Genomic Responses Associated with Drought Stress in Walnut as Revealed Transcriptome Sequencing

Walnut production is challenged by abiotic stresses. We investigated the leaf transcriptome responses of walnut under control and drought stress in 9 and 18 days. We identified 921, 1035 differentially expressed genes (DEGs) between control and drought stress groups in 9 and 18-day, respectively. In control and drought stress conditions DEGs were significantly enriched into the abscisic acid biosynthesis, regulation of stomata closure, leaf morphogenesis, carbohydrate metabolism, oxidative stress, cell wall macromolecule catabolism, and secondary metabolite biosynthesis pathways. We confirmed our RNA-Seq data using quantitative real-time PCR (qPCR) of six candidate genes. Our results indicated that more complicated transcript regulation of drought responses following prolong exposure to drought stress. In general, walnut activated more tolerance mechanisms 18 days after drought stress. Findings of this research would be useful for future studies on breeding for drought tolerance of Persian walnut and related species.

Genome-wide identification and expression profiling of durian CYPome related to fruit ripening

Durian (Durio zibethinus L.) is a major economic crop native to Southeast Asian countries, including Thailand. Accordingly, understanding durian fruit ripening is an important factor in its market worldwide, owing to the fact that it is a climacteric fruit with a strikingly limited shelf life. However, knowledge regarding the molecular regulation of durian fruit ripening is still limited. Herein, we focused on cytochrome P450, a large enzyme family that regulates many biosynthetic pathways of plant metabolites and phytohormones. Deep mining of the durian genome and transcriptome libraries led to the identification of all P450s that are potentially involved in durian fruit ripening. Gene expression validation by RT-qPCR showed a high correlation with the transcriptome libraries at five fruit ripening stages. In addition to aril-specific and ripening-associated expression patterns, putative P450s that are potentially involved in phytohormone metabolism were selected for further study. Accordingly, the expression of CYP72, CYP83, CYP88, CYP94, CYP707, and CYP714 was significantly modulated by external treatment with ripening regulators, suggesting possible crosstalk between phytohormones during the regulation of fruit ripening. Interestingly, the expression levels of CYP88, CYP94, and CYP707, which are possibly involved in gibberellin, jasmonic acid, and abscisic acid biosynthesis, respectively, were significantly different between fast- and slow-post-harvest ripening cultivars, strongly implying important roles of these hormones in fruit ripening. Taken together, these phytohormone-associated P450s are potentially considered additional molecular regulators controlling ripening processes, besides ethylene and auxin, and are economically important biological traits.

Ethylene Induced by Sound Stimulation Enhances Anthocyanin Accumulation in Grape Berry Skin through Direct Upregulation of UDP-Glucose: Flavonoid 3-O-Glucosyltransferase

Global warming has resulted in the loss of anthocyanin accumulation in berry skin. Sound stimulation can be used as a potential method for enhancing fruit color development since many plants recognize sound vibration as an external stimulus and alter their physiological status in response to it. Sound stimulation (sine wave sound at 1000 Hz) enhanced anthocyanin accumulation in grape cultured cells and berry skins in field-grown grapevines at the early stage of ripening. The transcription of UFGT and ACO2, which encode the key enzymes in anthocyanin and ethylene biosynthesis, respectively, was upregulated in grape cultured cells exposed to sound stimulation. In contrast, the transcription of MybA1 and NCED1, which encode a transcription factor for UFGT and a key enzyme in abscisic acid biosynthesis, respectively, was not affected by the sound stimulation. A treatment with an ethylene biosynthesis inhibitor, aminoethoxyvinyl glycine hydrochloride, revered the enhancement of anthocyanin accumulation by sound stimulation. As the promoter assay using a GUS reporter gene demonstrated that UFGT promoter was directly activated by the ethylene-releasing compound ethephon, which enhanced anthocyanin accumulation in grape cultured cells, we conclude that sound stimulation enhanced anthocyanin accumulation through the direct upregulation of UFGT by ethylene biosynthesis. Our findings suggest that sound stimulation contributes to alleviating poor coloration in berry skin as a novel and innovative practical technique in viticulture.

Class II LBD genes ZmLBD5 and ZmLBD33 regulate gibberellin and abscisic acid biosynthesis

Lateral organ boundaries domain (LBD) proteins are plant-specific transcription factors. Class I LBD members are widely reported to be pivotal for organ development, however, the role of class II members is unknown in cereal crops. Class II LBD proteins are distinguished from class I by the lack of a Gly-Ala-Ser (GAS) peptide and leucine-zipper-like coiled-coil domain, which is thought to be essential for protein dimerization. In this study, ZmLBD5 and ZmLBD33 form homo- and hetero-dimers, like class I members. At seedling stage, ZmLBD5 promoted biomass accumulation (shoot dry weight and root dry weight), root development (root length, root number, and root volume), and organ expansion (leaf area), while ZmLBD33 repressed these processes and display a dwarf phenotype. Both ZmLBD5 and ZmLBD33 displayed negative roles in drought tolerance mainly by increasing stomatal density and stomatal aperture. RNA sequencing, gene ontology enrichment analysis, and transient luciferase expression assays indicated that ZmLBD5 and ZmLBD33 are mainly involved in the regulation of the TPS-KS-GA2ox gene module, which comprises key enzymatic genes upstream of GA and ABA biosynthesis. GA1 content increased in ZmLBD5-overexpressing seedlings, while GA3 and abscisic acid content decreased in both transgenic seedlings. Consequently, exogenous GA1 or GA3 undoubtedly rescued the dwarf phenotype of ZmLBD33-overexpressing plants, with GA1 performing better. The study of ZmLBD5 and ZmLBD33 sheds light on the function of the class II LBD gene family in maize.

