Softening is an important part of the ripening process in many fruit and changes in cell wall composition and associated enzyme activity are thought to play central roles in the softening process. In this study, research was focused upon the phase of growth in which the grape berry (Vitis vinifera L.) softens.

A procedure was developed for the isolation of cell walls from mesocarp tissue of grape berries. This procedure included the use of filtration and phenol treatment to remove vascular tissue and adventitious proteins of cytoplasmic origin from the wall preparations. Compositional analysis of cell walls isolated from mature Gordo and Ohanez berries showed substantial differences in polygalacturonan and cellulose abundance. Subsequent analyses of the monosaccharide, polysaccharide and protein composition of walls isolated from Gordo berries at several stages of development revealed a number of changes that occurred during development. These included an increase in water-soluble pectins, a decrease in galactan/arabinogalactan-I and an increase in hydroxyproline-rich proteins.

The changes in cell wall polysaccharide composition during berry ripening suggested that a limited number of enzymes might be involved in the softening process. In an attempt to determine the importance of particular enzymes in berry softening, the activities of several enzymes were monitored throughout berry development. Overall, b-galactosidase and a-galactosidase activities were the most readily detectable and were found to increase markedly during berry softening. b-Galactosidase may be the enzyme responsible for the loss of galactan from cell walls as berries soften. Pectin methylesterase activity remained relatively low during berry development, consistent with the observation that the degree of esterification of pectins remained approximately the same during berry softening. Little or no activity was detected for polygalacturonase, galactanase, cellulase or xyloglucanase in ripening berries.

To further investigate the possible roles of enzymes in grape berry development, cDNAs encoding a number of cell wall-modifying enzymes were isolated, either by screening a grape berry cDNA library or by the polymerase chain reaction. The cDNAs were used as probes to monitor steady-state mRNA levels of the respective genes in developing grape berries and other grapevine tissues. The detection of b-galactosidase mRNA was consistent with the presence of high levels of b-galactosidase activity and the decrease in galactan content in walls. Similarly, the presence of polygalacturonase mRNA was consistent with the increased solubility of pectic polysaccharides during ripening, although no polygalacturonase activity had been detected. The presence of xyloglucan endotransglycosylase mRNA also suggested that this enzyme plays some role in berry softening. Some pectin methylesterase mRNA was detected during berry development, consistent with the low levels of pectin methylesterase activity during berry ripening. Despite the pectin methylesterase activity, the degree of esterification of pectic polysaccharides remained constant during berry softening. In contrast, mRNA transcripts encoding cellulases were not detected in ripening berries, in agreement with the absence of detectable cellulase activity.

In summary, no gross changes in the cell wall composition occurred during berry development and no single enzyme could be assigned as the major cause for grape berry softening. This supports the general theory that fruit softening is a complex process involving relatively minor changes in different components of the cell wall that can subsequently have a large impact on the overall wall structure.