Overexpression of the persimmon abscisic acid β‐glucosidase gene (DkBG1) alters fruit ripening in transgenic tomato

β‐Glucosidases (BG) are present in many plant tissues. Among these, abscisic acid (ABA) β‐glucosidases are thought to take part in the adjustment of cellular ABA levels, however the role of ABA‐BG in fruits is still unclear. In this study, through RNA‐seq analysis of persimmon fruit, 10 full‐length DkBG genes were isolated and were all found to be expressed. In particular, DkBG1 was highly expressed in persimmon fruits with a maximum expression 95 days after full bloom (DAFD). We verified that, in vitro, DkBG1 protein can hydrolyze ABA‐glucose ester (ABA‐GE) to release free ABA. Compared with wild‐type, tomato plants that overexpressed DkBG1 significantly upregulated the expression of ABA receptor PYL3/7 genes and showed typical symptoms of ABA hypersensitivity in fruits. DkBG1 overexpression (DkBG1‐OE) accelerated fruit ripening onset by 3–4 days by increasing ABA levels at the pre‐breaker stage and induced early ethylene release compared with wild‐type fruits. DkBG1‐OE altered the expression of ripening regulator NON‐RIPENING (NOR) and its target genes; this in turn altered fruit quality traits such as coloration. Our results demonstrated that DkBG1 plays an important role in fruit ripening and quality by adjusting ABA levels via hydrolysis of ABA‐GE.     

Regulation of the expression of lipoxygenase genes in <Emphasis Type="Italic">Prunus persica</Emphasis> fruit ripening

Three genes of the lipoxygenase (LOX) family in peach (Prunus persica var. compressa cv. Ruipan 4) were cloned, and their expression patterns during fruit ripening were analyzed using real-time quantitative PCR. All of the three peach LOX genes had been expressed during fruit ripening; however, their expression patterns were significantly different. During the normal ripening of peach fruits, the expression levels of PpLox1, PpLox2 and PpLox3 increased in varying degrees accompanying upsurge of ethylene evolution. After treated by methyl jasmonic acid (MeJA), the peak of ethylene releasing occurred in advance, and the declining rate of fruit hardness was accelerated, the expression level of the three peach LOX genes in fruits markedly enhanced at the early stage of storage, but significantly decreased at the late storage stage. So, it could be suggested that all three LOXs relate to fruit ripening; however, their functions might be different. PpLox1 expression increase along with the upsurge of ethylene evolution in both control and MeJA-treated peach fruits suggested that PpLox1 probably played a major role in the peach fruit ripening. Expression peak of PpLox2 appeared at the 1 DAH (days after harvest) in both control and MeJA-treated peach fruits, while obvious changes in ethylene evolution and fruit hardness was not observed, which suggested that the rise of PpLox2 expression can be induced by certain stimulation related to ripening, such as harvesting stress and MeJA treatment. The expression of PpLox3 kept a lower level in the natural ripening fruits, whereas raced up at the early stage of storage in the fruits treated with MeJA, which indicated that PpLox3 was expressed inductively and had minor roles during the normal ripening of peach fruits, but when encountered with external stimulation, its expression level would rapidly enhance and accelerate the ripening of peach fruit.     

ClSnRK2.3 negatively regulates watermelon fruit ripening and sugar accumulation

Watermelon(Citrullus lanatus) as non-climacteric fruit is domesticated from the ancestors with inedible fruits. We previously revealed that the abscisic acid(ABA) signaling pathway gene ClSnRK2.3 might infuence watermelon fruit ripening. However,the molecular mechanisms are unclear. Here,we found that the selective variation of ClSnRK2.3 resulted in lower promoter activity and gene expression level in cultivated watermelons than ancestors, which indicated ClSnRK2.3 might be a negative regulator ...     

