsfunctional prediction for orphan transporters,generating a complete picture of amino acid transportphysiology at the molecular level.

Five superfamilies of amino acid transporters

Amino acid-polyamine-choline (APC)transporter superfamilyAmong eukaryotes, amino acid transport is bestunderstood in yeast [5]. As unicellular organisms,yeasts use a variety of transporters for nutritionaluptake of amino acids and for intracellularcompartmentalization. Uptake across the plasmamembrane is mediated by 24 different amino acidtransporters belonging to the APC superfamily(SLC7, pfam00324). All 24 members contain12 putative membrane-spanning domains and havebeen functionally characterized; for example, CAN1functions as a H+–Arg symporter [6]. APC familymembers are not highly specific but insteadtransport several related, or even a wide spectrum of,structurally different amino acids, includingD-isomers [7]. Interestingly, SSY1, which contains acharacteristic N-terminal extension, acts as anextracellular amino acid sensor and transducer,controlling amino acid uptake activity [8]. APChomologs are also found in animals and plants.Nevertheless, phylogenetic analyses show that APCmembers can be grouped into three clustersreflecting the three kingdoms (yeast, plant andanimal) (Fig. 1). APC homologs have also been foundin bacteria.

In the absence of structural data, computer-aidedpredictions of secondary structures (Fig. 1) were usedto further categorize APC transporters from animalsand plants into two subgroups. Members of thecationic amino acid transporters (CATs) have14 putative transmembrane domains and are foundin both animals and plants. Mammalian CATs arerepresentatives of system y+ (Table 1), mediatingNa+-independent uptake of cationic amino acids [9]and, similar to their plant homologs (AtCAT1),mammalian CATs are characterized by a high affinityfor cationic amino acids [10]. Because both plants andyeast maintain a proton gradient over the plasma

When comparing the transporters of three completely sequenced eukaryotic

genomes – Saccharomyces cerevisiae, Arabidopsis thaliana and Homo sapiens

– transporter types can be distinguished according to phylogeny, substrate

spectrum, transport mechanism and cell specificity. The known amino acid

transporters belong to five different superfamilies. Two preferentially

Na+-coupled transporter superfamilies are not represented in the yeast and

Arabidopsis genomes, whereas the other three groups, which often function as

H+-coupled systems, have members in all investigated genomes. Additional

superfamilies exist for organellar transport, including mitochondrial and

plastidic carriers. When used in combination with phylogenetic analyses,

functional comparison might aid our prediction of physiological functions for

related but uncharacterized open reading frames.

Daniel Wipf

Uwe Ludewig

Wolfgang Koch

Wolf B. Frommer*

ZMBP, Plant Physiology,Auf der Morgenstelle 1,Eberhard-Karls-Universität Tübingen,D-72076 Tübingen,Germany.*e-mail: [email protected]

Mechthild Tegeder

Washington StateUniversity, School ofBiological Sciences,Pullman, WA 99164-4236,USA.

Doris Rentsch

Molecular PlantPhysiology, Institute ofPlant Sciences,Altenbergrain 21, CH-3013 Bern,Switzerland.


http://tibs.trends.com

140 Review

membrane, it was not surprising thatAtCAT1-mediated accumulation of histidine was pHdependent [10]. However, it remains unclear whetherAtCAT1 functions as a uniporter or exchanger, or as asecondary active proton cotransporter.

