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Suberin consisting of the polyaliphatic and polyphenolic domain confers the hydrophobic barrier of wound fruit and vegetables, providing essential protection from excessive water loss, ion outflow and pathogen invasion. The ω-hydroxyacid/fatty alcohol hydroxycinnamoyl transferase (FHT) is required for cross-linking of the polyaliphatic and polyphenolic domain in suberin biosynthesis. The transcription factor AchMYC2 acts as a positive activator in suberin biosynthesis but it is not known how it interacts with AchFHT in kiwifruit. In this study, overexpression of AchFHT in kiwifruit increased accumulation of suberin monomers, such as ferulate and ω-hydroxyacids. Yeast one-hybrid (Y1H) and dual-luciferase reporter (DLR) assays showed that AchMYC2 activated AchFHT transcript via interacting with the gene promoter. Overexpression of AchMYC2 in kiwifruit resulted in the upregulation of AchFHT , the accumulation of FHT protein and suberin monomers. Further, using yeast two-hybrid (Y2H) assay screened a jasmonate ZIM-domain protein (AchJAZ1, a key repressor of JA signaling pathway) which interacted with AchMYC2. Methyl jasmonate (MeJA) treatment increased expression levels of AchMYC2 and AchFHT , and the content of FHT protein, accompanying by the rapid wound healing process, as indicated by rapid suberin deposition, the reduced weight loss and relative electrolyte leakage. These results indicate that AchMYC2 activates AchFHT transcript to involve in MeJA-mediated suberin biosynthesis in wound healing of kiwifruit. • Methyl jasmonate (MeJA) accelerated suberin deposition in wound healing of kiwifruit. • MeJA stimulated expression levels of AchFHT and AchMYC2 and increased FHT content. • AchMYC2 activated AchFHT transcript by interacting with the gene promoter. [ABSTRACT FROM AUTHOR]

Background: Our previous study has demonstrated that the transcription of AchnKCS involved in suberin biosynthesis was up-regulated by exogenous abscisic acid (ABA) during the wound suberization of kiwifruit, but the regulatory mechanism has not been fully elucidated. Results: Through subcellular localization analysis in this work, AchnbZIP29 and AchnMYB70 transcription factors were observed to be localized in the nucleus. Yeast one-hybrid and dual-luciferase assay proved the transcriptional activation of AchnMYB70 and transcriptional suppression of AchnbZIP29 on AchnKCS promoter. Furthermore, the transcription level of AchnMYB70 was enhanced by ABA during wound suberization of kiwifruit, but AchnbZIP29 transcription was reduced by ABA. Conclusions: Therefore, it was believed that ABA enhanced the transcriptional activation of AchnMYB70 on AchnKCS by increasing AchnMYB70 expression. On the contrary, ABA relieved the inhibitory effect of AchnbZIP29 on transcription of AchnKCS by inhibiting AchnbZIP29 expression. These results gave further insight into the molecular regulatory network of ABA in wound suberization of kiwifruit. [ABSTRACT FROM AUTHOR]

Wound damage triggers the accumulation of abscisic acid (ABA), which induces the expression of a large number of genes involved in wound suberization in plants. Fatty acyl-CoA reductase (FAR) catalyzes the generation of primary fatty alcohols by the reduction of fatty acids in suberin biosynthesis. However, the regulatory effects of transcription factors (TFs) on AchnFAR in response to ABA are unexplored. In this study, kiwifruit AchnFAR displayed a biological function analogous to that of FAR in transiently overexpressed tobacco (Nicotiana benthamiana) leaves. The positive role of TFs, including AchnMYB41, AchnMYB107, and AchnMYC2, in the regulation of AchnFAR was identified. The three TFs could individually bind to the AchnFAR promoter to activate gene transcription in yeast one-hybrid and dual-luciferase assays. Transient overexpression of TFs in tobacco leaves resulted in the upregulation of aliphatic synthesis genes (including FAR) and the increase in aliphatics, including primary alcohols, α,ω-diacids, ω-hydroxyacids, and fatty acids. Moreover, exogenous ABA treatment elevated TF-mediated AchnFAR expression and the accumulation of primary alcohols. Conversely, fluridone, an inhibitor of ABA biosynthesis, suppressed the expression of AchnFAR and TF genes and reduced the formation of primary alcohols. The results indicate that AchnMYB41, AchnMYB107, and AchnMYC2 activate AchnFAR transcription to promote ABA-mediated primary alcohol formation in wound suberization in kiwifruit. [ABSTRACT FROM AUTHOR]

