Biotechnology of Fruit Quality - Fruit Ripening: Physiology, Signalling and Genomics - page 271

rot, a calcium-associated fruit disorder (de
Freitas et al. , 2012). These authors showed
that apoplastic calcium levels increased
due to reduced calcium binding to high
methoxyl pectin, a consequence of low
PME activity, and infl uenced the develop-
ment of blossom end rot symptoms in
tomato fruit (de Freitas et al. , 2012).
Preferential loss of galactose and/or
arabinose from cell walls during early fruit
ripening has led to the suggestion that
E -galactosidase plays an important role in
fruit textural changes (Gross and Sams,
1984). Among the seven E -galactosidase
genes ( SlTBG1-7 ) expressed in developing
fruit, only silencing of SlTGB4 (about 90%
reduction in extractable exogalactanase
activity) led to about 40% increase in fruit
fi rmness compared with the wild-type
fruits at comparable stages of ripening
(Smith and Gross, 2000; Smith et al. , 2002).
Total exogalactanase activity, cell-wall
galactose content and fruit softening were
not affected in transgenic fruit exhibiting
an approximate 90% reduction in SlTBG1
transcripts obtained by homology-
dependent gene silencing (Carey et al. ,
2001). The role of endo- E -mannanase,
which hydrolyses mannose in hemi-
cellulose polymers to mannobiose and
mannotriose, was tested by developing
transgenic plants expressing its antisense
RNA or a gene-specifi c hairpin RNAi gene.
These transgenic fruits exhibited reduced
endo- E -mannanase activity, but a clear
correlation between fruit fi rmness and
endo- E -mannanase activity was not found
(Bewley et al. , 2000).
The role of xyloglucan xyloglucosyl
transferase/endohydrolase ( XTH ) in fruit
textural changes was examined by over-
expressing SlXTH1 , a tomato homologue of
the Nicotiana tabacum NtXET-1 gene, under
the control of CaMV 35S promoter (Miedes
et al. , 2010). XTHs have been suggested to
play dual roles in cell-wall chemistry by
integrating newly secreted xyloglucan
chains into an existing cell-wall-bound
xyloglucan and by catalysing trans-
glucosylation during restructuring of exist-
ing cell-wall-bound xyloglucan molecules.
The transgenic fruits had a more than

fourfold increase in XET activity associated
with reduced xyloglucan depolymerization
and reduced fruit softening, suggesting its
role in maintaining the structural integrity
of cell walls (Miedes et al. , 2010). Most fruit
species contain multiple genes for pectate
lyase (PL), an enzyme that hydrolyses the
unesterifi ed galacturonosyl linkages by a
E -elimination reaction. Although expression
of several PL isozymes increases during
fruit ripening, understanding their role in
pectinolysis and fruit texture changes is still
in its early stages. Introduction of an
antisense gene of a strawberry PL ( njjs25 )
under the control of the CaMV 35S
promoter inhibited the expression of PL ,
and the transgenic strawberry fruit
registered a decrease in ripening-associated
fi rmness. These transgenic fruit showed an
extended postharvest shelf-life, a reduction
in pectin solubility, decreased depoly-
merization of bound polyuronides and loss
of cell-cell adhesion in the transgenic fruits
(Jiménez-Bermúdez et al. , 2002; Santiago-
Doménech et al. , 2008). Transgenic
inhibition of Cel1 and Cel2 , two endo- E -1,4-
glucanases (EGases, cellulases) present in
many fruits, had little effect on fruit
softening (Lashbrook et al. , 1998; Brummell
et al. , 1999a). Downregulation of Cel1 and
Cel2 in strawberry fruits yielded similar
results with little infl uence on fruit
softening, but a slightly higher abundance
of the larger hemicellulosic polymers was
present in the fruit (Mercado et al. , 2010;
Pang et al. , 2010).
Expansins are a family of proteins that
induce extension in isolated plant cell
walls, expressed during fruit development
and ripening, and their roles in fruit
textural changes have been examined using
molecular genetic techniques (Choi et al. ,
2006). The antisense RNA inhibition of a
ripening-specifi c expansin, SlExp1 , caused
a reduction in polyuronide depoly-
merization without affecting the break-
down of other structurally important
hemicelluloses, and the transgenic fruit
retained a fi rmer texture than the wild-type
fruit (Rose et al. , 1997). The constitutive
expression of SlExp1 caused an opposite
phenotype, and the transgenic fruit was