Download The ATP-Binding Cassette Transporters

The ATP-Binding Cassette Transporters: Structure,
Function, and Gene Family Comparison between
Rice and Arabidopsis1
Michal Jasinski2, Eric Ducos2, Enrico Martinoia, and Marc Boutry*
Unité de Biochimie Physiologique, Institut des Sciences de la Vie, Université catholique de Louvain, Croix
du Sud 2–20, B–1348 Louvain-la-Neuve, Belgium (M.J., E.D., M.B.); and Laboratory of Molecular Plant
Physiology, Institute of Plant Biology, Zurich University, CH–8008 Zurich, Switzerland (M.J., E.M.)
Cells are separated from their external environment by a membrane barrier, which ensures that
certain ions, metabolic intermediates, and macromolecules, such as proteins, remain within the cell. However, life is not possible without the exchange of
material and information with the external environment. Therefore, during evolution, efficient transport
systems have been developed to allow interaction
with the environment, and they ensure that essential
ions and metabolites enter the cell and other compounds leave it.
Many molecules are transported against a concentration gradient, requiring the use of energy. For
several transport systems, this energy is directly provided by ATP hydrolysis. Well known examples of
ATP-powered transport systems are cation pumps,
such as the Ca2⫹-, H⫹-, Na⫹/K⫹-, and H⫹/K⫹ATPases. The ATP-binding cassette (ABC) transporters, which constitute another major class of ATPpowered transporters, are less well characterized,
and the physiological function of many is still
unknown.
Here, we will review the structure and catalytic
mechanism of these enzymes, tackle their mysterious
capacity to transport a variety of substrates with
unrelated structures, compare the Arabidopsis and
rice (Oryza sativa subsp. japonica) gene family composition, and finish by mentioning examples of the
physiological roles of ABC transporters in plants.
Some of these topics have been reviewed in detail
(Theodoulou, 2000; Sanchez-Fernandez et al., 2001;
Martinoia et al., 2002; Rea et al., 2002).
1
This work was supported by grants from the Belgian Interuniversity Poles of Attraction Program, the European Commission
(Marie Curie Research Training Network), and the Belgian Fund
for Scientific Research and the Swiss National Foundation. E.
Ducos holds an individual Marie Curie European fellowship.
2
These two authors contributed equally to the paper.
* Corresponding author; e-mail [email protected]; fax
32–10 – 473872.
www.plantphysiol.org/cgi/doi/10.1104/pp.102.014720.
STRUCTURAL CHARACTERISTICS OF
ABC TRANSPORTERS
The ABC transporter family is very large (79 members in Escherichia coli, 29 in Saccharomyces cerevisiae,
and 131 in Arabidopsis) and its members are found
in all organisms. The encoded enzymes couple ATP
hydrolysis to the transport, across various membranes, of a great variety of substrates, including
lipids, heavy metal ions, inorganic acids, glutathione
conjugates, sugars, amino acids, peptides, secondary
metabolites, and xenomolecules, used as drugs (Higgins, 1992; Rea et al., 1998).
The ABC genes have been divided into several
clusters or subfamilies corresponding to phylogenetic pathways and structural features. The names
used to define the subfamilies are often historic and
related to the function of drug resistance, although
some members are clearly not involved in drug
transport and may, for example, function as a channel or channel regulator. The three best characterized subfamilies are the pleiotropic drug resistance
(PDR), multidrug resistance (MDR), and multidrug
resistance-associated protein (MRP) subfamilies.
MOTIFS AND DOMAINS
ABC proteins have a characteristic modular structure consisting of a double set of two basic structural
elements, a hydrophobic transmembrane domain
(TMD) usually made of six membrane-spanning
␣-helices, and a cytosolic domain, the latter containing a region involved in ATP binding and known as
the nucleotide-binding domain (NBD; for review, see
Higgins, 1992).
Although most prokaryotic ABC proteins are encoded as separate TMD and NBD subunits or as
one-half-size (one TMD and one NBD) transporters,
in the majority of eukaryotic ABC transporters, all
four domains (two TMDs and two NBDs) are contiguous on a single polypeptide, constituting a fulllength transporter in the “forward” TMD1-NBD1TMD2-NBD2 orientation in the MDR subfamily or
the “reverse” NBD1-TMD1-NBD2-TMD2 orientation
in the PDR subfamily (Fig. 1A). In the MRP subfam-
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Jasinski et al.
ily, the arrangement is in the forward orientation, but
some MRPs (e.g. MRP1) have five ␣ helices (sometimes designated as TMD0) at the N terminus.