Wound attack stimulates accumulation of abscisic acid (ABA) that activates a number of genes associated with wound suberization of plants. Cytochrome P450 fatty acid ω-hydroxylase CYP86A1 catalyzes ω-hydroxylation of fatty acids to form the ω-functionalized monomers that play a pivotal role in suberin synthesis. However, the transcriptional regulation of ABA signaling on AchnCYP86A1 has not been characterized in kiwifruit. In this study, AchnCYP86A1 , a kiwifruit homolog of Arabidopsis AtCYP86A1 , was isolated. AchnCYP86A1 -overexpressed N. benthamiana leaves displayed that the AchnCYP86A1 functioned as a fatty acid ω-hydroxylase associated with synthesis of suberin monomer. The regulatory function of three transcription factors (TFs, including AchnMYC2, AchnMYB41 and AchnMYB107) on AchnCYP86A1 was identified. All the three TFs were localized in nucleus and could individually interact with AchnCYP86A1 promoter to activate gene expression in yeast one-hybrid and dual-luciferase assays. The findings were further demonstrated in transient overexpressed N. benthamiana , in which all TFs notably elevated the expression of aliphatic synthesis genes including CYP86A1 and the accumulation of ω-hydroxyacids, α, ω-diacids, fatty acids and primary alcohols. Moreover, exogenous ABA induced the expression of AchnMYC2 , AchnMYB41 and AchnMYB107 that promoted AchnCYP86A1 involving in suberin monomer formation. Contrary to the inductive effects of ABA, however, fluridone (an inhibitor of ABA biosynthesis) inhibited the three TFs expression and suberin monomer formation. These results indicate that AchnMYC2, AchnMYB41 and AchnMYB107 positively regulate suberin monomer synthesis by activating AchnCYP86A1 promoter in response to ABA. [ABSTRACT FROM AUTHOR]

Suberin is a cell-wall biopolymer with aliphatic and aromatic domains that is synthesized in the wound tissues of plants in order to restrict water loss and pathogen infection. ω-hydroxyacid/fatty alcohol hydroxycinnamoyl transferase (FHT) is required for cross-linking of the aliphatic and aromatic domains. ABA is known to play a positive role in suberin biosynthesis but it is not known how it interacts with FHT. In this study, the kiwifruit (Actinidia chinensis) AchnFHT gene was isolated and was found to be localized in the cytosol. Transient overexpression of AchnFHT in leaves of Nicotiana benthamiana induced massive production of ferulate, ω-hydroxyacids, and primary alcohols, consistent with the in vitro ability of AchnFHT to catalyse acyl-transfer from feruloyl-CoA to ω-hydroxypalmitic acid and 1-tetradecanol. A regulatory function of four TFs (AchnABF2, AchnMYB4, AchnMYB41, and AchnMYB107) on AchnFHT was identified. These TFs localized in the nucleus and directly interacted with the AchnFHT promoter in yeast one-hybrid assays. Dual-luciferase analysis indicated that AchnABF2, AchnMYB41, and AchnMYB107 activated the AchnFHT promoter while AchnMYB4 repressed it. These findings were supported by the results of transient overexpression in N. benthamiana , in which AchnABF2 , AchnMYB41 , and AchnMYB107 induced expression of suberin biosynthesis genes (including FHT) and accumulation of suberin monomers, whilst AchnMYB4 had the opposite effect. Exogenous ABA induced the expression of AchnABF2 , AchnMYB41 , AchnMYB107 , and AchnFHT and induced suberin monomer formation, but it inhibited AchnMYB4 expression. In addition, fluridone (an inhibitor of ABA biosynthesis) was found to counter the inductive effects of ABA. Activation of suberin monomer biosynthesis by AchnFHT was therefore controlled in a coordinated way by both repression of AchnMYB4 and promotion of AchnABF2, AchnMYB41, and AchnMYB107. [ABSTRACT FROM AUTHOR]

