NAC transcription factor SlNOR-like1 plays a dual regulatory role in tomato fruit cuticle formation.

The plant cuticle is an important protective barrier on the plant surface, constructed mainly by polymerized cutin matrix and a complex wax mixture. Although the pathway of plant cuticle biosynthesis has been clarified, knowledge of the transcriptional regulation network underlying fruit cuticle formation remains limited. In the present work, we discovered that tomato fruits of the NAC transcription factor SlNOR-like1 knockout mutants (nor-like1) produced by CRISPR/Cas9 [clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9] displayed reduced cutin deposition and cuticle thickness, with a microcracking phenotype, while wax accumulation was promoted. Further research revealed that SlNOR-like1 promotes cutin deposition by binding to the promoters of glycerol-3-phosphate acyltransferase6 (SlGPAT6; a key gene for cutin monomer formation) and CUTIN DEFICIENT2 (SlCD2; a positive regulator of cutin production) to activate their expression. Meanwhile, SlNOR-like1 inhibits wax accumulation, acting as a transcriptional repressor by targeting wax biosynthesis, and transport-related genes 3-ketoacyl-CoA synthase1 (SlKCS1), ECERIFERUM 1-2 (SlCER1-2), SlWAX2, and glycosylphosphatidylinositol-anchored lipid transfer protein 1-like (SlLTPG1-like). In conclusion, SlNOR-like1 executes a dual regulatory effect on tomato fruit cuticle development. Our results provide a new model for the transcriptional regulation of fruit cuticle formation.

NAC transcription factor NOR-like1 regulates tomato fruit size.

MAIN CONCLUSION: NOR-like1 regulates tomato fruit size by targeting SlARF9, SlGRAS2, SlFW3.2, and SlFW11.3 genes involved in cell division and cell expansion. Fruit size is an important agricultural character that determines the yield of crops. Here, we found that NAC transcription factor NOR-like1 regulated fruit size by regulating cell layer number and cell area in tomato. Over-expressing NOR-like1 gene in tomato reduced fruit weight and size, whereas the knock-out of NOR-like1 increased fruit weight and size. At the molecular level, NOR-like1 binds to the promoter of SlGRAS2, SlFW3.2, and SlFW11.3 to repress their transcription, while it also binds to the promoter of ARF9 to activate its transcription. Overall, these results expand the biological function of NOR-like1 and deepen our understanding of the transcriptional network that regulates tomato fruit size.

The Ubiquitin-26S Proteasome Pathway and Its Role in the Ripening of Fleshy Fruits.

The 26S proteasome is an ATP-dependent proteolytic complex in eukaryotes, which is mainly responsible for the degradation of damaged and misfolded proteins and some regulatory proteins in cells, and it is essential to maintain the balance of protein levels in the cell. The ubiquitin-26S proteasome pathway, which targets a wide range of protein substrates in plants, is an important post-translational regulatory mechanism involved in various stages of plant growth and development and in the maturation process of fleshy fruits. Fleshy fruit ripening is a complex biological process, which is the sum of a series of physiological and biochemical reactions, including the biosynthesis and signal transduction of ripening related hormones, pigment metabolism, fruit texture changes and the formation of nutritional quality. This paper reviews the structure of the 26S proteasome and the mechanism of the ubiquitin-26S proteasome pathway, and it summarizes the function of this pathway in the ripening process of fleshy fruits.

Composition, metabolism and postharvest function and regulation of fruit cuticle: A review.

The cuticle of plants, a hydrophobic membrane that covers their aerial organs, is crucial to their ability to withstand biotic and abiotic stressors. Fruit is the reproductive organ of plants, and an important dietary source that can offer a variety of nutrients for the human body, and fruit cuticle performs a crucial protective role in fruit development and postharvest quality. This review discusses the universality and diversity of the fruit cuticle composition, and systematically summarizes the metabolic process of fruit cuticle, including the biosynthesis, transport and regulatory factors (including transcription factors, phytohormones and environmental elements) of fruit cuticle. Additionally, we emphasize the postharvest functions and postharvest regulatory technologies of fruit cuticle, and propose future research directions for fruit cuticle.