THE THREE-DIMENSIONAL STRUCTURE
Figure 1. Topology and putative catalytic mechanism of ABC transporters. A, Topology. The TMD, with the predicted membrane spans
(zig-zag), and the NBD, containing the nucleotide binding fold, are
shown for the three main types of full-size ABC transporters. B,
Putative catalytic mechanism. The figures schematize an ABC transporter with its four domains, two of which (TMDs) are embedded in
the membrane. A substrate (S) coming from the cytosol or already
within the membrane binds to a substrate binding site in one of the
TMDs (this site has not yet been localized precisely and might, as a
matter of fact, be one among several binding sites) (I3II). This is
followed by ATP (T) binding to the nucleotide-binding site localized
at the interface between both NBDs (note that only one of the two
nucleotide binding sites is represented) (II3III). ATP binding and
hydrolysis is accompanied by a structural rearrangement that allows
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Deciphering the molecular mechanism of the coupling between ATP hydrolysis and substrate transport represents a major challenge requiring the solution of the protein atomic structure. Despite the
tremendous problems involved in the crystallization
and x-ray analysis of membrane proteins, several
recent successes have been reported. The structure of
several NBDs has been determined in bacterial ABC
transporters as, in this case, these soluble subunits
are synthesized independently from the membrane
subunits and are thus more amenable to crystallization and x-ray diffraction analysis.
A high-resolution structure of two E. coli entire
ABC transporters has been determined: MsbA
(Chang and Roth, 2001) and BtuCD (Locher et al.,
2002) involved in lipid and vitamin B12 transport,
respectively. There are some discrepancies between
these structures, as well as between the structures
obtained for soluble NBDs. For instance, although
there is a consensus for the structure of a single NBD,
there are divergences concerning the interactions between a pair of NBDs. These are largely separated in
MsbA and are closely interacting in BtuCD. This
might be related to a crystallization artifact joining
together two one-half-size transporters in a nonnative conformation. Alternatively, these two conformations might represent two different states of the
catalytic cycle. The BtuCD model shows that a single
NBD does not constitute an independent functional
structure by itself, but that both NBDs interact, with
the ABC signature of one NBD interacting with the
other motifs from the other NBD to constitute an
ATP-binding site. Thus, there are two ATP-binding
sites lying at the interface between the two NBD
subunits. A similar interaction was found in Rad50,
an enzyme involved in DNA repair and sharing a
common ancestor with ABC NBDs (Hopfner et al.,
2000). In contrast to the BtuCD study, Rad50 was
crystallized in the presence of ATP. Comparison of
the two structures suggests that ATP binding induces
a closer interaction of the signature motif of one NBD
with the other NBD. This proposed interaction is also
in agreement with older biochemical observations
reporting cooperativity of ATP binding and hydrolysis. A recent 22 Å projection structure of the mouse
the substrate to access to, and to be translocated through, a membrane path (III3IV). Upon ADP (D) and phosphate dissociation, the
protein returns to the initial state (IV3I). This still putative model is
based on that proposed for the bacterial BtuCD (Locher et al., 2002)
except for the substrate path, which is inverted. Alternatively, the
central cavity might represent the substrate-binding site (discussed in
the text).
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Plant Physiol. Vol. 131, 2003
ATP-Binding Cassette Transporters
P-glycoprotein, an MDR-type ABC transporter, confirmed the close association between the two NBDs
(Lee et al., 2002).
Because of the large structural variety of substrates
transported by ABC carriers, the membrane domains,
which are involved in substrate binding and translocation, are poorly conserved at the primary structural
level and are expected to differ markedly at the threedimensional level. In the E. coli BtuCD and MsbA
structures mentioned above, a large central cavity
was found between the two TMDs within the membrane; this is lined with hydrophobic residues, has
two openings toward the membrane, and might represent the translocation pathway (Locher et al., 2002).
The BtuCD structure and others, as well as many
biochemical data, provide some clues about the catalytic cycle. The following succession of events can
be proposed (Fig. 1B). The transported substrate,
coming from the cytosol if it is hydrophilic or directly
from the inner membrane leaflet if it is hydrophobic
or amphiphilic, binds to a site in the membrane domain. This promotes the binding of ATP at the NBD
site. ATP hydrolysis (and possibly ATP binding) is
accompanied by a structural rearrangement of the
NBD that extends to the membrane domains and
induces translocation of the substrate through the
inner path toward the other face of the membrane.
