Ethylene Signal Transduction Pathway Research Articles

Genome-Wide Identification and Analysis of the EIN3/EIL Transcription Factor Gene Family in Doubled Haploid (DH) Poplar.

Ethylene (ET) is an important phytohormone that regulates plant growth, development and stress responses. The ethylene-insensitive3/ethylene-insensitive3-like (EIN3/EIL) transcription factor family, as a key regulator of the ET signal transduction pathway, plays an important role in regulating the expression of ET-responsive genes. Although studies of EIN3/EIL family members have been completed in many species, their role in doubled haploid (DH) poplar derived from another culture of diploid Populus simonii × P. nigra (donor tree, DT) remains ambiguous. In this study, a total of seven EIN3/EIL gene family members in the DH poplar genome were identified. Basic physical and chemical property analyses of these genes were performed, and these proteins were predicted to be localized to the nucleus. According to the phylogenetic relationship, EIN3/EIL genes were divided into two groups, and the genes in the same group had a similar gene structure and conserved motifs. The expression patterns of EIN3/EIL genes in the apical buds of different DH poplar plants were analyzed based on transcriptome data. At the same time, the expression patterns of PsnEIL1, PsnEIN3, PsnEIL4 and PsnEIL5 genes in different tissues of different DH plants were detected via RT-qPCR, including the apical buds, young leaves, functional leaves, xylem, cambium and roots. The findings presented above indicate notable variations in the expression levels of PsnEIL genes across various tissues of distinct DH plants. Finally, the PsnEIL1 gene was overexpressed in DT, and the transgenic plants showed a dwarf phenotype, indicating that the PsnEIL1 gene was involved in regulating the growth and development of poplar. In this study, the EIN3/EIL gene family of DH poplar was analyzed and functionally characterized, which provides a theoretical basis for the future exploration of the EIN3/EIL gene function.

Transcriptomic analysis to elucidate the molecular mechanisms underlying the growth advantage of Kluyveromyces marxianus XZ1 and the development of resistance in apple induced by XZ1

The preceding study demonstrated that Kluyveromyces marxianus XZ1 (XZ1) exhibits potential as a biological agent for combating disease by P. expansum in apple fruit. This potential is primarily attributed to its capacity for competing for resources and space, as well as activating certain genes involved in the transduction of plant hormone signals. In this study, transcriptomic analysis was performed on the interaction system of XZ1 and apple to investigate the detail molecular mechanisms of the yeast to develop competitive advantage and the molecular mechanisms of the apple to develop defense response action at the gene expression level. The transcriptome analysis of XZ1 in apple indicated that the meiosis pathway was activated to promote rapid proliferation of the yeast in apple. Autophagy, ubiquitin-mediated proteolysis, and the starch and sucrose metabolism pathway are induced to generate vital nutrients and energy for yeast proliferation. The transcriptome analysis of apple revealed upregulation in the expression levels of genes associated with transcription factors (TFS) involved in pattern-triggered immunity (PTI) and effector-triggered immunity (ETI) following treatment with XZ1. This finding suggests that XZ1 activates PTI and ETI in apple, thereby promoting disease resistance. Moreover, the yeast treatment induced the activation of ethylene and jasmonate signal transduction pathways, which play a crucial role in regulating disease resistance development in apple. Additionally, the yeast treatment resulted in the enhancement of lignin biosynthesis, serving as a protective mechanism for the apple against potential threats, as observed in the response to XZ1 treatment. The findings of this study establish a theoretical foundation for elucidating the molecular mechanism underlying the growth advantage of antagonistic yeast in the host and the induced resistance in apple. Additionally, this study offers a substantial repertoire of gene resources that can be utilized to enhance yeast biocontrol efficacy and bolster the disease resistance of apple through the application of molecular biology techniques.

OsJMJ718, a histone demethylase gene, positively regulates seed germination in rice.

Seed vigor has major impact on the rate and uniformity of seedling growth, crop yield, and quality. However, the epigenetic regulatory mechanism of crop seed vigor remains unclear. In this study, a (jumonji C) JmjC gene of the histone lysine demethylase OsJMJ718 was cloned in rice, and its roles in seed germination and its epigenetic regulation mechanism were investigated. OsJMJ718 was located in the nucleus and was engaged in H3K9 methylation. Histochemical GUS staining analysis revealed OsJMJ718 was highly expressed in seed embryos. Abiotic stress strongly induced the OsJMJ718 transcriptional accumulation level. Germination percentage and seedling vigor index of OsJMJ718 knockout lines (OsJMJ718-CR) were lower than those of the wild type (WT). Chromatin immunoprecipitation followed by sequencing (ChIP-seq) of seeds imbibed for 24 h showed an increase in H3K9me3 deposition of thousands of genes in OsJMJ718-CR. ChIP-seq results and transcriptome analysis showed that differentially expressed genes were enriched in ABA and ethylene signal transduction pathways. The content of ABA in OsJMJ718-CR was higher than that in WT seeds. OsJMJ718 overexpression enhanced sensitivity to ABA during germination and early seedling growth. In the seed imbibition stage, ABA and ethylene content diminished and augmented, separately, suggesting that OsJMJ718 may adjust rice seed germination through the ABA and ethylene signal transduction pathways. This study displayed the important function of OsJMJ718 in adjusting rice seed germination and vigor, which will provide an essential reference for practical issues, such as improving rice vigor and promoting direct rice sowing production.