Molecular and Genetic Events Determining the Softening of Fleshy Fruits: A Comprehensive Review.

Fruit softening that occurs during fruit ripening and postharvest storage determines the fruit quality, shelf life and commercial value and makes fruits more attractive for seed dispersal. In addition, over-softening results in fruit eventual decay, render fruit susceptible to invasion by opportunistic pathogens. Many studies have been conducted to reveal how fruit softens and how to control softening. However, softening is a complex and delicate life process, including physiological, biochemical and metabolic changes, which are closely related to each other and are affected by environmental conditions such as temperature, humidity and light. In this review, the current knowledge regarding fruit softening mechanisms is summarized from cell wall metabolism (cell wall structure changes and cell-wall-degrading enzymes), plant hormones (ETH, ABA, IAA and BR et al.), transcription factors (MADS-Box, AP2/ERF, NAC, MYB and BZR) and epigenetics (DNA methylation, histone demethylation and histone acetylation) and a diagram of the regulatory relationship between these factors is provided. It will provide reference for the cultivation of anti-softening fruits.

Transcription factor SlBEL2 interferes with GOLDEN2-LIKE and influences green shoulder formation in tomato fruits.

Chloroplasts play a crucial role in plant growth and fruit quality. However, the molecular mechanisms of chloroplast development are still poorly understood in fruits. In this study, we investigated the role of the transcription factor SlBEL2 (BEL1-LIKE HOMEODOMAIN 2) in fruit of Solanum lycopersicum (tomato). Phenotypic analysis of SlBEL2 overexpression (OE-SlBEL2) and SlBEL2 knockout (KO-SlBEL2) plants revealed that SlBEL2 has the function of inhibiting green shoulder formation in tomato fruits by affecting the development of fruit chloroplasts. Transcriptome profiling revealed that the expression of chloroplast-related genes such as SlGLK2 and SlLHCB1 changed significantly in the fruit of OE-SlBEL2 and KO-SlBEL2 plants. Further analysis showed that SlBEL2 could not only bind to the promoter of SlGLK2 to inhibit its transcription, but also interacted with the SlGLK2 protein to inhibit the transcriptional activity of SlGLK2 and its downstream target genes. SlGLK2 knockout (KO-SlGLK2) plants exhibited a complete absence of the green shoulder, which was consistent with the fruit phenotype of OE-SlBEL2 plants. SlBEL2 showed an expression gradient in fruits, in contrast with that reported for SlGLK2. In conclusion, our study reveals that SlBEL2 affects the formation of green shoulder in tomato fruits by negatively regulating the gradient expression of SlGLK2, thus providing new insights into the molecular mechanism of fruit green shoulder formation.

A MADS-box transcription factor, SlMADS1, interacts with SlMACROCALYX to regulate tomato sepal growth.

In flowering plants, sepals play important roles in the development of flowers and fruit, and both processes are regulated by MADS-box (MADS) transcription factors (TFs). SlMADS1 was previously reported to act as a negative regulator of fruit ripening. In this study, expression analysis shown that its transcripts were very highly expressed during the development of sepals. To test the role of SlMADS1, we generated KO-SlMADS1 (knock-out) tomato mutants by CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats and CRISPR-associated protein 9) technology and over-expression of SlMADS1 (OE-SlMADS1). The sepals and individual cells of KO-SlMADS1 mutants were significantly elongated, compared with the wild type (WT), whereas the sepals of OE-SlMADS1 tomatoes were significantly shorter and their cells were wider. RNA-seq (RNA-sequencing) of sepal samples showed that ethylene-, gibberellin-, auxin-, cytokinin- and cell wall metabolism-related genes were significantly affected in both KO-SlMADS1 and OE-SlMADS1 plants with altered sepal size. Since SlMACROCALYX (MC) is known to regulate the development of tomato sepals, we also studied the relationship between SlMC and SlMADS1 and the result showed that SlMADS1 interacts directly with SlMC. In addition, we also found that manipulating SlMADS1 expression alters the development of tomato plant leaves, roots and plant height. These results enrich our understanding of sepal development and the function of SlMADS1 throughout the plant.