Finally, the transporter returns to its initial configuration upon ADP and phosphate dissociation.
MULTISUBSTRATE TRANSPORTERS
One surprising observation regarding ABC transporters was the wide range of structurally and functionally unrelated compounds that are transported,
raising the question of how a single protein can bind
and transport such a remarkably diverse collection of
substrates. The modular construction of ABC transporters and the presence of two homologous TMDNBD units raise the possibility that each can contribute to the binding and transport of different classes of
compounds. Such a conclusion was drawn from
transport kinetics. For instance, Arabidopsis AtMRP2
and AtMRP3 can simultaneously transport endogenous substrates, such as chlorophyll catabolites, and
a model glutathionated compound, S-(2, 4-dinitrophenyl) glutathione, neither of which interferes
with the transport of the other (Lu et al., 1998;
Tommasini et al., 1998). However, detailed studies
of AtMRP2 transport multispecificity, using the
above-mentioned chlorophyll catabolites and S-(2,
4-dinitrophenyl) glutathione, together with other
substrates for plant MRPs, such as glucuronide 17
␤-estradiol, glutathionated metolachlor, and glutathione, showed more complex relationships at the
binding site and transport levels that were difficult to
explain on the basis of only two binding sites, leading
to the proposal that AtMRP2 has several substrate
binding sites (Klein et al., 1998; Liu et al., 2001).
Plant Physiol. Vol. 131, 2003
For some mammalian ABCs, binding sites for
drugs have been proposed on the membranespanning domains. Many drug substrates of multidrug transporters are lipophilic or amphiphilic molecules. On the basis of their preferential partitioning
in the membrane, it has been proposed that some
drugs directly access ABC transporters within the
membrane bilayer.
Neyfakh (2002) recently proposed that the large
membrane chamber mentioned above might actually
contain multisubstrate binding sites. Substrates
would establish a number of van der Wall’s interactions with surrounding hydrophobic residues. The
size of the binding pocket is large enough to accommodate different substrates interacting with different
residues forming the pocket walls. Considering the
fact that most of the ABC transporter substrates are
not genuinely hydrophobic and do possess polar,
besides hydrophobic, functionalities, Neyfakh (2002)
proposes that these molecules can interact with a few
charged and polar residues located within the walls
of the membrane cavity. The major interest of this
hypothesis is that it gives a simple explanation for
the multiple structurally dissimilar substrates of ABC
transporters. Clearly, molecular information about
substrate binding and transport must await the detailed three-dimensional structure determination of
several ABC transporters. This should uncover a
common mechanism for the coupling of ATP hydrolysis and substrate translocation. In addition, the determination of the three-dimensional structure of
ABC translocators crystallized with their substrates,
together with site-directed mutagenesis, should unravel the mystery of multiple substrates.
THE PLANT ABC GENE FAMILY
Several transporters have been characterized in
plants and, in some cases, their physiological roles
have been identified. However, the great majority of
plant ABC transporters remain uncharacterized. The
Arabidopsis ABC gene family alone contains 131
members, including 54 full-size transporters (consisting of two hydrophobic and two hydrophilic domains; Sanchez-Fernandez et al., 2001; Martinoia et
al., 2002) and, therefore, it is still too early to draw a
general picture of their roles. It might turn out that
the diversity of their roles equates with the size of the
gene family. The recent sequencing of 93% of the rice
genome (Goff et al., 2002) has allowed us to retrieve
putative ABC genes and to analyze how the ABC
gene family has evolved since monocots and dicots
diverged. We limited this analysis to the full-size
transporters, for which biochemical and physiological data are available.
We identified 45 rice sequences that putatively encode full-size ABC transporters. As in Arabidopsis,
almost all of these (44) belong to the three main PDR,
MDR, and MRP subfamilies. We also found one gene
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Jasinski et al.
homolog to PXA/PDE/CTS (Zolman et al., 2001;
Footitt et al., 2002; Hayashi et al., 2002). We did not
find any rice sequence homologous to AtAOH1, the
unique Arabidopsis representative of the so-called
ABC1 subfamily, which is highly represented in the
human genome.
