The overall goal of the research was to understand the processes involved in the development of woolliness in peaches and nectarines. Four specific hypotheses were proposed and in the course of the research evidence was gathered t support two of them and to not support two others. The hypotheses and a summary of the evidence are outlined below. 1. That woolliness arises from an imbalance between the activities of the cell wall pectin degrading enzymes. Using 'Flavortop' nectarines and 'Hermoza' peaches as model systems, storage regimes were manipulated to induce or prevent woolliness. The expression (mRNA abundance), protein content (Western blotting), and activity of polygalacturonase (PG) and pectin esterase (PE) were followed. Expression of the enzymes was not different, but activity and the ratio between PG and PE activities were quite different in fruits developing woolliness or ripening normally. This was also examined by looking at the substrate, the pectin moiety of the cell wall, and i woolly fruit there were more high molecular weight pectins with regions of non-methylated galacturonic acid residues. Taking an in vitro approach it was found a) that PE activity was stable at 0oC while PG activity decreased; b) incubating the calcium pectate fraction of the cell wall with PE extracted from peaches caused the polymers to form a gel characteristic of the visual woolly symptoms in peaches. 2. That continued cell wall synthesis occurs during storage and contributes to structural changes i cell walls and improper dissolution and softening after storage. We tried to adapt our technique of adding 13C-glucose to fruit discs, which was used successfully to follow cell wall synthesis during tomato ripening. However, the difference in sugar content between the two fruits (4% in tomato and 12% in peach) meant that the 13C-glucose was much more diluted within the general metabolite pool. We were unable to see any cell wall synthesis which meant that either the dilution factor was too great, or that synthesis was not occurring. 3. That controlled atmosphere (CA) prevents woolliness by lowering all enzyme activities. CA was found to greatly reduce mRNA abundance of the cell wall enzymes compared to regular air storage. However, their synthesis and activity recovered during ripening after CA storage and did not after regular air storage. Therefore, CA prevented the inhibition of enzyme activation found in regular air storage. 4. That changes in cell wall turgor and membrane function are important events in the development of woolliness. Using a micro pressure probe, turgor was measured in cells of individual 'O'Henry' and 'CalRed' peaches which were woolly or healthy. The relationship between firmness and turgor was the same in both fruit conditions. These data indicate that the development and expression of woolliness are not associated with differences in membrane function, at least with regard to the factors that determine cell turgor pressure. In addition, during the period of the grant additional areas were explored. Encoglucanase, and enzyme metabolizing hemicellulose, was found to be highly expressed air stored, but not in unstored or CA stored fruit. Activity gels showed higher activity in air stored fruit as well. This is the first indication that other components of the cell wall may be involved in woolliness. The role of ethylene in woolliness development was also investigated at it was found a) that woolly fruits had decreased ability to produce ethylene, b) storing fruits in the presence of ethylene delayed the appearance of woolliness. This latter finding has implication for an inexpensive strategy for storing peaches and nectarines.
Cell wall biosynthesis and the molecular mechanism of plant enlargement
