Cell-wall Metabolism and Softening during Ripening - Fruit Ripening: Physiology, Signalling and Genomics - page 53

developmental stage that corresponded to
when the mutant should normally ripen.
They demonstrated that PG2A does cause
depolymerization of polyuronides, but the
fruit did not signifi cantly soften. This
indicates that PG2A can fragment pectins
but that in the transgenic plants where
PG2A is suppressed there are other
enzymes that can also fragment pectin.
The results with overexpression of
PG2A in rin and suppression in Flavr Savr
further suggest that PG2A by itself may not
be enough to cause signifi cant alterations
in tomato fruit fi rmness; but before moving
on to a different topic, let us look at
another fruit. In peach, there are naturally
occurring varieties that produce fruit called
melting fl esh (MF) and non-melting fl esh
(NMF). Examination of NMF fruit found
that PGs were not expressed or were not
secreted into the cell wall (Callahan et al. ,
2004; Ghiani et al. , 2011). The NMF
varieties all had deletions of one form or
another in a genetic locus for a family of
PGs (Callahan et al. , 2004). In this
particular case, the correlation between MF
texture and the absence of PG was very
compelling. Moreover, what is also
important was that they found that fruit
fi rmness (compressibility) was not greatly
different between the MF and NMF
varieties (Ghiani et al. , 2011). These
authors concluded that cell turgor plays a
more important role in changes in fi rmness
but that PG activity is responsible for the
marked difference in fruit texture. To
restate, texture is defi ned as a sensory
attribute and is more diffi cult to measure
with instrumentation than fi rmness (Brook-
fi eld et al. , 2011).
E -Galactans are branch polymers on
rhamnogalacturonans, which are a major
component, in addition to polygalacturonic
acid, in the pectin matrix of the primary
cell wall (Carpita and Gibeaut, 1993).
Antisense suppression of E -galactosidase 4
reduced the amount of free galactose in
ripening fruit and increased fruit fi rmness
(Smith et al. , 2002). Another enzyme that
can affect pectin structure and frag-
mentation are pectin esterases (Rose et al. ,
2004). It is thought that polyuronides are

synthesized as a methyl or acetyl ester that
blocks the carboxyl acid group (Carpita
and Gibeaut, 1993; Rose et al. , 2004). After
the polyuronides are laid down in the cell
wall; the esterifi ed polyuronides are de-
esterifi ed by pectin esterases. The charged
carboxyl group can then interact with
calcium to form a stable egg-box-like
structure (Carpita and Gibeaut, 1993). It is
thought that de-esterifi cation of poly-
uronide is necessary before PGs can act on
this substrate (Tieman et al. , 1992).
Suppression of a pectin methyl esterase
(PME) in tomato fruit reduced the de-
esterifi cation of polyuronides and its de-
polymerization during ripening, but,
although an increase in soluble solids was
observed in the transgenic fruit, no change
in fi rmness or texture were noted (Tieman
et al. , 1992). A separate group suppressed a
different pectin esterase in tomato fruit and
also observed a signifi cant increase in
esterifi ed pectin but did not identify any
signifi cant differences in softening; how-
ever, unlike the earlier report on the
suppression of a PME, they did not observe
any difference in soluble solids (Hall et al. ,
1993).

4.3.2 Cellulose

Cellulose is another major component of
fruit cell walls, and it is the cellulose
microfi brils that provide the tensile
strength to the cell wall. Intuitively, it
makes sense that modifi cation of this
fraction of the cell wall would have
marked effects on softening. However, in
most fruits examined, the cellulose content
of the cell wall does not change much
during ripening (Maclachlan and Brady,
1994; Newman and Redgwell, 2002;
Brummell, 2006). Before discussing the
results for mutants with suppressed levels
of cellulases, it is worth noting that
analysis is complicated by the fact that we
do not know the true substrate for the plant
enzymes called cellulase. Cellulases are
defi ned by their ability to degrade and
reduce the viscosity of carboxymethyl-
cellulose (CMC) in an in vitro assay. CMC