To simplify the comparison, phylogenetic trees
were obtained separately for the PDR, MDR, and
MRP subfamilies (Fig. 2). It is clear that most of the
rice ABC genes identified have at least one Arabidopsis homolog with which they share more than
60% amino acid identity. This is reflected by the
observation that Arabidopsis and rice genes are
found together in many branches within a subfamily.
Furthermore, some of the exceptions might disappear when the sequence of the remaining 7% of the
rice genome is deciphered. Therefore, we can conclude that the overall organization of the full-size
ABC genes is similar in Arabidopsis and rice and
thus predates the separation between monocots and
dicots. However, because there are rice branches in
which no Arabidopsis gene was found, we have to
consider that, in some branches, rice or Arabidopsis
genes have been lost during evolution.
On the other hand, inspection of the various
branches within each subfamily shows that the number of genes within a subfamily branch is not the
same for both species, indicating that gene duplication or deletion occurred after monocots and dicots
diverged. For instance, five rice genes were identified
in the branch in which the single AtPDR9 is found,
whereas only one rice MRP gene was found in the
same branch as the four Arabidopsis genes AtMRP1,
2, 12, and 13. In the absence of any biochemical data
on most of these genes and their encoded products,
the reason for this asymmetry is not clear. A larger
number of genes might lead to enzymes with more
specialized functions related to species evolution or
simply to a higher expression level. It could also be
that a single gene with a broad expression is replaced
Figure 2. Phylogenetic trees showing the predicted relationships
between plant MDR, MRP, or PDR ABC transporters. CLUSTAL W
1.82 (Thompson et al., 1994) was used to align full-length ABC
transporters sequences belonging to the MDR, MRP, or PDR subfamilies. Distance matrix, bootstrap values (1,000 replicates) and phylogenetic trees were calculated with CLUSTAL W. Accession numbers of the ABC transporters sequences from rice are reported in
Table I. MDR and MRP Arabidopsis sequences were recovered according to the NCBI accession numbers described by SanchezFernandez et al. (2001). PDR sequences are named as described by
Martinoia et al. (2002), offering a more exhaustive list. Accession
numbers BAB85651, BAB62040, AAF23176, AAD10836, CAA71179,
CAA94437, CAC40990, and BAB92011, correspond, respectively, to
the proteins TaMDR1 (wheat, Triticum aestivum), CjMDR1 (Coptis
japonica), CMDR1 (cotton, Gossypium hirsutum), PMDR1 (potato,
Solanum tuberosum), HvMDR2 (barley, Hordeum vulgare), SpTUR2
(Spirodela polyrrhiza), NpABC1 (wild tobacco, Nicotiana plumbaginifolia) and NtPDR1 (cultivated tobacco, Nicotiana tabacum). Arabidopsis (At) ABC transporters are in green, Rice (Os) ABC transporters are in
red, and other species and bootstrap values are in black.
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Plant Physiol. Vol. 131, 2003
ATP-Binding Cassette Transporters
by several genes, each having a more restricted expression or a tighter regulation at the transcriptional
or posttranscriptional level. This kind of evolution is
expected to provide the plant with more flexibility.
For instance, the plant kingdom is characterized by a
large diversity in secondary metabolite production.
Therefore, adapted transporters might have been required to transfer these metabolites into the external
medium or the adequate internal compartment.
It is interesting to note that in some branches, rice
and Arabidopsis homologs are very closely related
(e.g. 78% identity between AtMDR14 with OsMDR10),
whereas this is not the case in other branches (e.g.
57.4% identity between AtMDR22 and OsMDR17). It
is tempting to speculate that the highly conserved
homologs are required for basic functions that have to
be fulfilled in both plants, whereas the more distantly
related homologs might have evolved more rapidly
and adapt to plant specialization. For instance, AtMDR11 mentioned above is proposed to be involved
in auxin transport, which might be considered as an
essential function. The high identity shared with the
rice OsMDR11 and OsMRD12 is in the same range as
that observed between rice and Arabidopsis genes
encoding plasma membrane H⫹-ATPases, which belong to a highly functionally conserved transporter
family (Arango et al., 2002).
Generally, intron organization confirms the general
subdivision within each tree (see “Supplementary
Material”). For instance, MDR genes cluster into two
main subdivisions: OsMDR3 to OsMDR9, all closely
related, and OsMDR10 to OsMDR16, more distant
among themselves. OsMDR1 and OsMDR2, on the
one hand, and OsMDR17, on the other hand, are
more individualized. In the MRP subfamily, OsMRP1
singles out by still possessing 26 introns, whereas
all the other OsMRPs have no more than 12 introns.
