Comparative genomics of N-acetyl-5-methoxytryptamine members in four Prunus species with insights into bud dormancy and abiotic stress responses in Prunus avium.

KEY MESSAGE: This study provides novel insights into the evolution, diversification, and functions of melatonin biosynthesis genes in Prunus species, highlighting their potential role in regulating bud dormancy and abiotic stresses. The biosynthesis of melatonin (MEL) in plants is primarily governed by enzymatic reactions involving key enzymes such as serotonin N-acetyltransferase (SNAT), tryptamine 5-hydroxylase (T5H), N-acetylserotonin methyltransferase (ASMT) and tryptophan decarboxylase (TDC). In this study, we analyzed Melatonin genes in four Prunus species such as Prunus avium (Pavi), Prunus pusilliflora (Ppus), Prunus serulata (Pser), and Prunus persica (Pper) based on comparative genomics approach. Among the four Prunus species, a total of 29 TDCs, 998 T5Hs, 16 SNATs, and 115 ASMTs within the genome of four Prunus genomes. A thorough investigation of melatonin-related genes was carried out using systematic biological methods and comparative genomics. Through phylogenetic analysis, orthologous clusters, Go enrichment, syntenic relationship, and gene duplication analysis, we discovered both similarities and variations in Melatonin genes among these Prunus species. Additionally, our study revealed the existence of unique subgroup members in the Melatonin genes of these species, which were distinct from those found in Arabidopsis genes. Furthermore, the transcriptomic expression analysis revealed the potential significance of melatonin genes in bud dormancy regulation and abiotic stresses. Our extensive results offer valuable perspectives on the evolutionary patterns, intricate expansion, and functions of PavMEL genes. Given their promising attributes, PavTDCs, PavT5H, PavNAT, and three PavASMT genes warrant in-depth exploration as prime candidates for manipulating dormancy in sweet cherry. This was done to lay the foundation for future explorations into the structural and functional aspects of these factors in Prunus species. This study offers significant insights into the functions of ASMT, SNAT, T5H, and TDC genes and sheds light on their roles in Prunus avium. Moreover, it established a robust foundation for further exploration functional characterization of melatonin genes in fruit species.

Horticulture crop under pressure: Unraveling the impact of climate change on nutrition and fruit cracking.

Climate change is increasingly affecting the nutritional content and structural integrity of horticultural crops, leading to challenges such as diminished fruit quality and the exacerbation of fruit cracking. This manuscript systematically explores the multifaceted impacts of these changes, with a particular focus on the nutritional quality and increased incidence of fruit cracking. An exhaustive review of current research identifies the critical role of transcription factors in mediating plant responses to climatic stressors, such as drought, temperature extremes, and saline conditions. The significance of transcription factors, including bHLH, bZIP, DOF, MDP, HD-ZIP, MYB, and ERF4, is highlighted in the development of fruit cracking, underscoring the genetic underpinnings behind stress-related phenotypic outcomes. The effectiveness of greenhouse structures in mitigating adverse climatic effects is evaluated, offering a strategic approach to sustain crop productivity amidst CO2 fluctuations and water scarcity, which are shown to influence plant physiology and lead to changes in fruit development, nutrient dynamics, and a heightened risk of cracking. Moreover, the manuscript delves into advanced breeding strategies and genetic engineering techniques, such as genome editing, to enhance crop resilience against climatic challenges. It also discusses adaptation strategies vital for sustainable horticulture, emphasizing the need to integrate novel genetic insights with controlled environment horticulture to counteract climate change's detrimental effects. The synthesis presented here underscores the urgent need for innovative breeding strategies aimed at developing resilient crop varieties that can withstand climatic uncertainty while preserving nutritional integrity.

Global increase in DNA methylation during orange fruit development and ripening.

DNA methylation is an important epigenetic mark involved in many biological processes. The genome of the climacteric tomato fruit undergoes a global loss of DNA methylation due to active DNA demethylation during the ripening process. It is unclear whether the ripening of other fruits is also associated with global DNA demethylation. We characterized the single-base resolution DNA methylomes of sweet orange fruits. Compared with immature orange fruits, ripe orange fruits gained DNA methylation at over 30,000 genomic regions and lost DNA methylation at about 1,000 genomic regions, suggesting a global increase in DNA methylation during orange fruit ripening. This increase in DNA methylation was correlated with decreased expression of DNA demethylase genes. The application of a DNA methylation inhibitor interfered with ripening, indicating that the DNA hypermethylation is critical for the proper ripening of orange fruits. We found that ripening-associated DNA hypermethylation was associated with the repression of several hundred genes, such as photosynthesis genes, and with the activation of hundreds of genes, including genes involved in abscisic acid responses. Our results suggest important roles of DNA methylation in orange fruit ripening.

