Endo-Polygalacturonases
Fruit ripening is accompanied by disassembly of several cell wall polymers,
including pectin and hemicellulose, which are primarily responsible for
ripening-associated changes in fruit texture. Extensive studies on ripeningassociated
pectin disassembly and the expression of the endo-polygalacturonase
(PG) gene family, suggested that tomato fruit texture could be modified by
transgenic modification of PG gene expression. The expression of both antisense
and sense constructs of the tomato fruit PG catalytic subunit (PG2) gene resulted
in greater than 95% reduction in PG activity (Sheehy et al
., 1988; Smith et al
.,
1998; 1990a). Fruit with reduced expression of PG were analyzed for alterations
in the expression of other cell wall hydrolases and none were detected (Smith et al., 1990b). Fruit with reduced PG activity provided the basis for testing the
significance of this PG during softening and ripening as well as the basis for the
commercial introduction of fresh and processed tomato fruit whose texture was
modified by this genetic modification. Analysis of cell wall polymers of these
fruit demonstrated that diminished PG expression contributed to reduced
depolymerization of the chelator-solubilized pectins and increased viscosity of
processed tomato products but did not reduce fruit softening (Taylor et al., 1991;
Carington et al., 1993; Fenwick et al., 1996; Brummell and Labavitch, 1997;
Porretta and Poli, 1997; Porretta et al., 1998).
The effect of antisense
suppression of a single fruit PG on fruit softening may be partially offset by
expression of other tomato PG genes in ripening fruit (Sitrit and Bennett, 1998).
Interestingly, transgenic plants with reduced expression of fruit PG did not
exhibit changes in leaf abscission, suggesting that PGs involved in abscission are
distinct from those that participate in fruit ripening (Tayloret al., 1991). The
tomato fruit PG gene has also been inactivated by transposition and stabilization
of a maize transposon,
DS, within the PG gene (Cooley and Yoder, 1998).
Suppression of the non-catalytic
β subunit of the PG1 isozyme complex in
transgenic tomato plants by expression of an antisense gene construct also
reduced pectin metabolism during fruit ripening (Watson et al., 1994).
Specifically, the reduced expression of the PG
β s1ubunit, a regulatory subunit,
reduced cell wall pectin solubilization and depolymerization, suggesting that the
dynamics of pectin associations and structure in the cell wall are determined by
several factors, perhaps some acting cooperatively (Watson et al., 1994).
Several other fruit characteristrics have been measured in tomato fruit with
suppressed PG gene expression. Transgenic tomato fruit were evaluated for
sensory characteristics and their color and flavor outperformed a similar variety
that was heterozygous for the
rin (ripening inhibited) locus, a variety that had
been bred for long shelf life (Sozzi Quiroga and Fraschina, 1997). The tomatine
content of transgenic fruit was unaffected by antisense suppression of PG (Furui
et al., 1998). Furthermore, PG antisense fruit generally had improved integrity
and were less susceptible to cracking and pathogen attack specifically at the
over-ripe stage (Krameret al., 1992; Had field and Bennett, 1998). However, the
susceptibility of PG suppressed transgenic tomato fruit to
Colletotrichum gloeosporioides was not measurably different than in wild-type fruit (Cooper
et al., 1998).