The other genes are divided into two clusters with
OsMRP2 to OsMRP7, on the one hand, and OsMRP8
to OsMRP11, on the other hand. Finally, the PDR
tree is too compact and the intron organization is
too conserved to clearly identify clusters. In agreement with this, the PDR subfamily is that for which
the highest global identity was found.
FUNCTIONS OF PLANT ABC TRANSPORTERS
Compared with our extensive knowledge of the
organization and sequences of the ABC gene family,
functional data are still sketchy. Multiple functions,
such as MDR, described for ABC transporters in
other taxa, triggered research into plant homologs.
Possible roles, such as cross-resistance to toxic compounds, including herbicides, have been postulated
for plant ABC proteins. Two main events started the
plant ABC odyssey, these being the cloning and characterization of Arabidopsis AtPGP1 encoding a homolog of human MDR1 and MDR3 (Dudler and Hertig, 1992), and biochemical studies on the uptake of a
Plant Physiol. Vol. 131, 2003
glutathione S-conjugate into barley vacuoles associated with an ABC transporter (Martinoia et al., 1993).
Since then, rapid progress in the identification and
characterization of plant ABC transporter genes has
resulted in the discovery of a large diversity of functions for the plant ABC family (for review, see Rea et
al., 1998, 2002; Rea, 1999; Davies and Coleman, 2000;
Theodoulou, 2000; Sanchez-Fernandez et al., 2001;
Martinoia et al., 2002). Several examples of characterized plant MDR-, MRP-, and PDR-type ABC transporters are described below.
THE MDR SUBFAMILY
Genome sequencing revealed 22 MDR-like genes in
Arabidopsis (Sanchez-Fernandez et al., 2001) and 17
in rice (Fig. 2). At this point, it is worth mentioning
that because they show sequence homology with
human MDR1, all ABC-type proteins are described as
MDRs, despite the fact that, in several cases, their
physiological role might be unrelated to drug resistance. This is, for instance, the case for STE6 from
Saccharomyces cerevisiae, which codes for a transporter
of the sex pheromone peptide (Kuchler et al., 1989).
One of the two MDR clusters mentioned above
contains 17 closely related genes (ⱖ 54.1 identity),
eight from Arabidopsis, seven from rice, and two
from other species: CjMDR1, which was isolated
from alkaloid-producing cultured cells of Coptis japonica (Yazaki et al., 2001), and TaMDR1, which is
induced in wheat root apices by aluminum (Sasaki et
al., 2002). The other cluster contains more divergent
MDRs, including some isoforms that seem to be species specific, because no close homolog was found in
the other species (i.e. OsMDR15 and OsMDR16).
Except for the Arabidopsis AtPGP1 gene mentioned above, few MDRs have been characterized in
detail at the functional level. AtPGP1 is expressed in
several plant organs, including the apical regions of
shoots and roots, and it has been suggested that
AtPGP1 might export a signal (peptide hormone)
from the shoot apical region. In situ immunofluorescence and subcellular fractionation methods have
shown that AtPGP1 is localized in the plasma membrane. It has been suggested to be involved in hypocotyl cell elongation (Sidler et al., 1998).
AtPGP1 (called AtMDR1 in Sanchez-Fernandez et
al., 2001 and in this paper) is the closest homolog to
AtMDR1 (called AtMDR11 in Sanchez-Fernandez et
al., 2001 and in this paper). Interestingly, Noh et al.
(2001) observed epinastic cotyledons with AtMDR1
knockout plants. Double AtPGP1 and AtMDR1
knockouts exhibit a more pronounced phenotype,
with a dwarf phenotype and twisted leaves. These
authors proposed that AtMDR1, probably in conjunction with AtPGP1, is required for the transport of
auxin from the apical meristematic regions, where
auxin synthesis takes place, to basal tissues.
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Jasinski et al.
