Plant Molecular Biology 41: 259–268, 1999.
© 1999 Kluwer Academic Publishers. Printed in the Netherlands.
Cloning and expression of amino acid transporters from broad bean
, Laurence Maurousset
, Mechthild Tegeder
Laboratoire de Physiologieet Biochimie V´eg´etales, ERS CNRS 6099, Bâtiment Botanique, UFR Sciences, Univer-
sit´e de Poitiers, 40 avenue du Recteur Pineau, 86022 Poitiers C´edex, France (
author for correspondence);
Physiology, Zentrum für Molekularbiologie der Pﬂanzen (ZMBP), Universität Tübingen, Auf der Morgenstelle 1,
72076 Tübingen, Germany
Received 22 March 1999; accepted in revised form 8 August 1999
Key words: amino acid transport, carrier, expression analysis, Vicia
This work describes the isolation of a full-length (VfAAP2) and three partial amino acid transporter genes (VfAAPa,
VfAAPb, VfAAPc) from broad bean (Vicia faba L.). The function of VfAAP2 was tested by heterologous expression
in a yeast mutant deﬁcient in proline uptake. VfAAP2 mediates proton-dependent proline uptake with an apparent
of about 1 mM. Analysis of substrate speciﬁcity by competition experiments showed that aromatic amino
acids, neutral aliphatic acids and L-citrulline are the best competitors, whereas basic amino acids do not compete
with proline. Northern analysis indicates that all VfAAPs exhibit different patterns of expression. VfAAP2 is most
strongly expressed in the stem and at a lower level in sink leaves and pods. VfAAPa, VfAAPb and VfAAPc are most
strongly expressed in the ﬂowers, but their expression in the other organs varies.
Abbreviations: PCMBS, p-chloromercuribenzene sulfonic acid; CCCP, carbonyl cyanide m-chlorophenyl hydra-
zone; DEPC, diethylpyrocarbonate
After uptake by the roots, nitrogen from the soil is in-
corporated into amino acids and ureides in the plant.
Exchange of nitrogen between the different organs
occurs both in the phloem and in the xylem mainly
in the form of amino acids and ureides. Although
asparagine/aspartate and glutamine/glutamate are usu-
ally the preferred transport form of nitrogen, all amino
acids are found in the phloem sap (Winter et al., 1992).
Long-distance transport of amino acids is necessary
not only for coordinated plant growth and develop-
ment, but also for efﬁcient storage of proteins in the
fruits, some amino acids being especially important
for the protein balance of human and cattle foods.
The nucleotide sequence data reported will appear in the
EMBL, GenBank and DDBJ Nucleotide Sequence Databases un-
der the accession numbers Y09591 (VfAAP2), AF061434 (VfAAPa),
AF061435 (VfAAPb) and AF061436 (VfAAPc).
Amino acid transport in plants has therefore been
the focus of numerous studies since many years.
Tracer experiments have resulted in detailed patterns
of nitrogen exchange among the different parts of the
plant along with the development of various species,
and in modelling of these exchanges (Pate, 1980,
1989). Early physiological studies showed that amino
acid transport across the plant plasma membrane is
mediated by several transporters exhibiting a wide
speciﬁcity, and energized by co-transport with protons
and possibly other cations (Etherton and Rubinstein,
1978; Kinraide and Etherton, 1980; Kinraide, 1981;
Despeghel and Delrot, 1983; Mounoury and Delrot,
1984; Wyse and Komor, 1984). Cell fractionation and
transport studies with puriﬁed plasma membrane vesi-
cles from sugar beet leaves indicated the presence of
at least 4 transport systems differing by their sub-
strate speciﬁcity (Li and Bush, 1990, 1991; Bush,
1993). Use of plasma membrane vesicles from Ricinus