The second subfamily, including the yeast APCtransporters, comprises proteins with 12 putativetransmembrane domains. Whereas the respectiveplant proteins AtLAT1–5 have not been analyzed,mammalian heteromeric amino acid transporters(HATs; for example, LAT-1, LAT-2, y+LAT1, y+LAT-2,asc-1, b0,+AT and two orphans) are composed of a lightsubunit and a heavy subunit [11,12]. The heavysubunit (SLC3) is required for the generation of activeplasma membrane amino acid transporters. Thissubunit is glycosylated, contains only one putativetransmembrane domain and plays a role in membranetargeting (e.g. in polarized epithelial cells) [11]. Twodifferent heavy subunits were identified in mammals:the widespread, strictly basolaterally localized (inepithelia) 4F2hc/CD98, and the brush border rBAT.So far, no similar proteins have been found in yeast orplants. Animals actively maintain Na+ andK+ gradients across the plasma membrane; thus,

amino acid accumulation is often coupled toNa+ gradients. Surprisingly, some of these Na+-coupledamino acid transporters fall into the HAT family [12],representing system y+L (y+LAT1, y+LAT2). The y+Lsystem is responsible for Na+-dependent transport ofneutral amino acids, whereas transport of basic aminoacids is not coupled to Na+ [13]. Taking theseobservations together, amino acid transport mediatedby members of the APC family is diverse, Na+- orH+-coupled, and has functions in uptake and nutrition.In multicellular organisms such as plants andmammals, these proteins might also function asexchangers important for both selective accumulationof specific amino acids, and redistribution andhomeostasis of the intracellular concentrations.

Sodium-dicarboxylate symporter (SDS) superfamilyIn animals, amino acids serve as nutrients or assignaling molecules; for example, duringneurotransmission. Fast removal of excitatory aminoacids from the synaptic cleft and recycling insurrounding neuroglia is crucial for terminatingpostsynaptic action and preventing accumulation ofexcitatory amino acids to neurotoxic levels (Fig. 2).

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Amino acid transporters have been characterized physiologicallyin animals, plants and yeast. Mulitple transporters exist, differingin substrate spectrum, transport mechanism and tissuespecificity. The text below defines the various types of transportand mammalian transport systems.

Mechanisms of transport

Active transport: often defined as transport up an electrochemicalgradient. More precisely defined as unidirectional or vectorialtransport produced within a membrane-bound protein complexby coupling energy-requiring processes to transport.Antiport (countertransport): coupled transport in whichmovement of ions (e.g. protons) drives active transport of asolute in the opposite direction.Exchanger: family of membrane proteins that perform theexchange of substrates across membranes.Primary active transport: the same membrane protein breaksdown ATP and uses energy released from this hydrolysis to movesolute across the membrane barrier against a chemical-potentialgradient.Secondary active transport: the movement of one species iscoupled to the movement of another species down anelectrochemical gradient established by primary activetransport.Symport (cotransport): membrane transport process thatcouples the transport of a substrate in one direction across amembrane (down its electrochemical gradient) to the transportof a different substrate in the same direction (up itselectrochemical gradient).Uniporters: a class of transmembrane transport proteins thattransports a single substrate across the plasma membrane alongits concentration gradient.

Amino acid transport systems of mammalian cells

System A: widespread, mediating transport of neutral amino acidswith broad selectivity including N-methyl derivatives, Na+-dependent.System asc: specific for small neutral amino acids.System ASC: ubiquitous, prefers neutral amino acids withoutbulky or side-branched chains, Na+-dependent.System B0,+: in blastocyts and probably also in brush-bordermembrane, broad specificity for zwitterionic and dibasic aminoacids, Na+-dependent.System b0,+: widespread, for dibasic and some neutral amino acids.System B0: brush border membrane of epithelia, broad substratespecificity, does not accept N-methyl amino acids, Na+-dependent.System BETA: widespread, transports β-alanine, taurine andGABA, Na+-dependent.System GLY: present in several tissues, transports glycine,Na+-dependent.System IMINO: in intestinal brush-border membrane, handlesproline, Na+-dependent.System L: widespread, for neutral-branched chains and aromaticamino acids.System N: cotransports glutamine and asparagine (and in someinstances histidine) with Na+ but, in contrast to system A,additionally countertransports protons.System PROT: proline-specific carriers, Na+-dependent.System T: aromatic amino acids.System VGT: vesicular glutamate transport.System X–

AG: brain and epithelial tissues, for acidic amino acids,

Na+-dependent.System X–

C: glutamate–cysteine exchanger.