YCF1 has been shown to be a vacuolar glutathione
conjugate pump conferring cadmium resistance (Szczypka et al., 1994; Li et al., 1996). Finally, the observation of the uptake of glutathione S-conjugates of
N-ethylmaleimide and metolachlor into barley vacuoles (Martinoia et al., 1993) triggered the identification of plant MRPs. In Arabidopsis and in rice, 15 and
12 MRP genes, respectively, have been identified. In
contrast to MDR transporters, MRPs are defined by a
hydrophobic N-terminal extension. Biochemical studies of the transport properties of AtMRP1, AtMRP2,
and AtMRP3 provided evidence for their role in the
export from the cytosol of exogenous or endogenous
compounds (Lu et al., 1997; Lu et al., 1998; Tommasini
et al., 1998). To date, vacuolar localization has only
been demonstrated for AtMRP2. During the cellular
detoxification process, potentially toxic compounds
are modified and transformed in more hydrophilic
compounds by conjugation to glutathione, Glc, glucuronide, or malonyl moieties (Kreuz et al., 1996) and
transported into the vacuole by plant MRPs (Martinoia et al., 2000).
Although AtMRP1, AtMRP2, and AtMRP3 are
all glutathione conjugate transporters, AtMRP2 and
AtMRP3 can also transport chlorophyll catabolites,
whereas AtMRP1, the closest homolog of AtMRP2,
cannot (Lu et al., 1998). This clearly suggests that
intraspecies and a fortiori interspecies sequence homology is not necessarily linked to the identical
function.
In animals, some ABC proteins have channel activity or regulate heterologous channels. This is the case
for the cystic fibrosis transmembrane conductance
regulator and the sulfonylurea receptor. Studies on
ion flux in guard cells using ion channel regulators
that alter ion conductance, e.g. glibenclamide, suggested the presence in plants of cystic fibrosis transmembrane conductance regulator- or/and sulfonylurea receptor-like proteins involved in stomata
movement (Leonhardt et al., 1997, 1999). Recently,
AtMRP5, expressed in the vascular bundle and epidermis, especially in guard cells, has been suggested
to function as an ion channel regulator (Gaedeke et
al., 2001) because in contrast to wild-type plants,
stomata in a knockout AtMRP5 mutant did not open
upon glibenclamide addition.
THE PDR SUBFAMILY
THE MRP SUBFAMILY
This subfamily was originally identified in drugresistant human lung cancer cell lines (MRP1; Cole et
al., 1992). MRPs are able to transport glutathione and
glucuronide conjugated to toxic compounds out of
the cytosol and have been studied in yeast, in which
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Starting from the observation of drug resistance in
the yeast, S. cerevisiae, another ABC transporter subfamily was identified and designated as PDR. The
yeast PDR5 gene was originally isolated as coding for
a transporter mediating resistance to cycloheximide
(Balzi et al., 1994), but PDR5 has now been shown to
mediate resistance to a wide range of structurally and
functionally unrelated compounds (Kolaczkowski et
al., 1996) and is considered a functional homolog of
human MDR1, despite the structural differences (Fig.
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Plant Physiol. Vol. 131, 2003
ATP-Binding Cassette Transporters
1). The Arabidopsis and rice genomes contain 15
PDR-like genes (Fig. 2). Compared with the other two
subfamilies, the PDR subfamily displays the least
divergence.
None of the Arabidopsis PDR genes has been characterized in detail. However, data from other plant
species are available. Smart and Fleming (1996) isolated a homolog of yeast PDR5, designated as TUR2,
from the aquatic plant, great duckweed (Spirodela
polyrrhiza). The SpTUR2 transcript is up-regulated by
abscisic acid and by environmental stresses, including low temperature, high salt, or cycloheximide.
Another member of this subfamily, NpABC1, has
been identified in wild tobacco. In vitro and in situ
immunodetection showed NpABC1 to be localized in
the plasma membrane. NpABC1 is expressed in the
leaf epidermis. In leaf tissues and in cell culture, its
expression is strongly enhanced by sclareol, an antifungal diterpene produced at the leaf surface of Nicotiana species, and, in parallel with NpABC1 induction,
cells acquire the ability to excrete a labeled sclareol
analog. These data suggest that NpABC1 is involved
in the secretion of secondary metabolites that play a
role in plant defense (Jasinski et al., 2001). Recently,
SpTUR2, mentioned above, was shown to confer resistance to sclareol in Arabidopsis; however, resistance was not extended to other drugs tested (van
den Brüle et al., 2002). This led the authors to suggest
that in contrast to the fungal PDR transporters, which
are characterized by a large substrate spectrum, plant
PDR transporters might have a much higher substrate specificity.
