Biotechnology of Fruit Quality - Fruit Ripening: Physiology, Signalling and Genomics - page 277

investigated primarily by the inheritance
pattern of fl ower colour and radiolabelling.
However, genetic engineering technology
has added a new dimension to our
understanding of fl avonoid biosynthetic
enzymes and substrates and their diversity
among various plant species (Table 16.2).
Mutants and transgenic plants have
provided direct evidence for the function
of various genes involved in fl avonoid
biosynthesis pathways (reviewed by
Ververidis et al. , 2007a). Flavonoids are
mainly synthesized from phenylalanine via
the phenylpropanoid pathway. Following
cinnamate hydroxylation by the cinnamate
4-hydroxylase and 4-coumarate:CoA ligase
step in phenylpropanoid pathway, the
fl avonoid biosynthesis pathway branches
out into phenolics (chlorogenic acid) and
fl avonols (naringenin, quercetin and their
derivatives) (Anterola and Lewis, 2002;
Ververidis et al. , 2007b).
Tomato fruits synthesize signifi cant
amounts of carotenoids but are poor in the
production of fl avonoids in fruit fl esh.
Flavonoid production in fruits is restricted
mainly to the peel with accumulation of
naringenin chalcone, the fl avonol rutin and
kaempferol 3- O -rutinoside (Crozier et al. ,
1997; Muir et al. , 2001; Bovy et al. , 2002).
The major focus of fl avonoid biotechnology
research is to increase fl avonoid accumu-
lation in fruit fl esh and determine the
potential to induce the production of new
fl avonoids (Table 16.2). Tomato fruit do not
have stilbene synthase gene ( StSy )
(Giovinazzo et al. , 2005) and cannot
normally produce resveratrol, a stilbenoid
fl avonoid. However, expression of a grape
StSy not only induced the production of
resveratrol in transgenic fruits but also led
to the accumulation of trans -resveratrol
and trans -resveratrol glucopyranosides
(piceid), further elevating the antioxidant
capacity in tomato fruit (D'Introno et al. ,
2009). Transgenic tomato lines were
developed that constitutively expressed
fl avonoid genes from different plant
species (Schijlen et al. , 2006). It was shown
that the expression of grape StSy produced
a higher amount of resveratrol and piceid
(a stilbenoid glucoside), while combined

expression of petunia chalcone synthase
and lucerne chalcone reductase induced
higher levels of butein and isoliquiritigenin
(deoxychalcones). Combined expression of
petunia chalcone isomerase and gerbera
fl avone synthase resulted in elevated
production of luteolin-7-glucoside, luteo-
linaglycon (fl avone) and quercetin glyco-
sides (fl avonol). Although the constitutive
overexpression of StSy produced up to
tenfold higher levels of resveratrol, it
resulted in complete male sterility,
probably due to a lack of coumaric and
ferulic acid production (Ingrosso et al. ,
2011). The seedless parthenocarpic fruit
phenotype resulting from male sterility is
of much interest because of its desirability
by both the consumer and food industry
(Rotino et al. , 1997; Ficcadenti et al. , 1999;
Pandolfi ni et al. , 2002).
The ectopic expression of petunia
chalcone isomersae in tomato resulted in a
78-fold increase in peel fl avonols, which
was mainly due to the accumulation of
rutin, a quercetin glycoside. After pro-
cessing the tomato fruit, the paste still
retained 65% of the total fl avonols present
in the fresh fruit (Muir et al. , 2001).
Although isofl avones are legume-specifi c
fl avonoids, tomato plants engineered to
constitutively overexpress soybean iso-
fl avone synthase ( 35S:GmIFS2 ) showed
signifi cant accumulation of genistin (a
major isofl avone metabolite) in leaves with
a marginal increase in fruit peel.
Naringenin chalcone biosynthesis was also
upregulated in these transgenic fruit,
indicating naringenin as a limiting factor
(substrate) for isofl avone biosynthesis in
fruit peel (Shih et al. , 2008).
In addition to the candidate gene
approach to enhance fl avonoid content,
transcription factors have also been tested
to achieve similar objectives. Coordinated
expression of maize MYB-type C1 and
MYC-type LC, transcription factors
implicated in anthocyanin production, in
tomato induced fl avonoid biosynthesis in
fruit fl esh the tissues where fl avonoids are
poorly synthesized (Bovy et al. , 2002).
Overall, a tenfold increase in total
fl avonoids and a 20-fold increase in total