System y+: widespread, for cationic and zwitterionic amino acids.System y+L: erythrocytes and placenta, transports basic aminoacids without Na+ but neutral amino acids with Na+.

Box 1. Transport terminology



141Review

SDS transporters mediate electrogenic glutamateand aspartate uptake by cotransport of Na+ andcounter-transport of K+, with no apparentdependence on Cl− gradients (GLAST [14], GLT-1 [15],EAAC1 [16]) (Fig. 2). Additionally, SDS membersshow non-stoichiometric Cl− channel activity gated bythe amino acid [17]. SDS members contain10 putative membrane-spanning domains withcytosolic C and N termini. Representatives of systemB0 [18], ASC (ASCT1; ASCT2) [19,20] and XAG

−, foundalso in non-brain tissues, also fall into the SDS family(SLC1; pfam00375) [16]. As rapid synaptictransmission and biochemically generated Na+

gradients are characteristic features of the animal

kingdom, it might not be surprising that no relatedproteins were found in yeast or plants, whereashomologs do exist in bacteria.

Neurotransmitter superfamily (NTS)Neurotransmitter transporters (Fig. 2) represent thethird large superfamily of transporters (SLC6;pfam00209). NTS members share a commonstructure with 12 putative transmembrane helicesand include plasma membrane carriers forγ-aminobutyric acid (GABA), proline, glycine andbetaine (e.g. GAT1, PROT), and probably representsystem B0,+ [21]. Amino acid transporters of the NTSdo not necessarily have a role in neurotransmissionbecause, for example, the GABA transporter (GAT-2),betaine/GABA transporter (BGT1), taurinetransporter (TAUT/BETA) and glycine transporter,are expressed in nonneuronal tissues. Becausecarriers belonging to this family are mainly found inbrain, and are probably involved in terminatingsynaptic transmission at inhibitory synapses(i.e. where large Na+ and Cl− gradients exist), NTStransporters couple amino acid uptake to electrogeniccotransport of Na+ and Cl−, thereby allowing cellularuptake of neurotransmitter molecules against aconcentration gradient. Similar to SDS, NTShomologs were not found in yeast and plant genomes.

Amino acid transporter superfamily 1 (ATF1)In contrast to other superfamilies, ATF1 members(SLC38; pfam01490) were first described in plants.Structurally related proteins were only recentlyidentified in yeast and animals [5]. The superfamilycontains plant-specific sub-branches, and branchesthat are more structurally related to yeast andhuman transporters (Fig. 3). ATF1 members contain9–11 putative membrane-spanning domains withcytosolic N and extracellular C termini [22] (Fig. 3).

The best-characterized members of the ATF1superfamily are ArabidopsisAAPs (amino acidpermeases) (Fig. 4) which, when expressed in yeast oroocytes, mediate Na+-independent, H+-coupleduptake of a wide spectrum of amino acids [23].AtAAP1–5 represent low affinity transporters withlow selectivity towards amino acid side chains; onlyAAP3 and 5 efficiently transport basic amino acids.AAP6 has a 10-fold higher affinity towards allsubstrates when compared with other AAPs and, sofar, is the only known transporter mediatingtransport of aspartate, which is involved in longdistance transport, with physiologically relevantaffinity [23]. This highlights the relevance ofbiophysical transport properties for physiology:although AAP3 and 5 prefer basic amino acids, in vivocompetition showed that at physiological glutamateor glutamine concentrations AAP3 and 5 will nottransport basic amino acids efficiently but insteadserve as general amino acid transporters. Thus, forexact assessment of in vivo function, imaging toolsneed to be devised to determine amino acid

