Transcription factors DkBZR1/2 regulate cell wall degradation genes and ethylene biosynthesis genes during persimmon fruit ripening.

BRASSINAZOLE RESISTANT (BZR) transcription factors are critical components of the brassinosteroid signalling pathway, but their possible roles in fruit ripening have rarely been reported. In this study, four BZR sequences were isolated from persimmon fruit. Among the four BZR genes, DkBZR1/2 were expressed in persimmon fruit; DkBZR1 protein amount decreased and dephosphorylated DkBZR2 gradually accumulated during the storage period. DkBZR1/2 proteins were localized in both the nucleus and cytoplasm and accumulated in the nucleus after 24-epibrassinolide treatment. DkBZR1 suppressed the transcription of Diospyros kaki endo-1,4-betaglucanase 1 (DkEGase1) and 1-aminocyclopropane-1-carboxylate synthase 1 (DkACS1) by binding to the BR response element (BRRE) in their promoters, and DkBZR2 activated the transcription of pectate lyase 1 (DkPL1) and 1-aminocyclopropane-1-carboxylate oxidase 2 (DkACO2) by binding to the E-box motif in their promoters. Transient overexpression of DkBZR2 promoted the conversion of acid-soluble pectin to water-soluble pectin and increased ethylene production in persimmon fruit. Our findings indicate that DkBZR1 and DkBZR2 serve as repressors and activators of persimmon fruit ripening, respectively.

Genome-wide analysis of the bZIP gene family and the role of AchnABF1 from postharvest kiwifruit (Actinidia chinensis cv. Hongyang) in osmotic and freezing stress adaptations.

Chilling injury (CI) is a barrier to the refrigeration of kiwifruit, resulting in decreased fruit quality and increased nutrient loss during storage. Understanding the molecular basis underlying the cold response and its regulation in refrigerated kiwifruit is therefore highly important. Basic (region) leucine zipper (bZIP) transcription factors (TFs) have been widely studied for their roles in abiotic stress resistance in various species. In this study, we identified 81 bZIP family proteins in kiwifruit and classified them into 11 groups. Further transcriptome analysis revealed that the expression of members of the AREB/ABF family was strongly induced by low temperature and abscisic acid (ABA). Ectopic expression of AchnABF1 enhanced plant cold tolerance by upregulating the expression of several key genes associated with ABA-dependent and ABA-independent pathways in Arabidopsis thaliana. Reactive oxygen species (ROS) metabolism was suggested to be involved in the AchnABF1-mediated osmotic stress response. For instance, enhanced ROS-scavenging ability was observed in transgenic plants with enhanced activity of catalase (CAT) and peroxidase (POD), which resulted in decreased in situ O2.- and H2O2 accumulation, ion leakage, and malondialdehyde (MDA) content under various abiotic stresses. In addition, AchnABF1 also participated in the osmotic stress response during both the germination and postgermination stages. We concluded that AchnABF1 may play an important role in kiwifruit during refrigeration.

Functional characterization of persimmon ß-galactosidase gene DkGAL1 in tomato reveals cell wall modification related to fruit ripening and radicle elongation.

Cell wall metabolism during fruit ripening is a highly organized process that involves complex interplay among various cell wall hydrolases. Among these cell wall hydrolases, ß-galactosidase has been identified to participate in cell wall metabolism via its ability to catalyze galactosyl metabolism from the large and complex side chains of cell walls. In this study, the galactose content in the pericarp increased during persimmon fruit ripening, but cell wall galactosyl residues decreased, indicating a relationship between galactose metabolism and persimmon fruit ripening. Expression of a previously isolated ß-galactosidase gene, DkGAL1, increased 25.01-fold during fruit ripening. Heterologous expression of DkGAL1 under the CaMV 35S promoter in tomato accelerated on-plant and postharvest fruits ripening. The fruit firmness of one of transgenic line, OE-18, was 23.83% lower than that of WT at the breaker stage. The transgenic fruits produced more ethylene by promoting the expression of ethylene synthesis-related genes and cell wall degradation-related genes. Overexpression of DkGAL1 in tomato also reduced cell-to-cell adhesion and promoted both wider intercellular spaces and less cell compaction in transgenic fruit structures. Moreover, DkGAL1 was involved in seed germination and radicle elongation in transgenic tomato seeds. These results confirm the role of DkGAL1 in fruit ripening and suggest that this gene alters galactose metabolism in the fruit, which can promote ripening and reduce cellular adhesion. In addition, the role of DkGAL1 is not limited to fruit softening; DkGAL1 was also involved in seed germination and radicle elongation in transgenic tomato seeds.

Overexpression of persimmon DkXTH1 enhanced tolerance to abiotic stress and delayed fruit softening in transgenic plants.

KEY MESSAGE: DkXTH1 promoted cell elongation and more strength to maintain structural integrity by involving in cell wall assembly, thus enhanced tolerance to abiotic stress with broader phenotype in transgenic plants. Xyloglucan endotransglucosylase/hydrolase (XTH) is thought to play a key role in cell wall modifications by cleaving and re-joining xyloglucan, and participates in the diverse physiological processes. DkXTH1 was found to peak in immature expanding persimmon fruit, and its higher expression level exhibited along with firmer fruit during storage. In the present study, transgenic Arabidopsis and tomato plants were generated with DkXTH1 constitutively expressed. Overexpression of DkXTH1 enhanced tolerance to salt, ABA and drought stresses in transgenic Arabidopsis plants with respect to root and leaf growth, and survival. Transgenic tomatoes collected at the mature green stage, presented delayed fruit softening coupled with postponed color change, a later and lower ethylene peak, and higher firmness in comparison with the wild-type tomatoes during storage. Furthermore, broader leaves and tomato fruit with larger diameter were gained in transgenic Arabidopsis and tomato, respectively. Most importantly, transgenic plants exhibited more large and irregular cells with higher density of cell wall and intercellular spaces, resulting from the overactivity of XET enzymes involving in cell wall assembly. We suggest that DkXTH1 expression resulted in cells with more strength and thickness to maintain structural integrity, and thus enhanced tolerance to abiotic stress and delayed fruit softening in transgenic plants.

DkXTH8, a novel xyloglucan endotransglucosylase/hydrolase in persimmon, alters cell wall structure and promotes leaf senescence and fruit postharvest softening.

Fruit softening is mainly associated with cell wall structural modifications, and members of the xyloglucan endotransglucosylase/hydrolase (XTH) family are key enzymes involved in cleaving and re-joining xyloglucan in the cell wall. In this work, we isolated a new XTH gene, DkXTH8, from persimmon fruit. Transcriptional profiling revealed that DkXTH8 peaked during dramatic fruit softening, and expression of DkXTH8 was stimulated by propylene and abscisic acid but suppressed by gibberellic acid and 1-MCP. Transient expression assays in onion epidermal cells indicated direct localization of DkXTH8 to the cell wall via its signal peptide. When expressed in vitro, the recombinant DkXTH8 protein exhibited strict xyloglucan endotransglycosylase activity, whereas no xyloglucan endohydrolase activity was observed. Furthermore, overexpression of DkXTH8 resulted in increased leaf senescence coupled with higher electrolyte leakage in Arabidopsis and faster fruit ripening and softening rates in tomato. Most importantly, transgenic plants overexpressing DkXTH8 displayed more irregular and twisted cells due to cell wall restructuring, resulting in wider interstitial spaces with less compact cells. We suggest that DkXTH8 expression causes cells to be easily destroyed, increases membrane permeability and cell peroxidation, and accelerates leaf senescence and fruit softening in transgenic plants.