The leucoanthocyanidins can be oxidized by
anthocyanidin synthase (ANS), also called leu-
coanthocyanidin dioxygenase (LDOX), a non-
heme iron 2-oxoglutarate-dependent oxygenase
to form anthocyanidin. The glycosylation of this
compound (along with methylation and acyla-
tion) forms the stable color pigments anthocya-
nins. The anthocyanidins are also substrates for a
second PA-specific enzyme, anthocyanidin
reductase (ANR) that catalyzes their reduction to
2,3-cis flavan-3-ols, for example epicatechin,
which is the predominant extension unit in PAs
ofLotusspp. The ANR enzyme is also a member
of the RED super family and wasfirst identified
in Arabidopsis (Xie et al. 2003 ). That anthocy-
anidins are a substratefits with the observation
that ANS is essential for PA biosynthesis in
Arabidopsis (Abrahams et al. 2003 ). TwoANR
genes have been identified inL. corniculatus;
LcANR2appears to be a pseudogene as it lacks
several exons, whileLcANR1when expressed in
E. coli was able to catalyze the formation of
epicatechin (Paolocci et al. 2007 ). InL. japonicus,
the gene corresponding toLcANR1is probably
Chr4.CM1616.680.
Theflavan-3-ol precursors of PAs accumulate
on the cytoplasmic side of the ER, but PA poly-
merization is believed to occur in the vacuole. The
process of glycosylation, transport, and polymer-
ization of PAs is still poorly understood, and we
only mention it briefly. The first transporter
identified in PA biosynthesis was the Arabidopsis
TT12, a multidrug and toxic compound extrusion
(MATE) family protein (Debeaujon et al. 2001 ).
AM. truncatulatransporter, MATE1, comple-
ments the Arabidopsistt12mutant phenotype and
both Arabidopsis TT12 and Medicago MATE1
appear to prefer epicatechin 3′-O-glucoside as
substrate (Zhao and Dixon 2009 ). A glycosyl-
transferase with activity toward epicatechin,
UGT72L1, was identified by transcript profiling
of Medicago hairy roots expressing the Arabid-
opsis transcription factor TT2 (Pang et al. 2008 ).
The closest related gene toMATE1in the genome
ofL. japonicusis chr2.LjT36E17.20.
14.3 Isoflavonoids and Their Role
in Plant–Microbe InteractionsIsoflavonoids represent a different branch of the
phenylpropanoid pathway and are considered
characteristic defense compounds of legumes,
although they may also act as signaling mole-
cules in symbiotic interactions. Like PAs, isofl-
avonoids have gained interest because of their
proposed health benefits in humans, such as a
reduced incidence of hormone-related cancers
attributed to their phytoestrogen properties
(Cornwell et al. 2004 ). The estrogenic activity of
isoflavonoids in forage legumes can lead to
breeding and infertility problems in farm ani-
mals, a condition known as clover disease
(Mustonen et al. 2006 ).
Isoflavonoids differ from otherflavonoids by
the phenolic B-ring being attached to the C-3 of
the C-ring instead of to the C-2 like in other
flavonoids (Fig. 14.2). Although considered a
characteristic class of compounds for the Faba-
ceae, isoflavonoids are not unique to them and
have been reported in plant species from at least
59 other plant families (Reynaud et al. 2005 ;
Lapcik 2007 ). For example, a wide variety of
isoflavones and their glycosides occur in the
genusIris, also demonstrating their presence in
both monocots and dicots (Wang et al. 2010 ). It
should, however, be noted that some reports of
the occurrence of isoflavonoids in non-legume
species relate to the presence of trace amounts
that may result from side reactions of enzymes
with related primary functions. Such enzymatic
promiscuity is common in plant-specialized
metabolism and provides the evolutionary origin
for the selection of alternative enzymatic func-
tionalities and the emergence of new biosynthetic
pathways (Khersonsky and Tawfik 2010 ).
Isoflavonoids function as phytoalexins, and
their biosynthesis is induced in response to, for
instance, microbial infection or experimental
treatments with elicitors such as reduced gluta-
thione. Isoflavonoids known to be produced by
L. japonicus, and the relatedL. corniculatus, are152 A.M. Takos and F. Rook