Cell-wall Metabolism and Softening during Ripening - Fruit Ripening: Physiology, Signalling and Genomics - page 54

is an artifi cial water-soluble cellulose. For
example, purifi ed avocado cellulase readily
degrades CMC but when added to
microfi brils or crystalline cellulose, the
enzyme is incapable of signifi cantly
degrading these substrates (O'Donoghue et
al. , 1994). A similar result has been
obtained with other plant cellulases
(Urbanowicz et al. , 2007; Vicente et al. ,
2007). It has been suggested that the native
substrate for plant cellulases might be
xyloglucans (Hatfi eld and Nevins, 1986);
however, subsequent studies with purifi ed
avocado cellulase demonstrated that this
cellulase could not degrade purifi ed
xyloglucans (O'Donoghue et al. , 1994).
What is the substrate for cellulases? We
still do not know for certain; however,
incubation of avocado cellulase with cell
walls purifi ed from unripe avocado fruit
caused a loss in the cohesiveness of the
microfi brils (O'Donoghue et al. , 1994).
These authors concluded that avocado
cellulase does not cleave the xyloglucans
that tether the cellulose microfi brils but
rather cleaves cellulose at accessible sites
on the periphery of the microfi brils,
which causes a loss of integrity within
the fi bril structure and alters the binding
of associated cell-wall matrix poly-
saccharides, such as xyloglucans. In
avocado, which produces an inordinate
amount of cellulase, it was concluded that,
although cellulase might not depolymerize
crystalline cellulose microfi brils, cellulase
might modify the cellulose polymers
exposed to the surface of the microfi bril,
which would then affect the tethering and
organization of the microfi brils in the wall
(O'Donoghue et al. , 1994). Unfortunately,
transformation of avocado to suppress
cellulase is not currently possible.
Tomato fruit includes two cellulases
that are expressed during fruit ripening,
Cel1 and Cel2 (Lashbrook et al. , 1994).
When each gene - Cel1 (Lashbrook et al. ,
1998) and Cel2 (Brummell et al. , 1999a) -
was suppressed individually, there was no
signifi cant effect on fruit ripening or
softening. This result left open the
possibility that both genes needed to be
suppressed in order to have an effect on

softening. However, in a subsequent study
where both Cel1 and Cel2 were
suppressed simultaneously, no obvious
change in softening was reported, but
there was a clear effect on infection by
Botrytis cinerea (a common fungal
pathogen of fruit), which suggested to the
authors that these cellulases altered the
cell wall in a manner that made the fruit
more susceptible to infection (Flors et al. ,
2007). Interestingly, expression of Cel2 in
the non-ripening rin mutant, which does
not normally express Cel2, did cause the
fruit to partially soften but not ripen
(Flors et al. , 2007). What this suggests is
that Cel2 might affect tethering of the
cellulose microfi brils but that, in a normal
ripening process, there are other enzymes
or processes that have a similar effect on
tethering of microfi brils.
Before moving on to look at other
components of the cell wall, it is of interest
to examine research with strawberry fruit.
Suppression of PGs had a clear effect on
fruit fi rmness when measured by extrusion
of the pulp through an orifi ce during fruit
compression (Quesada et al. , 2009). The
measurement of fruit fi rmness for straw-
berry was done differently from that for
tomato fruit and might be better compared
with the decrease in viscosity of the tomato
paste prepared from transgenic tomato
with reduced PG activity and also the MF
characteristic of peach. Also of com-
parative interest is that suppression of a
cellulase (Cel1) in strawberry, like tomato,
had no signifi cant effect on fruit fi rmness
(Woolley et al. , 2001).
Whilst on the topic of untethering of
microfi brils, it is necessary to discuss work
on a protein called expansin. Expansins
were fi rst discovered in the stems of
elongating seedlings (McQueen-Mason et
al. , 1992; McQueen-Mason and Cosgrove,
1994) and have since been identifi ed in
many plant tissues (Rose et al. , 1997; Rose
et al. , 2000; Cosgrove et al. , 2002).
Expansins are an interesting class of
protein because although they have no
known in vitro enzyme activity, they
clearly play a role in loosening of the cell
wall during cell elongation (Cosgrove,