Cold stress triggers premature fruit abscission through ABA-dependent signal transduction in early developing apple

Fruit abscission is a complex physiological process that is regulated by internal and environmental factors. During early development, apple fruit are exposed to extreme temperature fluctuations that are associated with premature fruit drop; however, their effect on fruit abscission is largely unknown. We hypothesized that fruit abscission is triggered by cold stress and investigated the molecular basis of premature fruit drop using RNA-Seq and metabolomics data from apple fruit undergoing abscission following cold stress in the field. Genes responsive to abscisic acid signaling and cell wall degradation were upregulated during abscission, consistent with the increased abscisic acid concentrations detected by liquid chromatography-mass spectrometry. We performed ex vivo cold shock experiments with excised tree subunits consisting of a branch, pedicel, and fruit. Abscission induction occurred in the cold-stressed subunits with concurrent upregulation of abscisic acid biosynthesis (MdNCED1) and metabolism (MdCYP707A) genes, and ethylene biosynthesis (MdACS1) and receptor (MdETR2) genes in the pedicel. Another key finding was the activation of cytoplasmic streaming in abscission-zone cells detected by electron microscopy. Our results provide a novel insight into the molecular basis of fruit abscission physiology in response to cold stress in apple.

Ascorbic acid and prunasin, two candidate biomarkers for endodormancy release in almond flower buds identified by a nontargeted metabolomic study

Temperate fruit trees belonging to Prunus species have the ability to suspend (induce dormancy) and resume growth periodically in response to environmental and seasonal conditions. Endodormancy release requires the long-term accumulation of chill. Upon accumulation of cultivar-specific chill requirements, plants enter the state of ecodormancy, which means the ability to grow has been restored, depending on the fulfilment of heat requirements. As many different metabolic pathways are implicated in endodormancy release, we have performed a metabolomic analysis, using the ultra-high-performance liquid chromatography–quadrupole time-of-flying (UPLC–QToF) technique. We assayed flower buds in different stages of endodormancy in four almond cultivars with different flowering times: the extra-early Desmayo Largueta, the late Antoñeta, the extra-late Penta, and the ultra-late Tardona. An orthogonal projection to latent-structure discriminant-analysis model was created to observe differences between endodormant and ecodormant flower buds. The metabolites showing the most significant variation were searched against the Metlin, HMDB, and KEGG libraries, which allowed us to identify 87 metabolites. These metabolites were subsequently assigned to specific pathways, such as abscisic acid biosynthesis, phenylpropanoid biosynthesis, and D-sorbitol metabolism, among others. The two metabolites that exhibited the most significant variations in all the cultivars studied with fold changes of up to 6.49 were ascorbic acid and prunasin. For the first time, these two metabolites have been proposed as potential biomarkers for endodormancy release in almond. Given the high synteny present between the Rosaceae species, these results could be extrapolated to other important crops like peach, plum, cherry, or apricot, among others.

Transcriptional Analysis of Carotenoids Accumulation and Metabolism in a Pink-Fleshed Lemon Mutant

Pink lemon is a spontaneous bud mutation of lemon (Citrus limon, L. Burm. f) characterized by the production of pink-fleshed fruits due to an unusual accumulation of lycopene. To elucidate the genetic determinism of the altered pigmentation, comparative carotenoid profiling and transcriptional analysis of both the genes involved in carotenoid precursors and metabolism, and the proteins related to carotenoid-sequestering structures were performed in pink-fleshed lemon and its wild-type. The carotenoid profile of pink lemon pulp is characterized by an increased accumulation of linear carotenoids, such as lycopene, phytoene and phytofluene, from the early stages of development, reaching their maximum in mature green fruits. The distinctive phenotype of pink lemon is associated with an up-regulation and down-regulation of the genes upstream and downstream the lycopene cyclase, respectively. In particular, 9-cis epoxycarotenoid dioxygenase genes were overexpressed in pink lemon compared with the wild-type, suggesting an altered regulation of abscisic acid biosynthesis. Similarly, during early development of the fruits, genes of the carotenoid-associated proteins heat shock protein 21, fibrillin 1 and 2 and orange gene were overexpressed in the pulp of the pink-fleshed lemon compared to the wild-type, indicating its increased capacity for sequestration, stabilization or accumulation of carotenes. Altogether, the results highlighted significant differences at the transcriptomic level between the pink-fleshed lemon and its wild-type, in terms of carotenoid metabolism and the capacity of stabilization in storage structures between the two accessions. Such changes may be either responsible for the altered carotenoid accumulation or in contrast, a metabolic consequence.