Recently, another member of the PDR subfamily,
NtPDR1 from cultivated tobacco, has been identified
and its expression shown to be induced by various
microbial elicitors (Sasabe et al., 2002), which is further support for the idea that some PDR genes might
be involved in plant defense. In the same branch as
NpABC1, NtPDR1, and SpTUR2, a single Arabidopsis
gene, AtPDR9, and five rice genes have been identified (Fig. 2).
paired fatty acid catabolism and, consequently, developmental arrest unless Suc is added to the growth
medium, probably due to strongly reduced peroxisomal import of fatty acyl-, indole-3-butyric acid-, and
2,4-dichlorophenoxybutyric acid-CoA derivatives.
The same mutant (called cts by Footitt et al., 2002) was
identified genetically as having a very strong effect on
germination. cts mutants do not germinate and exhibit
a forever dormant phenotype. These authors suggest
that CTS could have an additional function to the
transport of acylCoAs, which involves the removal of
dormancy and promotion of germination potential.
OTHER SUBFAMILIES
Several other subfamilies of ABC transporters have
been identified in Arabidopsis (described in detail in
Sanchez-Fernandez et al., 2001). One-half-size ABC
transporters with a single TMD-NBD module are as
numerous as the full-size transporters (two TMDNBD modules). Among these, a large group (29
members) of WBC-type transporters has been identified by homology with the White, Scarlet, and Brown
ABC transporters from Drosophila, which transport
pigment precursors (Sanchez-Fernandez et al., 2001).
However, these are currently poorly characterized,
except for Arabidopsis STA1, which is localized in
mitochondria, is involved in the maturation of cytosolic Fe/S proteins, and represents a functional ortholog of yeast Atm1p (Kushnir et al., 2001).
Finally, 26 putative soluble ABC proteins containing one or two NBDs, but no TMD, have been predicted in Arabidopsis (Sanchez-Fernandez et al.,
2001) and are expected to work in association with
other proteins. Only one has been studied, AtABC1,
which is a plastid protein with a single NBD. Disruption of the gene results in reduced responsiveness to
far-red light and the accumulation of protoporphyrin
IX, suggesting that AtABC1 transports this precursor
and is involved in intercompartmental communication of light signaling (Moller et al., 2001).
PEROXISOMAL ABC-TYPE TRANSPORTERS
Yeast and animal peroxisomal membranes contain
forward-orientation, one-half-molecule ABC transporters that catalyze the transport of long-chain acylcoenzyme A (CoA) substrates. In plants, a peroxisomal full-size ABC transporter, PXA1 or PED3, has
very recently been described (Zolman et al., 2001;
Hayashi et al., 2002), and a rice homolog was found in
the rice genome. In Arabidopsis, the corresponding
mutants were originally isolated on the basis of the
resistance of their roots to growth inhibition and the
promotion of lateral root initiation by indole-3-butyric
acid and its analog 2,4-dichlorophenoxybutyric acid.
Mapping of the insertion revealed that it had inactivated a full-size ABC transporter gene. Arabidopsis
seedlings with a nonfunctional PXA1/PED3 show imPlant Physiol. Vol. 131, 2003
CONCLUSION
Because of the large variety of substrates translocated and, as a consequence, their multiple physiological roles, ABC transporters have been at the center of interest for many years. Major questions for the
future concern their structure-function relationships,
which will require the elucidation of their threedimensional structures and multisubstrate binding
properties and of the membrane transport pathway.
Although the general organization of the ABC gene
family is conserved between rice and Arabidopsis,
there are differences at a more detailed level. In the
phylogenetic tree, some branches are specific to or
more subdivided in either species, possible related to
functional specialization. Therefore, this observation
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Jasinski et al.
shows the interest to combine experimental approaches on several species.
The prediction of a large number of ABC proteins
in Arabidopsis and in rice, the largest number reported to date, implies the versatility of these enzymes
and their importance in plants. Being immobile, plants
have been forced to develop a sophisticated enzymatic
machinery to cope with the environment, and the
ABCs appear to be an important part of this machinery. In contrast to the situation in other organisms,
studies of plant ABC proteins are still at an early stage,
and new information on members of the plant ABC
family is impatiently awaited.
ACKNOWLEDGMENTS
The authors thank the Torrey Mesa Research Institute and, in particular,
Dr. S. Goff for providing them with access to the Rice Database search
program.
Received September 17, 2002; returned for revision October 30, 2002; accepted December 10, 2002.
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