Suppressing ABA uridine diphosphate glucosyltransferase (SlUGT75C1) alters fruit ripening and the stress response in tomato

Abscisic acid (ABA) glucose conjugation mediated by uridine diphosphate glucosyltransferases (UGTs) is an important pathway in regulating ABA homeostasis. In the present study, we investigated three tomato SlUGTs that are highly expressed in fruit during ripening, and these SlUGTs were localized to the cytoplasm and cell nucleus. Among these three UGTs, SlUGT75C1 catalyzes the glucosylation of both ABA and IAA in vitro; SlUGT76E1 can only catalyze the conjugation of ABA; and SlUGT73C4 cannot glycosylate either ABA or IAA. Therefore, SlUGT75C1 was selected for further investigation. SlUGT75C1 RNA interference significantly up‐regulated the expression level of SlCYP707A2, which encodes an ABA 8′‐hydroxylase but did not affect the expression of SlNCED1, which encodes a key enzyme in ABA biosynthesis. Suppression of SlUGT75C1 significantly accelerated fruit ripening by enhancing ABA levels and promoting the early release of ethylene. SlUGT75C1‐RNAi altered the expression of fruit ripening genes (genes involved in ethylene release and cell wall catabolism). SlUGT75C1‐RNAi seeds showed delayed germination and root growth compared with wild‐type as well as increased sensitivity to exogenous ABA. SlUGT75C1‐RNAi plants were also more resistant to drought stress. These results demonstrated that SlUGT75C1 plays a crucial role in ABA‐mediated fruit ripening, seed germination, and drought responses in tomato.     

Changes in the levels of abscisic acid and its metabolites resulting from chilling of tomato fruits

The 6,6,6-[²H]-analogues of abscisic acid (ABA), phaseic (PA) and dihydrophaseic (DPA) acids were used in GC-MS-SIM determination of free and total alkali hydrolyzable ABA, PA and DPA in the pericarp of tomato (Lycopersicon esculentum L. cv. Pik Red) fruit. Determinations were made on breaker-stage fruit stored 1, 2, 3 or 4 weeks at 2.5°C or at 10°C, and after subsequent ripening for 1 week in darkness at 20°C. Two-fold increases in levels of ABA occurred after storage at low temperatures with a slightly but significantly greater increase in ABA level occurring with 2.5°C storage. These increases in ABA levels were not associated with the appearance of damage symptoms that occurred with storage at the chilling temperature (2.5°C). Differences in ABA metabolism were found resulting from storage at the two temperatures, 2.5 or 10°C. Significantly greater DPA levels were found after 10°C storage than after 2.5°C storage (2 weeks). Levels of ABA ester-conjugates increased with 20°C ripening only after 10°C storage while free ABA levels decreased after both storage temperature conditions. Levels of DPA conjugates also increased only after 20°C ripening following 10°C storage. A longer period of storage resulted in decreases of free DPA levels after 10°C storage but increased DPA levels were found after 2.5°C storage.Abbreviations ABA abscisic acid - PA phaseic acid - DPA dihydrophaseic acid - GC-MS-SIM gas chromatography-mass spectrometry-selected ion monitoring - HPLC high pressure liquid chromatography - fw. fresh weight author for correspondence     

Abscisic acid plays an important role in the regulation of strawberry fruit ripening

The plant hormone abscisic acid (ABA) has been suggested to play a role in fruit development, but supporting genetic evidence has been lacking. Here, we report that ABA promotes strawberry (Fragaria ananassa) fruit ripening. Using a newly established Tobacco rattle virus-induced gene silencing technique in strawberry fruit, the expression of a 9-cis-epoxycarotenoid dioxygenase gene (FaNCED1), which is key to ABA biosynthesis, was down-regulated, resulting in a significant decrease in ABA levels and uncolored fruits. Interestingly, a similar uncolored phenotype was observed in the transgenic RNA interference (RNAi) fruits, in which the expression of a putative ABA receptor gene encoding the magnesium chelatase H subunit (FaCHLH/ABAR) was down-regulated by virus-induced gene silencing. More importantly, the uncolored phenotype of the FaNCED1-down-regulated RNAi fruits could be rescued by exogenous ABA, but the ABA treatment could not reverse the uncolored phenotype of the FaCHLH/ABAR-down-regulated RNAi fruits. We observed that down-regulation of the FaCHLH/ABAR gene in the RNAi fruit altered both ABA levels and sugar content as well as a set of ABA- and/or sugar-responsive genes. Additionally, we showed that exogenous sugars, particularly sucrose, can significantly promote ripening while stimulating ABA accumulation. These data provide evidence that ABA is a signal molecule that promotes strawberry ripening and that the putative ABA receptor, FaCHLH/ABAR, is a positive regulator of ripening in response to ABA.     