NAC Transcription Factor Family Regulation of Fruit Ripening and Quality: A Review.

The NAC transcription factor (TF) family is one of the largest plant-specific TF families and its members are involved in the regulation of many vital biological processes during plant growth and development. Recent studies have found that NAC TFs play important roles during the ripening of fleshy fruits and the development of quality attributes. This review focuses on the advances in our understanding of the function of NAC TFs in different fruits and their involvement in the biosynthesis and signal transduction of plant hormones, fruit textural changes, color transformation, accumulation of flavor compounds, seed development and fruit senescence. We discuss the theoretical basis and potential regulatory models for NAC TFs action and provide a comprehensive view of their multiple roles in modulating different aspects of fruit ripening and quality.

A tomato NAC transcription factor, SlNAM1, positively regulates ethylene biosynthesis and the onset of tomato fruit ripening.

Fruit ripening in tomato (Solanum lycopersicum) is the result of selective expression of ripening-related genes, which are regulated by transcription factors (TFs). The NAC (NAM, ATAF1/2, and CUC2) TF family is one of the largest families of plant-specific TFs and members are involved in a variety of plant physiological activities, including fruit ripening. Fruit ripening-associated NAC TFs studied in tomato to date include NAC-NOR (non-ripening), SlNOR-like1 (non-ripening like1), SlNAC1, and SlNAC4. Considering the large number of NAC genes in the tomato genome, there is little information about the possible roles of other NAC members in fruit ripening, and research on their target genes is lacking. In this study, we characterize SlNAM1, a NAC TF, which positively regulates the initiation of tomato fruit ripening via its regulation of ethylene biosynthesis. The onset of fruit ripening in slnam1-deficient mutants created by CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats and CRISPR-associated protein 9) technology was delayed, whereas fruit ripening in OE-SlNAM1 lines was accelerated compared with the wild type. The results of RNA-sequencing (RNA-seq) and promoter analysis suggested that SlNAM1 directly binds to the promoters of two key ethylene biosynthesis genes (1-aminocyclopropane-1-carboxylate synthase: SlACS2 and SlACS4) and activates their expression. This hypothesis was confirmed by electrophoretic mobility shift assays and dual-luciferase reporter assay. Our findings provide insights into the mechanisms of ethylene production and enrich understanding of the tomato fruit ripening regulatory network.

[Fruit cracking: a review].

Fruit cracking is a common physiological disease. Many fruits such as tomato, sweet cherry, apple, jujube, pomegranate, and litchi are liable to crack, causing considerable economic loss and agricultural resources waste. The mechanisms of fruit cracking are comprehensive. Some correlations have been observed between susceptibility of fruit cracking and some fruit traits (genetic, fruit size, fruit shape, fruit growth rate, water content, fruit skin characteristics, related gene expression, etc). Also, environmental condition (temperature, light, rainfall, etc) and orchard management (irrigation, sun-shade, mineral, growth regulator, etc) can influence fruit cracking. Here, progress in studies on fruit cracking is reviewed to provide a reference for prevention and control of fruit cracking.

[Analysis of apple postharment damage under high CO2 concentration by transcriptome combined with metabolome].