Highlights • Kiwifruit water loss was spatial movement of water from interior tissues to outwards. • Vibration caused excess water loss although there was an undamaged fruit surface. • Vibration accelerated water loss process due to intracellular damage in kiwifruit. Abstract Fruit usually suffers from water loss after transportation and storage, which largely blemishes the quality characteristics including appearance, saleable weight and texture. This study was conducted to interpret the potential relations between fruit water loss and transport vibration. Kiwifruit were subjected to vibration using an electrodynamic shaker followed by storing at 25 °C and 75% RH (relative humidity) in dark for 12 d. Fruit weight loss and water content in the epidermis (EP), outer pericarp (OP), inner pericarp (IP) and core tissues were determined. Shrinkage of EP cells and water movement were evaluated using laser scanning confocal microscopy and magnetic resonance imaging, respectively. Cell damage was investigated by observing the ultrastructure and electrolyte leakage. The results showed that fruit water loss was accompanied with the spatial movement of water from interior IP, OP and external EP. Vibration damaged the fruit cells with plasmolysis, membrane contracting and increased electrolyte leakage, which accelerated movement of water from the tissues to outward resulting in severe water loss of fruit. However, the electrolyte leakage and water content in the core were little changed in both control and vibrated fruit. Shrinkage of EP cells coupled with fruit shrivel appeared by 4 d of storage after vibration. However, the symptom in control fruit was not observed until 8 d of storage. It could be concluded that the simulated transport vibration caused intracellular damage in fruit tissues, which accelerated the water loss and shriveling process. [ABSTRACT FROM AUTHOR]

Abstract: BACKGROUND: Rapid wound healing would be critical for successful long‐term storage of fruits and vegetables. However, there was no direct evidence for the requirement and efficiency of oxygen in the fruit wound‐healing process. This study was conducted to investigate the role of oxygen in wound‐induced suberization by analyzing melanin, suberin polyphenolics (SPPs) and related enzymes in half‐cut kiwifruits exposed to 100%, 50%, 21% and 0% oxygen. RESULTS: By 3 days after wounding, the wound surface of kiwifruit in high (50 and 100%) oxygen appeared as a continuous layer of melanin and SPPs underneath, which effectively prevent excessive water vapor loss from the fruit halves. In contrast, melanin and SPPs deposition in the wound surface in 0% oxygen was significantly reduced, with high water vapor loss. Rapid decrease of soluble phenolic acids (caffeic, p‐coumaric, ferulic acids) was coupled with the increase of bound ferulic acid (coniferyl diacetate) especially in high oxygen by 9 days after wounding. Meanwhile, high oxygen enhanced peroxidase, catalase, phenylalanine ammonia‐lyase, and polyphenol oxidase activities. CONCLUSION: Oxygen is required for wound‐induced melanin and SPPs formation, and high oxygen is effective in promoting wound suberization in postharvest kiwifruit. © 2017 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