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Fig. 1. Phylogenetic tree of the amino acid-polyamine-choline (APC) superfamily (SLC7). Maximumparsimony analyses were performed using PAUP 4.0b4a (http://paup.csit.fsu.edu/index.html), with allDNA characters unweighted and gaps scored as missing characters. Heuristic tree searches wereexecuted using 100 random sequence additions and the tree bisection-reconnection branch-swapping algorithm with random sequence analysis. The complete alignment was based on 894 sites;834 were phylogenetically informative. The APC superfamily can be divided into five different clusters(shadowed areas). The graphs at the bottom correspond to hydrophobicity plots obtained by TMHMManalysis [47]; the red curves correspond to predicted transmembrane domains, the blue tointracellular domains and the pink to extracellular domains. According to predicted membranetopologies, the genes can be divided into two clusters: (1) 12 transmembrane domains in plant andhuman LATs; and (2) 14 transmembrane domains in plant and human CATs and in yeast APCmembers. Abbreviations: At, Arabidopsis thaliana; Hs, Homo sapiens; Sc, Saccharomyces cerevisiae.



142 Review

concentrations at the sites of transport, and to correlate dynamic analysis of amino acidconcentration changes in subcellular compartmentswith transporter characteristics and localization.

Compared with yeast and animals, for whichcell-wall-free cell cultures that retain the properties oftissues are available, plant cells, with their rapiddedifferentiation, presence of cell walls and largevacuoles, are much more difficult to handle. Thus,much less is known about cell-specific amino acidtransport physiology in plants. Despite detailedknowledge about molecular properties of plant aminoacid transporters, direct characterization of cellularproperties as in animals is impossible. Only noveltools such as knockout mutants will allow light to beshed on the role of the numerous plant amino acidtransporters. Physiological studies using wholetissues, individual cells or plasma membrane vesiclesindicated that amino acid transport in plants ismediated by carriers with overlapping specificity,coupling an electrochemical potential gradient (of H+)to secondary active accumulation of amino acids[5,24]. Plants take up amino acids as a nitrogensource and, more importantly, amino acids are theprincipal, long distance transport forms of organicnitrogen. In accordance, plant amino acid transportergenes are preferentially expressed in vascular tissue(W.B. Frommer et al., unpublished). Althoughproteogenic amino acids accumulate in phloem(100–200 mM) and xylem sap (~10-fold less than inphloem), amines and acidic amino acids predominate.Intracellular, apoplasmic and phloem composition ofamino acids is similar; thus, the properties of AAPsare consistent with functions in cellular uptake ofamino acids from xylem and phloem loading (Fig. 4).The seven AAP genes characterized to date(AtAAP1–6 and AtAAP8) show distinct expressionpatterns, suggesting specific functions [5].

Promoter–reporter studies showed that AtAAP1 isexclusively expressed in seeds supplying embryos[25], whereas AtAAP2 was in vascular tissue,potentially functioning in exchange between xylemand phloem. Antisense experiments support the roleof AAPs in phloem transport (W. Koch et al.,unpublished). AtLHT1, a member of a second ATF1subfamily (Fig. 3) [26] transports lysine and histidine,but also other amino acids. LHT1 is present in alltissues, with expression being strongest in youngleaves, flowers and siliques. A third branch comprisesputative transporters (Fig. 3) for the plant hormoneauxin, which is structurally related to tryptophan[27]. ANT1, a member of the fourth branch (Fig. 3),mediates transport of neutral amino acids [28]. Incontrast to the plant carriers just described, whichtransport diverse amino acids, ProTs (prolinetransporters; Fig. 3) preferentially transport proline,betaine and GABA [29,30]. ProTs are ubiquitouslyexpressed but accumulate under salt stress, implyinga role in stress adaptation [29]. Proline-specificcarriers from humans were found as members of theNa+-coupled NTS superfamily (PROT [31]); however,the proline-specific, Na+-dependent system IMINOhas not yet been cloned. One can thus speculate thatthe transport system IMINO, characterized solely byfunctional studies, is encoded by animal ATF1homologs that remain to be characterized. Takentogether, all investigated plant ATF1s function asH+-coupled systems, probably playing a role inaccumulating amino acids within the plant cell.