The mechanism and function of active DNA demethylation in plants.

DNA methylation is a conserved and important epigenetic mark in both mammals and plants. DNA methylation can be dynamically established, maintained, and removed through different pathways. In plants, active DNA demethylation is initiated by the RELEASE OF SILENCING 1 (ROS1) family of bifunctional DNA glycosylases/lyases. Accumulating evidence suggests that DNA demethylation is important in many processes in plants. In this review, we summarize recent studies on the enzymes and regulatory factors that have been identified in the DNA demethylation pathway. We also review the functions of active DNA demethylation in plant development as well as biotic and abiotic stress responses. Finally, we highlight those aspects of DNA demethylation that require additional research.

A DEMETER-like DNA demethylase governs tomato fruit ripening.

In plants, genomic DNA methylation which contributes to development and stress responses can be actively removed by DEMETER-like DNA demethylases (DMLs). Indeed, in Arabidopsis DMLs are important for maternal imprinting and endosperm demethylation, but only a few studies demonstrate the developmental roles of active DNA demethylation conclusively in this plant. Here, we show a direct cause and effect relationship between active DNA demethylation mainly mediated by the tomato DML, SlDML2, and fruit ripening- an important developmental process unique to plants. RNAi SlDML2 knockdown results in ripening inhibition via hypermethylation and repression of the expression of genes encoding ripening transcription factors and rate-limiting enzymes of key biochemical processes such as carotenoid synthesis. Our data demonstrate that active DNA demethylation is central to the control of ripening in tomato.

Unveiling the effect of gibberellin-induced iron oxide nanoparticles on bud dormancy release in sweet cherry (Prunus avium L.).

Hydrogen cyanide has been extensively used worldwide for bud dormancy break in fruit trees, consequently enhancing fruit production via expedited cultivation, especially in areas with controlled environments or warmer regions. A novel and safety nanotechnology was developed since the hazard of hydrogen cyanide for the operators and environments, there is an urgent need for the development of novel and safety approaches to replace it to break bud dormancy for fruit trees. In current study, we have systematically explored the potential of iron oxide nanoparticles, specifically α-Fe2O3, to modulate bud dormancy in sweet cherry (Prunus avium). The synthesized iron oxide nanoparticles underwent meticulous characterization and assessment using various techniques, including Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), transmission electron microscopy (TEM), and ultraviolet-visible infrared (UV-Vis) spectroscopy. Remarkably, when applied at a concentration of 10 mg L-1 alongside gibberellin (GA4+7), these iron oxide nanoparticles exhibited a substantial 57% enhancement in bud dormancy release compared to control groups, all achieved within a remarkably short time span of 4 days. Our RNA-seq analyses further unveiled that 2757 genes within the sweet cherry buds were significantly up-regulated when treated with 10 mg L-1 α-Fe2O3 nanoparticles in combination with GA, while 4748 genes related to dormancy regulation were downregulated in comparison to the control. Moreover, we discovered an array of 58 transcription factor families among the crucial differentially expressed genes (DEGs). Through hormonal quantification, we established that the increased bud burst was accompanied by a reduced concentration of abscisic acid (ABA) at 761.3 ng/g fresh weight in the iron oxide treatment group, coupled with higher levels of gibberellins (GAs) in comparison to the control. Comprehensive transcriptomic and metabolomic analyses unveiled significant alterations in hormone contents and gene expression during the bud dormancy-breaking process when α-Fe2O3 nanoparticles were combined with GA. In conclusion, our findings provide valuable insights into the intricate molecular mechanisms underlying the impact of iron oxide nanoparticles on achieving uniform bud dormancy break in sweet cherry trees.

Chromosome-level genome assembly provides insights into the genetic diversity, evolution, and flower development of Prunus conradinae.

