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).