5-Azacytidine (AZ) is a DNA methylation inhibitor that has recently demonstrated potential in regulating fruit quality through exogenous application. In this study, we treated mandarin fruits for 4-day storage. Noteworthy were the induced degreening and the enhanced citrus aroma of fruits under AZ treatment, involving the promotion of chlorophyll degradation, carotenoid biosynthesis, and limonene biosynthesis. Key genes associated with these processes exhibited expression level increases of up to 123.8 times. Additionally, AZ treatment activated defense-related enzymes and altered phenylpropanoid carbon allocation towards lignin biosynthesis instead of flavonoid biosynthesis. The expression levels of lignin biosynthesis-related genes increased by nearly 100 times, leading to fortified lignin that is crucial for citrus defense against Penicillium italicum. Currently, the underlying mechanisms of such intense AZ-induced changes in gene expressions remain unclear and further research could help establish AZ treatment as a viable strategy for citrus preservation.

Seed longevity, the capacity to remain alive during dry storage, is pivotal to germination performance and is essential for preserving genetic diversity. It is acquired during late maturation concomitantly with seed degreening and the de-differentiation of chloroplasts into colorless, non-photosynthetic plastids, called eoplasts. As chlorophyll retention leads to poor seed performance upon sowing, these processes are important for seed vigor. However, how these processes are regulated and connected to the acquisition of seed longevity remains poorly understood. Here, we show that such a role is at least provided by ABSCISIC ACID INSENSITIVE 4 (ABI4) in the legume Medicago truncatula. Mature seeds of Mtabi4 mutants contained more chlorophyll than wild-type seeds and exhibited a 75% reduction in longevity and reduced dormancy. MtABI4 was necessary to stimulate eoplast formation, as evidenced by the significant delay in the dismantlement of photosystem II during the maturation of mutant seeds. Mtabi4 seeds also exhibited transcriptional deregulation of genes associated with retrograde signaling and transcriptional control of plastid-encoded genes. Longevity was restored when Mtabi4 seeds developed in darkness, suggesting that the shutdown of photosynthesis during maturation, rather than chlorophyll degradation per se, is a requisite for the acquisition of longevity. Indeed, the shelf life of stay green mutant seeds that retained chlorophyll was not affected. Thus, ABI4 plays a role in coordinating the dismantlement of chloroplasts during seed development to avoid damage that compromises the acquisition of seed longevity. Analysis of Mtabi4 Mtabi5 double mutants showed synergistic effects on chlorophyll retention and longevity, suggesting that they act via parallel pathways.

Peel degreening is the most conspicuous aspect of fruit ripening in many citrus fruits because of its importance for marketability. In this study, peel degreening in response to propylene (an ethylene analog) and at varying storage temperatures was characterized in Satsuma mandarin (Citrus unshiu Marc.) fruit. Propylene treatment triggered rapid peel degreening (within 4-6 days), indicated by an increase in the citrus color index (CCI) and chlorophyll loss. Peel degreening was also observed in fruit at 10°C and 15°C after 28-42 days, with gradual CCI increase and chlorophyll reduction. However, fruit at 5°C, 20°C, and 25°C remained green, and no substantial changes in peel CCI and chlorophyll content were recorded during the 42-day storage duration. The transcriptomes of peels of fruit treated with propylene for 4 days and those stored at varying temperatures for 28 days were then analyzed by RNA-Seq. We identified three categories of differentially expressed genes that were regulated by (i) propylene (and by analogy, ethylene) alone, (ii) low temperature (5°C, 10°C, or 15°C vs. 25°C) alone, and (iii) either propylene or low temperature. Gene-encoding proteins associated with chlorophyll degradation (such as CuSGR1, CuNOL, CuACD2, CuCAB2, and CuLHCB2) and a transcription factor (CuERF114) were differentially expressed by propylene or low temperature. To further examine temperature-induced pathways, we also monitored gene expression during on-tree fruit maturation vs. postharvest. The onset of on-tree peel degreening coincided with autumnal drops in field temperatures, and it was accompanied by differential expression of low temperature-regulated genes. On the contrary, genes that were exclusively regulated by propylene (such as CuCOPT1 and CuPOX-A2) displayed insignificant expression changes during on-tree peel degreening. These findings indicate that low temperatures could be involved in the fruit ripening-related peel degreening independently of ethylene.

The effects of low temperature (LT) on chlorophyll (Chl) degradation in peel of apple fruit during ripening were investigated. Apples collected at commercial maturity were stored at 4 ± 0.5°C. Our data indicated that LT treatment reduced respiration rate and ethylene production and slowed down softening of apple fruit during ripening. The LT treatment delayed increase in L*, a*, and b* values and decrease in Chl content compared with controls. The LT treatment reduced hydrogen peroxide (H2 O2 ) and malondialdehyde (MDA) contents and decelerated superoxide anion (O2 ·- ) production rate in chloroplast of peel compared with controls during ripening. The LT treatment differentially reduced activities of pheophytin pheophorbide hydrolase (PPH), Mg-dechelatase (MDcase), chlorophyll-degrading peroxidase (Chl-POX), and Chl oxidase, while enhanced SOD activity in chloroplast of peel during ripening. Expression levels of MdHCARa, MdNYC1, MdNYC3, MdNYE1, MdRCCR2, MdPPH1, MdPAO6, MdPAO8, and MdNOL2 in peel were differentially reduced by LT treatment during ripening. Our results indicated that LT treatment might delay Chl degradation through inhibiting PAO pathway and Chl oxidation during ripening of apple fruit. PRACTICAL APPLICATIONS: The LT is a common practice used to extend storage life of apple fruit. Degreening caused by Chl degradation is an integral part of fruit ripening, and elucidating its mechanism is an important subject for fruit quality maintenance. Our data indicated that LT delayed degreening of apple fruit by inhibiting PAO pathway and Chl oxidation during ripening. These results will provide useful information for clarifying molecular mechanisms of LT in regulation of degreening and also for quality maintenance of apple fruit.