• AchLOX3 and AchOPR3 were involved in JA biosynthesis of kiwifruit. • ABA-induced AchABF1-1 was a nucleus-localized transcription factor. • AchABF1-1 activated AchLOX3 and AchOPR3 expression by binding to their promoters. • ABA promoted SPP formation by the induction of JA biosynthesis. Mechanical damage is a central limiting factor in the maintenance of the postharvest fruit quality during harvest, transportation, and processing. In this study, wounded kiwifruit (Actinidia chinensis cv. Xuxiang) were respectively treated with abscisic acid (ABA), fluridone (FLD, an inhibitor of ABA biosynthesis), methyl jasmonate (MeJA), diethyldithiocarbamic acid (DIECA, an inhibitor of JA biosynthesis), and ABA+DIECA to investigate the potential interaction between ABA and JA in the process of wound healing. The results showed that blocking JA biosynthesis significantly reduced the positive effect of ABA on the accumulation of suberin polyphenolic (SPP). Besides, transiently overexpressed the AchLOX3 and AchOPR3 in tobacco (Nicotiana benthamiana) leaves confirmed that these two genes respectively encoded lipoxygenase and 12-oxo-phytodienoic acid reductase involved in JA biosynthesis. Notably, the positive regulation of transcription factor (TF) AchABF1-1 on AchLOX3 and AchOPR3 was identified. ABA-induced AchABF-1 located in the nucleus and activated the transcriptional expression of AchLOX3 and AchOPR3 through binding to their promoters. These results were further confirmed in the tobacco leaves overexpressing AchABF1-1 in which AchABF1-1 promoted NbLOX and NbOPR expression with the increases of JA content. In addition, exogenous ABA increased endogenous JA content, and enhanced LOX and OPR activities as well as their related gene expression, while FLD reduced these indices. These results suggested that the promotion effect of JA on the SPP accumulation was enhanced through the promotion of JA biosynthesis by ABA-responsive transcription factor AchABF1-1 during the wound healing process of kiwifruit. [ABSTRACT FROM AUTHOR]

• AchnCYP86A1 and AchnFAR were involved in wound suberinzation of kiwifruit. • AcnMYB4 repressed AchnCYP86A1 and AchnFAR expression by binding to gene promoter. • ABA induced the AchnCYP86A1 and AchnFAR expression by inhibition of AchnMYB4. Suberin biosynthesis involves a large number of genes, and many of them are induced by abscisic acid (ABA). However, the regulation of transcription factor (TF) in response to ABA on suberin biosynthetic genes in kiwifruit has been unexplored. In this study, two genes, AchnCYP86A1 and AchnFAR respectively encoding a fatty acid ω-hydroxylase and fatty acyl-CoA reductase involved in suberin monomer biosynthesis were demonstrated in transient overexpressed tobacco (Nicotiana benthamiana). Notably, the negative regulation of AchnMYB4 on AchnCYP86A1 and AchnFAR was identified. AchnMYB4 could repress AchnCYP86A1 and AchnFAR transcript by directly binding to the gene promoter in yeast one-hybrid and dual-luciferase assays. These results were further confirmed in transient overexpressed tobacco leaves in that AchnMYB4 significantly down-regulated suberin biosynthetic genes including CYP86A1 , FAR2 and FAR3 , and reduced accumulation of ω-hydroxyacids and primary alcohols. Moreover, exogenous ABA could induce the expression of AchnCYP86A1 and AchnFAR , and the accumulation of corresponding suberin monomers by inhibition of AchnMYB4 in wound-tissue of kiwifruit. However, fluridone (an inhibitor of ABA biosynthesis) was found to counter the inductive effects of ABA. Taken together, the results suggest that ABA activates AchnCYP86A1 and AchnFAR to promote suberin monomers biosynthesis by inhibiting AchnMYB4. [ABSTRACT FROM AUTHOR]

Quick suberin-based healing after wounding played a protective role for plant to prevent further damage. In this study, the stimulative effect of exogenous abscisic acid (ABA) on wound suberization in postharvest kiwifruit was evaluated through suberin staining with toluidine blue O as well as the determination of suberin phenolics and aliphatics in wound tissue. Furthermore, to reveal the regulatory involvement of ABA in wound suberization, comparative quantitative proteomics and transcriptomics analyses based on iTRAQ and qRT-PCR technique were performed. In proteomics levels, a total of 95 protein species consistently showed differential abundance between ABA and control, including 29 down-regulated and 66 up-regulated protein species. The Kyoto Encyclopedia of Genes and Genomes (KEGG) with protein-protein interaction analyses revealed that ABA mainly affected the antioxidant system, phenylpropanoid metabolism and lipid metabolism associated with wound suberization. Based on the data of proteomics analysis, the differential expressions of genes encoding 11 selected protein species were confirmed by qRT-PCR analyses. GSH-Px, MDHAR, SOD, APX, POD, PAL, CCR, PPO, CYP86B1, DGGT and KCS11 were likely to be the key enzymes that involved the response of ABA to stimulate wound suberization by mediating the antioxidant system, phenylpropanoid metabolism and lipid metabolism. Biological significance Kiwifruit is susceptible to physical injury causing postharvest deterioration during harvest, transportation and storage. Therefore, quick healing is important for maintaining the postharvest quality of injured fruit. This work elucidated the potential role of ABA and the proteomic mechanism of its regulation in wound suberization of postharvest kiwifruit. [ABSTRACT FROM AUTHOR]

