Controlling the rate of softening to extend shelf life was a key target for researchers engineering genetically modified (GM) tomatoes in the 1990s, but only modest improvements were achieved. Hybrids grown nowadays contain 'non-ripening mutations' that slow ripening and improve shelf life, but adversely affect flavor and color. We report substantial, targeted control of tomato softening, without affecting other aspects of ripening, by silencing a gene encoding a pectate lyase.

Polygalacturonase activity is not detectable in mature green tomato fruits but appears as fruits begin to change colour and continues to increase during the ripening period. There is a sequential appearance of two isoenzymes, polygalacturonase 1 and 2, during ripening. These isoenzymes have been purified and their properties compared. Polygalacturonase 1 has a Mr of 100000, is 50% inactivated at 78° C and has a density of 1.343 g cm−3 in caesium chloride. Polygalacturonase 2 has a Mr of 42000, is 50% inactivated at 57° C and has a density of 1. 300 g cm−3 in caesium chloride. Fruits from isogenic lines homozygous for the ‘Neverripe’ (Nr) mutation do not ripen normally and contain reduced amounts of polygalacturonase. Only polygalacturonase 1 is produced in Nr fruit. Tomatoes from isogenic lines homozygous for the ‘ripening inhibitor’ (rin) mutation do not ripen normally and produce very little detectable polygalacturonase. Although polygalacturonases 1 and 2 have different properties they both give rise to a single polypeptide on electrophoresis in polyacrylamide gels in the presence of sodium dodecylsulphate (Mr= 46000). A comparison of the major fragments produced by limited proteolysis of polygalacturonase 1 and 2 with chymotrypsin suggests that the polypeptides from the two isoenzymes are similar. The same conclusion was reached from a comparison of polygalacturonase 1 and 2 by radioimmunoassay, using antibody prepared against polygalacturonase 2 and 125I‐labelled polygalacturonase 2. The results from radioimmunoassay of extracts from green and ripening fruits suggest that the increase in polygalacturonase activity during ripening is due to net synthesis of protein.

The Colorless non-ripening (Cnr) mutation in tomato (Solanum lycopersicum) results in mature fruits with colorless pericarp tissue showing an excessive loss of cell adhesion (A.J. Thompson, M. Tor, C.S. Barry, J. Vrebalov, C. Orfila, M.C. Jarvis, J.J. Giovannoni, D. Grierson, G.B. Seymour [1999] Plant Physiol 120: 383-390). This pleiotropic mutation is an important tool for investigating the biochemical and molecular basis of cell separation during ripening. This study reports on the changes in enzyme activity associated with cell wall disassembly in Cnr and the effect of the mutation on the program of ripening-related gene expression. Real-time PCR and biochemical analysis demonstrated that the expression and activity of a range of cell wall-degrading enzymes was altered in Cnr during both development and ripening. These enzymes included polygalacturonase, pectinesterase (PE), galactanase, and xyloglucan endotransglycosylase. In the case of PE, the protein product of the ripening-related isoform PE2 was not detected in the mutant. In contrast with wild type, Cnr fruits were rich in basic chitinase and peroxidase activity. A microarray and differential screen were used to profile the pattern of gene expression in wild-type and Cnr fruits. They revealed a picture of the gene expression in the mutant that was largely consistent with the real-time PCR and biochemical experiments. Additionally, these experiments demonstrated that the Cnr mutation had a profound effect on many aspects of ripening-related gene expression. This included a severe reduction in the expression of ripening-related genes in mature fruits and indications of premature expression of some of these genes in immature fruits. The program of gene expression in Cnr resembles to some degree that found in dehiscence or abscission zones. We speculate that there is a link between events controlling cell separation in tomato, a fleshy fruit, and those involved in the formation of dehiscence zones in dry fruits.Colorless non-ripening (Cnr) is a pleiotropic dominant mutation of tomato (Solanum lycopersicum) that results in fruits with a white pericarp displaying much reduced cell-to-cell adhesion (Thompson et al., 1999;Fraser et al., 2001). The Cnr locus has been mapped to the middle of the long arm of chromosome 2 and is currently the subject of a map-based cloning exercise (Tor et al., 2002). We have recently isolated and sequenced a region of tomato chromosome 2 that cosegregates with the Cnr locus and are testing a candidate gene at this locus (K. Manning, J.J. Giovannoni, and G.B. Seymour, unpublished data).The loss of cell adhesion in Cnr appears to be due principally to modifications in cell wall structure. Sections of Cnr pericarp tissue show obvious changes in comparison with wild-type fruits, including larger intercellular spaces and thinner cell walls in ripe fruits (Orfila et al., 2001). Mechanical tests on pericarp tissue have revealed that the force required for cell wall failure is greater in Cnr, while tests on cell wall preparations showed ...

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