Regulation Of Citrus Fruit Acidity By Genes Related To Citrate Metabolism And Transportation

Fruit acidity is one of the most important factors of fruit quality, and is regulated by the metabolism and transportation of organic acids in fruit cells. The main organic acid in citrus fruit is citrate. Gaocheng (Citrus sp.), Satsuma mandarin (Citrus unshiu Marc.) and Ponkan (Citrus reticulata Blanco cv. Ponkan) fruit were used as materials to investigate the expression of genes related to citrate metabolism and transportation by RNA-Seq, the function of genes related to citrate transport were further analyzed. This study was to reveal the mechanism of fruit acidity regulated by genetic factor, gardening practices and postharvest treatment in citrus fruit. The main results are as follows:1. Three transport proteins related to citrate degradation were identified. Gaocheng and Satsuma mandarin were selected for study and the degradation of citrate was deduced to be the main cause of the difference in acidity in fully mature fruits. RNA-Seq analysis was carried out on Gaocheng and Satsuma mandarin fruit of different developmental stages and the results indicated that citrate degradation occurred mainly through the glutamine pathway, catalyzed by CitAco3, CitGS2and CitGDUl, and also three transport-related genes, CitCHX, CitAL-MT and CitDIC, were shown to be associated with citrate degradation. In postharvest Gaocheng fruit, the expressions of these three transport-related genes were induced under hot air treatment, accompanied by a reduction of7%-9%in total acid degradation. Transient expression of CitCHX, CitAL-MT and CitDIC in tobacco leaves was performed, and the citrate content was decreased to24%,31%and27%of the control, respectively, and the activity of aconitase were significantly increased compared to the control. It can be concluded that the transport proteins were important factors to promote the citrate degradation pathway, but the mechanism needs further research.2. The mechanism of citrate metabolism in Ponkan fruit during delayed harvest was investigated. Compared to the open field fruit, the fruit cultivated in greenhouse showed lower titratable acid (TA) content, and the solids to acids ratio was increased by6%-19%, which indicated better flavor in greenhouse fruit. The citrate content was higher in open field fruit, this was regulated by CitPEPCs and CitCSs, which was significantly increased in open field fruit compared to greenhouse fruit. However, CitAco3and CitGAD4, which were the key genes of gamma-aminobutyric acid (GABA) shunt, were also up regulated, indicating that the GABA shunt was the main citrate degradation pathway under stress in Ponkan fruit.3. The impacts of low temperature and water stress on citrate metabolism in Ponkan fruit during delayed harvest were investigated. The citrate contents in low temperature-treated fruit were significantly increased, by1.4-1.9folds, compared to the control; it showed no significant difference in fruit with water stress treatment in the early30days, but was significantly increased, by about1.5folds, in the following30days compared to the control fruit. In addition, the expressions of CitPEPCs, CitCSs, CitAco3and CitGAD4were significantly induced by low temperature treatment, but showed no significant difference in water stress compared to the control fruit. The cultivation time for delayed harvest is generally maintained for1-2months, thus, low temperature may be the main factor influencing citrate metabolism during delayed harvest in open field fruit.In summary, two steps were proposed for citrate degradation in citrus fruits under different conditions. Firstly, the citrate is transported out of the vacuole via CitCHX, CitAL-MT and CitDIC. The exported citrate is then available to be metabolized through glutamine pathway during fruit development or GABA shunt under low temperature stress.