Prunus conradinae, a valuable flowering cherry belonging to the Rosaceae family subgenus Cerasus and endemic to China, has high economic and ornamental value. However, a high-quality P. conradinae genome is unavailable, which hinders our understanding of its genetic relationships and phylogenesis, and ultimately, the possibility of mining of key genes for important traits. Herein, we have successfully assembled a chromosome-scale P. conradinae genome, identifying 31,134 protein-coding genes, with 98.22% of them functionally annotated. Furthermore, we determined that repetitive sequences constitute 46.23% of the genome. Structural variation detection revealed some syntenic regions, inversions, translocations, and duplications, highlighting the genetic diversity and complexity of Cerasus. Phylogenetic analysis demonstrated that P. conradinae is most closely related to P. campanulata, from which it diverged ~ 19.1 million years ago (Mya). P. avium diverged earlier than P. cerasus and P. conradinae. Similar to the other Prunus species, P. conradinae underwent a common whole-genome duplication event at ~ 138.60 Mya. Furthermore, 79 MADS-box members were identified in P. conradinae, accompanied by the expansion of the SHORT VEGETATIVE PHASE subfamily. Our findings shed light on the complex genetic relationships, and genome evolution of P. conradinae and will facilitate research on the molecular breeding and functions of key genes related to important horticultural and economic characteristics of subgenus Cerasus.

DNA cytosine methylation dynamics and functional roles in horticultural crops.

Methylation of cytosine is a conserved epigenetic modification that maintains the dynamic balance of methylation in plants under the regulation of methyltransferases and demethylases. In recent years, the study of DNA methylation in regulating the growth and development of plants and animals has become a key area of research. This review describes the regulatory mechanisms of DNA cytosine methylation in plants. It summarizes studies on epigenetic modifications of DNA methylation in fruit ripening, development, senescence, plant height, organ size, and under biotic and abiotic stresses in horticultural crops. The review provides a theoretical basis for understanding the mechanisms of DNA methylation and their relevance to breeding, genetic improvement, research, innovation, and exploitation of new cultivars of horticultural crops.

Oxygenation alleviates waterlogging-caused damages to cherry rootstocks.

Waterlogging has occurred more frequently in recent years due to climate change, so it is a huge threat to crop yield and quality. Sweet cherry, a fruit tree with a high economic value, is sensitive to waterlogging stress. One of the most effective methods for enhancing the waterlogging tolerance of sweet cherries is to select waterlogging-tolerant rootstocks. However, the waterlogging tolerance of different cherry rootstocks, and the underlying mechanism remains uncharacterized. Thus, we first evaluated the waterlogging resistance of five sweet cherry rootstocks planted in China. The data showed that 'Gisela 12' and 'Colt' were the most waterlogging-sensitive and -tolerant among the five tested varieties, respectively. Oxygenation effectively alleviated the adverse impacts of waterlogging stress on cherry rootstocks. Moreover, we found that the waterlogging group had lower relative water content, Fv/Fm value, net photosynthetic rate, and higher antioxidant enzyme activities, whereas the oxygenated group performed better in all these parameters. RNA-Seq analysis revealed that numerous DEGs were involved in energy production, antioxidant metabolism, hormone metabolism pathways, and stress-related transcription factors. These findings will help provide management strategies to enhance the waterlogging tolerance of cherry rootstocks and thereby achieve higher yield and better quality of cherries.

Chromosome-scale genome assembly of Prunus pusilliflora provides novel insights into genome evolution, disease resistance, and dormancy release in Cerasus L.

Prunus pusilliflora is a wild cherry germplasm resource distributed mainly in Southwest China. Despite its ornamental and economic value, a high-quality assembled P. pusilliflora genome is unavailable, hindering our understanding of its genetic background, population diversity, and evolutionary processes. Here, we de novo assembled a chromosome-scale P. pusilliflora genome using Oxford Nanopore, Illumina, and chromosome conformation capture sequencing. The assembled genome size was 309.62 Mb, with 76 scaffolds anchored to eight pseudochromosomes. We predicted 33 035 protein-coding genes, functionally annotated 98.27% of them, and identified repetitive sequences covering 49.08% of the genome. We found that P. pusilliflora is closely related to Prunus serrulata and Prunus yedoensis, having diverged from them ~41.8 million years ago. A comparative genomic analysis revealed that P. pusilliflora has 643 expanded and 1128 contracted gene families. Furthermore, we found that P. pusilliflora is more resistant to Colletotrichum viniferum, Phytophthora capsici, and Pseudomonas syringae pv. tomato (Pst) DC3000 infections than cultivated Prunus avium. P. pusilliflora also has considerably more nucleotide-binding site-type resistance gene analogs than P. avium, which explains its stronger disease resistance. The cytochrome P450 and WRKY families of 263 and 61 proteins were divided into 42 and 8 subfamilies respectively in P. pusilliflora. Furthermore, 81 MADS-box genes were identified in P. pusilliflora, accompanying expansions of the SVP and AGL15 subfamilies and loss of the TM3 subfamily. Our assembly of a high-quality P. pusilliflora genome will be valuable for further research on cherries and molecular breeding.