The environmental gas concentration affects the storage period and quality of fruits and vegetables. High concentration CO2 treating for a long time will cause damage to fruits, However, the specific molecular mechanism is unclear. To analyze the mechanism of CO2 injury in apple, high-throughput sequencing technology of Illumina Hiseq 4000 and non-targeted metabolism technology were used to analyze the transcriptome sequencing and metabolomics analysis of browning flesh tissue of damage fruit and normal pulp tissue of the control group. A total of 6 332 differentially expressed genes were obtained, including 4 187 up-regulated genes and 2 145 down regulated genes. Functional analysis of the differentially expressed genes confirmed that the occurrence of CO2 injury in apple was related to redox process, lipid metabolism, hormone signal transduction process and energy metabolism process. Twenty candidate browning genes were successfully screened, among which grxcr1 (md14g1137800) and gpx (md06g1081300) participated in the reactive oxygen species scavenging process, and pld1_ 2 (md15g1125000) and plcd (md07g1221900) participated in phospholipid acid synthesis and affected membrane metabolism. mdh1 (md05g1238800) participated in TCA cycle and affected energy metabolism. A total of 77 differential metabolites were obtained by metabolomic analysis, mainly organic acids, lipids, sugars and polyketones, including 35 metabolites related to browning. The metabolism of flavonoids was involved in the browning process of apple. Compared with the control tissue, the content of flavonoids such as catechin and quercetin decreased significantly in the damaged apple tissue, the antioxidant capacity of cells decreased, the redox state was unbalanced, and the cell structure was destroyed, resulting in browning. The results of this study further enrich the theoretical basis of CO2 damage, and provide reference for the practical application of high concentration CO2 preservation technology.

Inhibition of downy blight and enhancement of resistance in litchi fruit by postharvest application of melatonin.

Litchis are tasty fruit with economic importance. However, the extreme susceptibility of harvested litchis to litchi downy blight caused by Peronophythora litchii leads to compromised quality. This study aimed to study the effects of melatonin on postharvest resistance to P. litchii in 'Feizixiao' litchis. Results showed that melatonin restricted lesion expansion in litchis after P. litchi inoculation. Melatonin enhanced the activities of phenylalanine ammonia-lyase, cinnamate-4-hydroxylase and 4-hydroxycinnamate CoA ligase while promoting the accumulations of phenolics and flavonoids. Nicotinamide adenine dinucleotide phosphate content and glucose-6-phosphate dehydrogenase and 6-phosphogluconic acid dehydrogenase activities were higher in treated fruit than control fruit. Higher energy status along with elevated H+-ATPase, Ca2+-ATPase, succinate dehydrogenase and cytochrome C oxidase activities were observed in treated fruit. Ultrastructural observation showed reduced damage in mitochondria in treated fruit. The results suggest that melatonin induced resistance in litchis by modulating the phenylpropanoid and pentose phosphate pathways as well as energy metabolism. .

Melatonin Enhances Cold Tolerance by Regulating Energy and Proline Metabolism in Litchi Fruit.

Melatonin (MLT) is a vital signaling molecule that regulates multiple physiological processes in higher plants. In the current study, the role of MLT in regulating chilling tolerance and its possible mechanisms in litchi fruit during storage at ambient temperatures after its removal from refrigeration was investigated. The results show that the application of MLT (400 µM, dipping for 20 min) to 'Baitangying' litchi fruit effectively delayed the development of chilling injury (CI) while inhibiting pericarp discoloration, as indicated by higher chromacity values (L*, a*, b*) and anthocyanin levels. MLT treatment suppressed the enhancements of the relative electrical conductivity (REC) and malondialdehyde (MDA) content, which might contribute to the maintenance of membrane integrity in litchi fruit. MLT treatment slowed the decline in cellular energy level, as evidenced by higher adenosine triphosphate (ATP) content and a higher energy charge (EC), which might be ascribed to the increased activities of enzymes associated with energy metabolism including H+-ATPase, Ca2+-ATPase, succinate dehydrogenase (SDH), and cytochrome C oxidase (CCO). In addition, MLT treatment resulted in enhanced proline accumulation, which was likely a consequence of the increased activities of ornithine-δ-aminotransferase (OAT) and Δ1-pyrroline-5-carboxylate synthase (P5CS) and the suppressed activity of proline dehydrogenase (PDH). These results suggest that the enhanced chilling tolerance of litchi fruit after MLT treatment might involve the regulation of energy and proline metabolism.