Mammalian ATF1 homologs are most closelyrelated to plant genes that have not yet beencharacterized (Fig. 3); however, they also functionallyresemble plant transporters for cellular uptake ofamino acids. As might be expected, couplingmechanisms are different; that is, animal transporterscouple amino acid uptake to Na+ gradients, whereas

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Table 1. Amino acid transport systems of mammalian cellsa,b

HUGO Mammalian transport system Example gene Family designation GenBank identifier (gi)

SLC1 ASC ASCT1 SDS 1173365SLC1 B0 ATB0 SDS 1478281SLC1 X–

AG EAAT1 SDS 383572SLC6 B0,+ ATB0,+ NTS 5732680SLC6 BETA GAT1 NTS 204222SLC6 GLY GLYT1 NTS 204434SLC6 PROT PROT NTS 7657589SLC7 y+ CAT1 APC 1706186SLC7 + SLC3 asc HAT (asc1 + 4F2hc) APC + 4F2hc 8394322SLC7 + SLC3 b0,+ HAT (b0,+AT + rBAT) APC + rBAT 12585187SLC7 + SLC3 L HAT (LAT1 + 4F2hc) APC + 4F2hc 12643412SLC7 + SLC3 X–

C xCT APC + 4F2hc 12585386SLC7 + SLC3 y+L HAT (y+LAT1 + 4F2hc) APC + 4F2hc 4507055SLC16 T TAT1 Monocarboxylate 14090278SLC17 VGT BNP1 VGT 9945322SLC38 A ATA3 ATF1 13876616SLC38 N SN1 ATF1 5870893aSystems are classified in the SLC (solute carrier) series according to the Human Gene (HUGO) Nomenclature Committee Database (http://www.gene.ucl.ac.uk/nomenclature).bSubstrate: +, cationic; 0, neutral; – , anionic.



143Review

plant transporters couple amino acid uptake toH+ symport. Molecularly identified transporters such as SA1 represent system A [32], whereas theclosely related SN1 and SN2 correspond to system Nin brain (Fig. 2). Interestingly, system N cotransportsglutamine and asparagine (in some instanceshistidine) with Na+ but, in contrast to system A,additionally countertransports H+ ions [33]. In certain physiological conditions this might also lead to amino acid efflux. Interestingly, thisH+ antiport mechanism is conserved in anothersubgroup, the vesicular/vacuolar VGAT andAVT1 transporters.

Besides a role in plasma membrane amino acidtransport, some ATF1 members fulfil function in

import or export of solutes in intracellular vesiclesand vacuoles. In animals, amino acids areconcentrated in intracellular vesicles (in lysosomal orsynaptic vesicles) by proton antiport. For example,VGAT concentrates GABA in synaptic vesicles ofinhibitory synapses [34] (Fig. 2). Yeast VGAThomologs (AVT; Fig. 3) mediate amino acid transportinto and out of vacuoles [35]; compartmentation ofneutral amino acids into the vacuole is accomplishedby AVT1, an H+-antiporter, whereas amino acidexport is mediated by AVT3, 4 and 6, probablyfunctioning as H+ cotransporters. A structurally andfunctionally similar animal transporter wasidentified (LYAAT1; Fig. 3) that mediated H+-coupledexport of amino acids from lysosomes [36]. H+-coupledsymporters were described in epithelial cells; thus,one might speculate that these are also related toplant AAPs.

In summary, a multitude of transport mechanismsis realized in the ATF1 superfamily, making themattractive for comparative structure–functionanalyses. Bioinformatics suggests plastidiclocalization for some uncharacterized transporters(see ‘Organellar transporters’ section). ATF1members are involved in both uptake andredistribution of amino acids between organs andcellular compartments.

Amino acid transporters within the majorfacilitator superfamily (MFS)Amino acid transporters were found in two MFSsubfamilies and are related to H+–monocarboxylate

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144 Review

or phosphate transporters. MFS proteins transport awide spectrum of substances unrelated to amino acidsand use multiple transport mechanisms such asuniport or H+ cotransport.