Wound-induced suberization is an essentially protective healing process for fruit to reduce water loss and avoid infecting. However, cognate mechanisms that regulate this process are little known. To expand our knowledge of suberization induced by wounding, a wound-healing investigation together with metabolite profiling study was conducted in postharvest kiwifruit ( Actinidia deliciosa ). The development of suberization in wounded fruit was demonstrated by autofluorescence observation and toluidine blue staining at 1–4 day (d) after wounding. Activities of phenylalanine ammonia-lyase (PAL), cinnamyl-alcohol dehydrogenase (CAD) and peroxidase (POD) in wound-healing tissue were enhanced by abscisic acid (ABA). The constituent analysis of suberin including polyphenolics (SPP) and polyaliphatics (SPA) proved that exogenous ABA increased the content levels of total phenols, total flavonoids and alkanes, alkenes, alcohols, alkane acids, olefine acids, esters, glycerides and vitamin E in wound-healing tissue. Results suggested that ABA stimulated suberization through the activation of PAL, CAD and POD to accelerate wound-healing of wounded kiwifruit. [ABSTRACT FROM AUTHOR]

Suberin polyphenolic (SPP), a kind of agronomically significant biopolymer, is composed primarily of phenolic acids generally synthesized by phenylpropanoid metabolism. In this study, the transcriptional activation of suberin polyphenolic biosynthesis by jasmonic acid (JA) inducible-transcription factor AchMYC2 was confirmed in wound tissues of kiwifruit (Actinidia chinensis). Three general phenylpropanoid metabolism-related genes, AchPAL , AchC4H and Ach4CL respectively encoding phenylalanine ammonia-lyase (PAL), cinnamic acid-4-hydroxylase (C4H) and 4-coumaric-CoA ligase were determined in the kiwifruit core tissues overexpressing these genes. Besides, the positive regulation of AchMYC2 on AchPAL , AchC4H and Ach4CL was observed. JA-induced AchMYC2 locating in the nucleus activated the transcriptional expression of AchPAL , AchC4H and Ach4CL through binding to their promoters. The results were further evidenced in the kiwifruit core tissues overexpressing of AchMYC2, in which AchMYC2 enhanced the expression levels of AchPAL , AchC4H and Ach4CL and increased the accumulation of phenolic acids including cinnamic acid, p -coumaric acid, caffeic acid, ferulic acid and sinapic acid. Meanwhile, the contents of five phenolic acids in the kiwifruit wound tissues were increased by methyl jasmonate (MeJA), but reduced by diethyldithiocarbamic acid (DIECA, an inhibitor of JA biosynthesis). Together results indicated that AchMYC2 activated AchPAL , AchC4H and Ach4CL transcription to promote JA-mediated suberin polyphenolic accumulation in the wound healing of kiwifruit. • AchPAL, AchC4H, and Ach4CL involved in suberin polyphenolic precursors synthesis. • MeJA-induced AchMYC2 was a nucleus-localized transcription factor. • AcnMYC2 activates AchPAL, AchC4H, and Ach4CL expression by binding to their promoters. • AchMYC2 promotes JA-mediated suberin polyphenolic accumulation of wounded kiwifruit. [ABSTRACT FROM AUTHOR]