Effect of Abscisic Acid on Banana Fruit Ripening in Relation to the Role of Ethylene

Abstract The role of abscisic acid (ABA) in banana fruit ripening was examined with the ethylene binding inhibitor, 1-methylcyclopropene (1-MCP). ABA (0, 10⁻⁵, 10⁻⁴, or 10⁻³ mol/L) was applied by vacuum infiltration into fruit. 1-MCP (1 μL/L) was applied by injecting a measured volume of stock gas into sealed glass jars containing fruit. Fruit ripening, as judged by ethylene evolution and respiration associated with color change and softening, was accelerated by 10⁻⁴ or 10⁻³ mol/L ABA. ABA at 10⁻⁵ mol/L had no effect. The acceleration of ripening by ABA was greater at 10⁻³ mol/L than at 10⁻⁴ mol/L. ABA-induced acceleration of banana fruit ripening was not observed in 1-MCP treated fruit, especially when ABA was applied after exposure to 1-MCP. Thus, ABA's promotion of ripening in intact banana fruit is at least partially mediated by ethylene. Exposure of ABA-treated fruit to 0.1 μL/L ethylene for 24 h resulted in increased ethylene production and respiration, and associated skin color change and fruit softening. Control fruit (no ABA) was unresponsive to similar ethylene treatments. The data suggest that ABA facilitates initiation and progress in the sequence of ethylene-mediated ripening events, possibly by enhancing the sensitivity to ethylene. Received 29 January 1999; accepted 16 January 2000     

Cloning of 9-cis-epoxycarotenoid dioxygenase (NCED) gene and the role of ABA on fruit ripening

In order to understand more details about the role of abscisic acid (ABA) in fruit ripening and senescence, six 740 bp cDNAs (LeNCED1, LeNCED2, PpNCED1, VVNCED1, DKNCED1 and CMNCED1) which encode 9-cis-epoxycarotenoid dioxygenase (NCED) as a key enzyme in ABA biosynthesis, were cloned from fruits of tomato, peach, grape, persimmon and melon using an RT-PCR approach. A Blast homology search revealed a similarity of amino acid 85.76% between the NCEDs. A relationship between ABA and ethylene during ripening was also investigated. At the mature green stage, exogenous ABA treatment increased ABA content in flesh, and promoting ethylene synthesis and fruit ripening, while treatment with nordihydroguaiaretic acid (NDGA), inhibited them, delayed fruit ripening and softening. However, ABA inhibited the ethylene synthesis obviously while NDGA promoted them when treated the immature fruit with these chemicals. At the breaker, NDGA treatment cannot block ABA accumulation and ethylene synthesis. Based on the results obtained in this study, it was concluded that ABA plays different role in ethylene synthesis system in different stages of tomato fruit ripening.Key words: tomato, NCED gene, ABA, ethylene, fruit ripening, peach, grape, persimmon, melon     

Abscisic acid levels in tomato ovaries are regulated by <Emphasis Type="Italic">LeNCED1</Emphasis> and <Emphasis Type="Italic">SlCYP707A1</Emphasis>

Although the hormones, gibberellin and auxin, are known to play a role in the initiation of fruits, no such function has yet been demonstrated for abscisic acid (ABA). However, ABA signaling and ABA responses are high in tomato (Solanum lycopersicum L.) ovaries before pollination and decrease thereafter (Vriezen et al. in New Phytol 177:60–76, 2008). As a first step to understanding the role of ABA in ovary development and fruit set in tomato, we analyzed ABA content and the expression of genes involved in its metabolism in relation to pollination. We show that ABA levels are relatively high in mature ovaries and decrease directly after pollination, while an increase in the ABA metabolite dihydrophaseic acid was measured. An important regulator of ABA biosynthesis in tomato is 9-cis-epoxy-carotenoid dioxygenase (LeNCED1), whose mRNA level in ovaries is reduced after pollination. The increased catabolism is likely caused by strong induction of one of four newly identified putative (+)ABA 8′-hydroxylase genes. This gene was named SlCYP707A1 and is expressed specifically in ovules and placenta. Transgenic plants, overexpressing SlCYP707A1, have reduced ABA levels and exhibit ABA-deficient phenotypes suggesting that this gene encodes a functional ABA 8′-hydroxylase. Gibberellin and auxin application have different effects on the LeNCED1 and SlCYP707A1 gene expression. The crosstalk between auxins, gibberellins and ABA during fruit set is discussed.