MFS: H+–monocarboxylate transporters (MCTs)TAT1, a MCT member (SLC16) specifically mediatesuptake of aromatic amino acids, thus displayingfeatures of System T [37]. This finding might not besurprising as amino acids contain at least onecarboxylate group. As compared with othercharacterized members, TAT1 seems independent of Na+ and pH. MCTs are distantly related toorganic anion transporters (OATs), also belonging tothe MFS. Closer homologs not yet analyzed werealso found in yeast.

MFS: Vesicular glutamate transporters (VGTs)Surprisingly, VGTs [e.g. BNPI [38] (Fig. 2)] are closelyrelated to inorganic phosphate transporters (SLC17;pfam00083). Homologs are present in yeast(e.g. allantoate transporters; DAL5) and inArabidopsis, all with unknown function (Fig. 5). Itwill be interesting to test phosphate and glutamatetransport activities in all family members. Thus,cross-species comparison might aid our elucidation ofthe function of orphans.

MFS: Ionotrophic glutamate receptors (iGluRs)Amino acid derivatives serve as important signalingmolecules. Furthermore, cells determine theextracellular availability of substrates so as toregulate uptake and release systems accordingly.Thus, specific receptors or sensors are required at theplasma membrane. An emerging concept in yeast isthat members of the transporter families haveevolved as sensors (e.g. SSY1 [8]; see ‘APCsuperfamily’). A second well-characterized receptortype is represented by ionotrophic receptors: inanimal brains, ionotrophic glutamate receptors(iGluRs) function as glutamate-activated ionchannels in rapid synaptic transmission. A largefamily of putative iGluRs was also found in plants[39]. The existence of bacterial homologs indicatesthat signaling via iGluRs is a primitive signalingmechanism pre-dating the divergence of animals andplants. Phylogenetic analyses indicate that thedivergence of animal iGluRs and plant GLR genespre-dates the divergence of iGluR subtypes inanimals (N-methyl-D-aspartate versus kainate andα-amino-3-hydroxy-5-methylisoxazole-4-propionateas agonist) [39]. Although the involvement of plantiGluRs in amino acid signaling remains to beestablished, glutamate-dependent Ca2+ influx wasfound in wild-type plants [40], and in plants in which

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Fig. 3. Phylogenetic treeof the amino acidtransporter (ATF1)superfamily (SLC38).Phylogenetic analyseswere performed asdescribed for Fig. 1. Thecomplete alignment wasbased on 1038 sites;566 were phylogeneticallyinformative. The ATF1superfamily can bedivided into five differentclusters. The graphscorrespond tohydrophobicity plotsobtained by TMHMManalysis [47]; the redcurves correspond topredicted transmembranedomains, the blue tointracellular domains andthe pink to extracellulardomains. Green indicatesa plant gene, red ananimal gene and brown afungal gene. Nine or 11transmembrane domainsare predicted for all ATF1genes. Abbreviations:At, Arabidopsis thaliana;Hs, Homo sapiens;Sc, Saccharomycescerevisiae.



145Review

expression of ionotrophic glutamate homologs hadbeen knocked out (these plants show symptoms ofcalcium deficiency) [41]. However, with the typicallymillimolar concentrations of acidic amino acids foundin the phloem, compared with micro- orsubmicromolar concentrations in the synaptic cleftsof animals, affinities of the receptors and potentiallyalso signal transduction mechanisms must differ [36].

Organellar transporters

Mitochondrial transporter family (MCF)Mitochondria play an important role in amino acidmetabolism and, therefore, need multiple transportsystems in their membranes. Porins in the outermembrane allow passage of a wide spectrum ofcompounds, including amino acids. Inner membranecarriers comprising six-membrane-spanning domainsbelong to the mitochondrial carrier family(MCF; SLC25). In yeast, arginine and ornithinecarriers (arg11p and bac1p) were identified [42,43].MCF members potentially involved in amino acidtransport are also present in animals and plants.Suppression cloning in yeast mutants could serve as a tool for characterizing these homologs.

