# Molecular and Hormonal Mechanisms Regulating Fleshy Fruit Ripening
## 2 Main Physiological and Biochemical Changes Are Regulated by Ripening Genes
### Fruit Softening  
It is generally accepted that fruit softening during ripening is, at least partly, due to cell wall changes catalysed by wall-modifying enzymes. Investigations with many fruits have shown that there are at least 10 different types of cell wall-modifying enzymes that are expressed during ripening [56], and these are believed to collectively contribute to the change in texture, leading to alterations in the structure and hydration of cell wall polymers, leading to softening. These include the pectin-modifying enzymes: pectinesterase (PE), pectate lyase (PL), polygalacturonase (PG) and \(\beta\)-galactosidase (\(\beta\)-GAL), and the hemicellulose/cellulose-modifying enzymes: 1,4-\(\beta\)-glucanase, xyloglucan transglycosylation-hydrolase (XTH) and expansin (EXP). It is clear that several of these enzymes have specific actions affecting cell wall structure that contribute to texture changes, although there is no single enzyme that makes a major contribution to softening [56; 57]. Expansins allow cellulose microfibrils to slide over one another without covalent modification, and PG reduces the chain length of pectins, which does have some effect because it decreases the viscosity of tomato extracts [58]. Softening of intact fruit is largely unaffected, although some small increase in firmness has been detected in antisense-PG tomatoes [59]. Furthermore, an analysis of _PG/EXP1_ knockdown tomato fruit showed that they were firmer, had higher total soluble solids, denser cell walls and thicker cuitics than fruit of other genotypes [60], and also showed reduced skin cracking [60]. More recently, RNAi knockdown and CRISPR/Cas9 knockout experiments showed that eliminating _PL_ produced firmer tomatoes, although softening was not abolished [61]. The question of the regulation of cell wall metabolising enzymes was addressed early on and there are differences between fruits. _PG_ transcripts in tomato are very sensitive to low levels of ethylene [62]. The situation varies in different fruits, however, and in melon, which expresses three _PG_ genes, accumulation of _CmPG1_ transcripts is dependent on ethylene, whereas regulation of _CmPG2_ is ethylene-independent and expression of _CmPG3_ is regulated by both ethylene-dependent and ethylene-independent factors [63; 64]. Attempts to delay fruit softening and improve fruit quality by manipulating stress responses during storage have provided new insights into the role of a group of cell wall-modifying enzymes in causing texture change. Deterioration of peaches is delayed if they are stored at 0 \({}^{\circ}\)C. Upon removal from storage, however, they fail to undergo normal softening, and consequently, the texture is disliked by consumers due to the failure to produce the normal levels of a group of cell wall-modifying enzymes. A low-temperature conditioning (LTC) treatment at 8 \({}^{\circ}\)C, however, prior to 0 \({}^{\circ}\)C storage, results in much improved softening upon return to room temperature after 0 \({}^{\circ}\)C storage. The explanation for this is that after transfer to room temperature, transcripts for _PG, PL, PE, \(\beta\)-1,4-endoglucanase, xylosidase (XYL), EXP1, GAL_ and _XTH_ are all substantially increased, provided the peaches were acclimated at 8 \({}^{\circ}\)C prior to 0 \({}^{\circ}\)C storage [65]. Similarly, in persimmon fruit, many cultivars are asringent, due to the high soluble tannins content, which consumers find unacceptable. There are several postharvest treatment methods for removing these soluble tannins. One successful approach is to store the fruits in a 95% CO\({}_{2}\) + 1% O\({}_{2}\) atmosphere. This postharvest high CO\({}_{2}\)/anaerobic storage leads to acetaldehyde production, which precipitates the soluble tannins, reducing or eliminating the astringency. Unfortunately, however, persimmon treated in this way tend to soften too much.