Transcriptome Analysis of Eggplant under Salt Stress: AP2/ERF Transcription Factor SmERF1 Acts as a Positive Regulator of Salt Stress.

Salt stress, a type of abiotic stress, impedes plant growth and development and strongly reduces crop yield. The molecular mechanisms underlying plant responses to salt stress remain largely unclear. To characterize the enriched pathways and genes that were affected during salt treatment, we performed mRNA sequencing (mRNA-seq) in eggplant roots and identified 8509 differentially expressed genes (DEGs) between the mock and 24 h under salt stress. Among these DEGs, we found that the AP2/ERF transcription factor family member SmERF1 belongs to the plant-pathogen interaction pathway, which was significantly upregulated by salt stress. We found that SmERF1 localizes in the nuclei with transcriptional activity. The results of the virus-induced gene silencing assay showed that SmERF1 silencing markedly enhanced the susceptibility of plants to salt stress, significantly downregulated the transcript expression levels of salt stress defense-related marker genes (9-cis-epoxycarotenoid dioxygenase [SmNCED1, SmNCED2], Dehydrin [SmDHN1], and Dehydrin (SmDHNX1), and reduced the activity of superoxide dismutase and catalase. Silencing SmERF1 promoted the generation of H2O2 and proline. In addition, the transient overexpression of SmERF1 triggered intense cell death in eggplant leaves, as assessed by the darker diaminobenzidine and trypan blue staining. These findings suggest that SmERF1 acts as a positive regulator of eggplant response to salt stress. Hence, our results suggest that AP2/ERF transcription factors play a vital role in the response to salt stress.

GhBEE3-Like gene regulated by brassinosteroids is involved in cotton drought tolerance.

Brassinosteroids (BRs) are important phytohormones that play a vital role in plant drought tolerance, but their mechanisms in cotton (Gossypium hirsutum L.) are poorly understood. Numerous basic helix-loop-helix (bHLH) family genes are involved in the responses to both BRs and drought stress. GhBEE3-Like, a bHLH transcription factor, is repressed by both 24-epi-BL (an active BR substance) and PEG8000 (drought simulation) treatments in cotton. Moreover, GhBZR1, a crucial transcription factor in BR signaling pathway, directly binds to the E-box element in GhBEE3-Like promoter region and inhibits its expression, which has been confirmed by electrophoretic mobility shift assay (EMSA) and dual luciferase reporter assay. Functional analysis revealed that Arabidopsis with GhBEE3-Like overexpression had drought sensitive phenotype, while GhBEE3-Like knock-down cotton plants obtained by virus-induced gene silencing (VIGS) technology were more tolerant to drought stress. Furthermore, the expression levels of three stress-related genes, GhERD10, GhCDPK1 and GhRD26, were significantly higher in GhBEE3-Like knock-down cotton than in control cotton after drought treatment. These results suggest that GhBEE3-Like is inhibited by BRs which elevates the expressions of stress-related genes to enhance plant drought tolerance. This study lays the foundation for understanding the mechanisms of BR-regulated drought tolerance and establishment of drought-resistant cotton lines.

Cell suspension culture-mediated incorporation of the rice bel gene into transgenic cotton.

Cotton plants engineered for resistance to the herbicides, glyphosate or glufosinate have made a considerable impact on the production of the crop worldwide. In this work, embryogenic cell cultures derived from Gossypium hirsutum L. cv Coker 312 hypocotyl callus were transformed via Agrobacterium tumefaciens with the rice cytochrome P450 gene, CYP81A6 (bel). In rice, bel has been shown to confer resistance to both bentazon and sulfanylurea herbicides. Transformed cells were selected on a liquid medium supplemented alternately or simultaneously with kanamycin (50mg/L) and bentazon (4.2 µmol). A total of 17 transgenic cotton lines were recovered, based on the initial resistance to bentazon and on PCR detection of the bel transgene. Bel integration into the cotton genome was confirmed by Southern blot and expression of the transgene was verified by RT-PCR. In greenhouse and experimental plot trials, herbicide (bentazon) tolerance of up to 1250 mg/L was demonstrated in the transgenic plants. Transgenic lines with a single copy of the bel gene showed normal Mendelian inheritance of the characteristic. Importantly, resistance to bentazon was shown to be stably incorporated in the T1, T2 and T3 generations of self-fertilised descendents and in plants outcrossed to another upland cotton cultivar. Engineering resistance to bentazon in cotton through the heterologous expression of bel opens the possibility of incorporating this trait into elite cultivars, a strategy that would give growers a more flexible alternative to weed management in cotton crops.