To reveal genetic factors or pathways involved in the pod degreening, we performed transcriptome and metabolome analyses using a yellow pod cultivar of the common bean \"golden hook\" ecotype and its green pod mutants yielded via gamma radiation. Transcriptional profiling showed that expression levels of red chlorophyll catabolite reductase (RCCR, Phvul.008G280300) involved in chlorophyll degradation was strongly enhanced at an early stage (2 cm long) in wild type but not in green pod mutants. The expression levels of genes involved in cellulose synthesis was inhibited by the pod degreening. Metabolomic profiling showed that the content of most flavonoid, flavones, and isoflavonoid was decreased during pod development, but the content of afzelechin, taxifolin, dihydrokaempferol, and cyanidin 3-O-rutinoside was remarkably increased in both wild type and green pod mutant. This study revealed that the pod degreening of the golden hook resulting from chlorophyll degradation could trigger changes in cellulose and flavonoids biosynthesis pathway, offering this cultivar a special color appearance and good flavor.

Chlorophyll (Chl) degradation is one of the most obvious signs of leaf senescence and fruit ripening. Stay-green (SGR) homologs that can remove magnesium from Chl a are the most important components in Chl degradation pathway in green plants. SGR homologs are not only universally involved in Chl breakdown during the senescence of green organs, but also play crucial roles in other organs during plant growth and development, such as fruit mature and nodule development. In this review, we focus on the diverse functions of SGR homologs in plant growth and development. A better understanding of SGR would be helpful for providing a theoretical basis for further illustrating the regulatory mechanism of SGR homologs.

Degreening is widely used in citrus fruit to extend the market season for economic gains and increase the consumer acceptance. Elevated CO2 was exogenously applied to Satsuma mandarins (Citrus unshiu Marc.) for degreening and its effect on pigment metabolism was investigated. The results revealed 15% CO2 treatment accelerated the citrus fruit peel color change along with the chlorophyll degradation, β-cryptoxanthin and flavonoids accumulation. The expression of CitSGR, CitNYC, CitChlase, CitPPH, CitPAO and CitRCCR genes involved in chlorophyll metabolism and a set of genes involved in producing β, β-xanthophylls were up-regulated by elevated CO2. For flavonoid metabolism, the up-regulated expressions of CitPAL, CitCHS and CitCHI partly explained the increased total flavonoids content. These results showed that 15% CO2 treatment improved the visual appearance of citrus fruits due to its impact on pigment metabolism and also maintained their nutritional value, thus could be employed as a potential commercial technique for citrus degreening.

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In table grape production, protected cultivation in a vineyard in different regions and climates is currently a commonly used practice. The aims of this study were to provide key approaches to sustainably produce two crops of grape without overlap under protected environment in a single year. Spraying the degreening chemicals 400 mg/L ethephon +0.4% sulfur at 4 weeks of vine nutrient restoration after the harvest of the summer crop resulted in the highest percentage of sprouting inflorescence. The retention of 7-10 buds in the base shoot results in the high percentage of sprouting inflorescence. Bud breaking chemicals with 2.5% hydrogen cyanamide+2.0% Baoguoliang +0.02% Shenzhonggen significantly led to sprout inflorescence more efficiently. Cluster and fruit weights of the winter crop weighed significantly less than those of the summer crop. However, the contents of total soluble sugar and titratable acidity were higher than those of the summer crop. The anthocyanin content in the peel of the winter fruit was significantly higher than that in the summer fruit. The yield of the winter crop is controlled by the yield of the summer fruit. To maintain the stability of the two crops for one year, the ratio of yield in the winter to the summer should be controlled from 2:5 to 3:5 to ensure the sustainable production of two crops without overlap for \'Summer Black\' grape. These results may help grape growers to overcome the impacts of rainy and hot climates with the help of protected facilities, and it could enable the use of solar radiation and heat resources in subtropical and tropical areas.

Citrus fruit postharvest degreening is a critical stage in marketing, carried out by exposure to ethylene or ethephon. Genome-wide screening of the AP2/ERF superfamily indicated that a novel ERF-II (CitERF6) was shown to trans-activate the CitPPH promoter. Expression of CitERF6 is associated with both developmental and postharvest degreening in citrus fruit. Transient and stable over-expression of CitERF6 in Nicotiana tabacum leaves and \'Ponkan\' fruit also results in rapid chlorophyll degradation. Auto- and mutual-regulation was also found between CitERF6 and the previously characterized CitERF13 using the dual-luciferase and yeast one-hybrid assays. Moreover, substitution of the 35S promoter for endogenous promoters showed that both pCitERF6::CitERF6 and pCitERF13::CitERF13 were effective in trans-activating their promoters or triggering chlorophyll degradation. It is proposed that ethylene is one of the triggers activating promoters of CitERF6 and CitERF13, and subsequent auto- and mutual-regulation between CitERF6 and CitERF13 might facilitate the effect of ethylene, leading to fruit degreening.