Plastidic transportersA major part of amino acid assimilation andbiosynthesis of plants occurs in plastids. Amino acidspass the outer envelope via OEP proteins (OEP16 and24), which form cation-selective high conductancechannels permeable to amines and amino acids [44,45].Amino acids cross the inner chloroplast envelope but,so far, transporters have not been identified. Recently,however, a member of the plasma membrane glucose

transporter family was identified as the chloroplastglucose transporter [46]. As the protein acquired atypical chloroplast-targeting sequence, one canspeculate that certain members of one of the five aminoacid transporter superfamilies described here mighthave acquired respective signal peptides targetingthem to the inner envelope. Bioinformatic analysesmight help us identify candidates for plastidic andmitochondrial transport systems.

Perspectives

Previously, a multitude of genes encoding amino acidtransporters was identified by functional expressioncloning. Together with the availability of the completegenome sequences of fungi, plant and humans, it ispossible to provide an almost complete insight intothe inventory of transporters of all three kingdoms. Incombination with phylogenetic analyses of amino acidtransporter superfamilies, full genome comparisonprovides evidence for the existence of a multitude oftransporters differing in localization, transportmechanisms and tissue distribution. As a minimum,yeast use 24, Arabidopsis 53 and humans 46, putativeamino acid transporters, not counting the largeorganellar carrier families. However, the largenumber of transporters mediating vesicular/vacuolaror plasma membrane amino acid transport is notunexpected considering the complex requirements forintercellular and long distance transport, and themultitude of physiological systems described inanimals (Table 1). The known amino acidtransporters fall into at least five differentsuperfamilies, which use diverse couplingmechanisms even within a family to determine thepolarity of transport. Two transporter superfamilies(SDS and NTS) were found exclusively in animals,whereas the other three groups are also representedin fungi and plants, potentially reflecting a differencein nutritional and signaling functions. Most plantstry to exclude Na+ to prevent toxicity, mainly usingH+ gradients generated by H+-ATPases as drivingforces for secondary active transport. By contrast,animals and humans only rarely use H+ gradients,instead coupling transport via Na+–K+ ATPases toNa+ gradients. The phylogenetic trees and structuralanalyses shown here, in combination with the widespectrum of analysis tools available for functionalcharacterization, will undoubtedly help us to assignfunctions to uncharacterized orphans. In contrast toother transporter superfamilies that consist ofproteins with similar topology (e.g. MFStransporters), the various amino acid transporterfamilies (e.g the ATF1 and APC superfamilies) areevidently divergent regarding topology. They alsodiffer with respect to coupling mechanisms andsubcellular distribution. However, the currentlylimited information about subfamily localization andtransport mechanisms does not yet allow for a clearcorrelation of the membrane topology of the orphansas revealed by genome comparisons, or their

Review

Acknowledgements

We are grateful toR. Panford for criticalreading of themanuscript. This workwas supported by grantsto W.B.F. from DeutscheForschungsgemeinschaft(Gottfried-Wilhelm Leibniz,Aminosäuretransport) and BMBF.

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s p e c t r u m o f p r o t e o g e n i c

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i n p h l o e m ( 1 0 0 – 2 0 0 m

M)

a n d x y l e m s a p ( ~ 1 0 - f o l d

l o w e r c o n c e n t r a t i o n t h a n

p h l o e m ) , a m i n e s a n d

a c i d i c a m i n o a c i d s

p r e d o m i n a t e . T h e a c t u a l

i n t r a c e l l u l a r , a p o p l a s m i c

a n d p h l o e m c o m p o s i t i o n

o f a m i n o a c i d s i s t h o u g h t

t o b e s i m i l a r ; t h u s , t h e

r e l a t i v e l y u n s p e c i f i c A A P s

m i g h t b e r e s p o n s i b l e f o r

f u n c t i o n s s u c h a s u p t a k e

o f a m i n o a c i d s f r o m t h e

x y l e m i n t o t h e c e l l a n d

p h l o e m l o a d i n g . N o

o r g a n e l l a r t r a n s p o r t e r h a s

b e e n i d e n t i f i e d t o d a t e .



146 Review

distribution and functional mechanisms. Althoughsequence and structural similarity searches areuseful to classify genes into families, the differencesobserved (e.g. in the case of MCTs regarding substraterecognition) indicate that transport function,specificity and localization need to be established foreach single protein.

In most cases, transporters recognize a broadspectrum of amino acids and derivatives. It will thusbe interesting to study recognition and transport inrelation to protein structure in more detail.Comparative analysis of the amino acid transportersuperfamilies not only yields valuable information

with respect to mechanistic and structural properties,but is also of central importance for ourunderstanding of nutrition and signaling functions inthe different classes of organisms. In the intestine,different sets of transporters are localized onbrush-border and basolateral membranes. Bycoupling to different mechanisms, these allowpolarized amino acid transport. Here, polarizedtranscellular uptake of amino acids across intestinalabsorptive cells into the bloodstream might serve as amodel for intercellular transport of amino acids; forexample, from palisade parenchyma into the vascularsystem in plants (Fig. 4).

Review

References

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18 Lynch, A.M. and McGivan, G.M. (1987) Evidencefor a single common Na+-dependent transportsystem for alanine, glutamine, leucine andphenylalanine in brush-border membranevesicles from bovine kidney. Biochim. Biophys.Acta 899, 176–184

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neurotransmitter transporters. J. Neurochem. 71,1785–1803

Ti BS

AtA

B011475

AtT45634

AtAAC35230

AtT01534

HsBNP1

HsDNP1

HsSLC17A1

HsSLC17A4

HsSLC17A2

HsSLC17A3

HsSLC17A5

ScYLL055W

SCYLR004C

ScDal5

ScTNA1

ScYIL166C

ScFEN2

ScSEO1

ScVHT1

Yeast allantoatetransporter familyAtAAD32766

1.21.00.80.6

Pro

babi

lity

0.40.20.0

0 100 200

HsBNP1

300 400 500

1.21.00.80.6

Pro

babi

lity

0.40.20.0

0 100 200

AtAAC35230

300 400 500

1.21.00.80.6

Pro

babi

lity

0.40.20.0

0 10050 200

HsSLC17A2

300150 250 350 400

Fig. 5. Phylogenetic treeof BNP1-related aminoacid transporters [38].Phylogenetic analyseswere performed asdescribed for Fig. 1. Thecomplete alignment wasbased on 695 sites; 615were phylogeneticallyinformative. The BNP1homologs can be dividedinto three differentclusters (shadowed areas).The graphs correspond tohydrophobicity plotsobtained by TMHMManalysis [47]; the redcurves correspond topredicted transmembranedomains, the blue tointracellular domains andthe pink to extracellulardomains. Green indicatesa plant gene, red an animalgene and brown a fungalgene. A high variabilityexists between the BNP1homologs in which 8–12transmembrane domainswere predicted fordifferent members.Abbreviations:At, Arabidopsis thaliana;Hs, Homo sapiens;Sc, Saccharomycescerevisiae.



147Review

22 Chang, H.C. and Bush, D.R. (1997) Topology ofNAT2, a prototypical example of a new family ofamino acid transporters. J. Biol. Chem. 272,30552–30557

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27 Bennet, M.J. et al. (1996) The Arabidopsis AUX1gene: a permease like regulator of rootgravitropism. Science 273, 948–950

28 Chen, L. et al. (2001) ANT1, an aromatic andneutral amino acid transporter in Arabidopsis.Plant Physiol. 125, 1813–1820

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Review

Pete Jeffs is a freelancer working in Paris, France.

review trends in biochemical sciences vol.27 no.3 march ...€¦ · (hats; for example, lat-1,...

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