Biocontrol fungi induced stem-base rot disease resistance of Morinda officinalis How revealed by transcriptome analysis.

Morinda officinalis How (MO) is a Rubiaceae plant, and its medicinal part is dried root, which is one of the "Four Southern Medicines" in China. At present, the plant MO breed seedlings mainly by cutting methods. Long-term asexual propagation makes pathogenic fungi accumulate in MO, leading to stem-base rot, which is caused by Fusarium oxysporum (Fon). In this study, we used Trichoderma harzianum and Pestalotiopsis sp. as biocontrol fungi to investigate their antagonistic ability to Fon through in vitro antagonism and pot experiments, and combined with transcriptome sequencing to explore the mechanism of biocontrol. The results showed that both Trichoderma harzianum and Pestalotiopsis sp. could inhibit the growth of Fon. In addition, Trichoderma harzianum and Pestalotiopsis sp. could also enhance the basic immunity to Fon by increasing the activities of defensive enzymes such as POD and SOD, chlorophyll content, soluble sugar content, and oligosaccharide content of MO. The mechanism of biological control of stem-base rot of MO was discussed by transcriptome technology. MO was treated with two treatments, root irrigation with biocontrol fungi or inoculation with Fon after root irrigation with biocontrol fungi. Transcriptome sequencing revealed that nearly 11,188 differentially expressed genes (DEGs) were involved in the process of inducing MO systemic resistance to Fon by biocontrol fungi. Meanwhile, Gene Ontology (GO) classification and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, as well as transcription factor (TFs) prediction showed that there were significant differences in the expression levels of MO roots under different treatments. Also, the genes of the "MAPK signaling pathway" and "plant hormone signaling pathway" were analyzed, in which the ERFs gene of the ethylene signal transduction pathway participated in the metabolism of glycosyl compounds. It is speculated that the ethylene signal may participate in the immune response of the sugar signal to the infection of Fon. After qRT-PCR verification of 10 DEGs related to the ethylene signal transduction pathway, the expression trend is consistent with the results of transcriptome sequencing, which proves the reliability of transcriptome sequencing. In conclusion, this study preliminarily identified the molecular mechanism of the biological control of MO stem-base rot and provided a scientific basis for further research on the prevention and control mechanism of MO stem-base rot.

New insights into the occurrence of continuous cropping obstacles in pea (Pisum sativum L.) from soil bacterial communities, root metabolism and gene transcription

BackgroundContinuous cropping is a significant obstacle to sustainable development in the pea (Pisum sativum L.) industry, but the underlying mechanisms of this remain unclear. In this study, we used 16 S rDNA sequencing, transcriptomics, and metabolomics to analyze the response mechanism of roots and soil bacteria to continuous cropping and the relationship between soil bacteria and root phenotypes of different pea genotypes (Ding wan 10 and Yun wan 8).ResultsContinuous cropping inhibited pea growth, with a greater effect on Ding wan 10 than Yun wan 8. Metabolomics showed that the number of differentially accumulated metabolites (DAMs) in pea roots increased with the number of continuous cropping, and more metabolic pathways were involved. Transcriptomics revealed that the number of differentially expressed genes (DEGs) increased with the number of continuous cropping. Continuous cropping altered the expression of genes involved in plant-pathogen interaction, MAPK signal transduction, and lignin synthesis pathways in pea roots, with more DEGs in Ding wan 10 than in Yun wan 8. The up-regulated expression of genes in the ethylene signal transduction pathway was evident in Ding wan 10. Soil bacterial diversity did not change, but the relative abundance of bacteria significantly responded to continuous cropping. Integrative analysis showed that the bacteria with significant relative abundance in the soil were strongly associated with the antioxidant synthesis and linoleic acid metabolism pathway of pea roots under continuous cropping once. Under continuous cropping twice, the bacteria with significant relative abundance changes were strongly associated with cysteine and methionine metabolism, fatty acid metabolism, phenylpropanoid biosynthesis, terpenoid backbone biosynthesis, linoleic acid, and amino sugar and nucleotide sugar metabolism.ConclusionDing wan 10 was more sensitive to continuous cropping than Yun wan 8. Continuous cropping times and pea genotypes determined the differences in root metabolic pathways. There were common metabolic pathways in the two pea genotypes in response to continuous cropping, and the DEGs and DAMs in these metabolic pathways were strongly associated with the bacteria with significant changes in relative abundance in the soil. This study provides new insights into obstacles to continuous cropping in peas.

Regulation and mechanism of ethylene treatment on storage quality of fresh-cut lotus (Nelumbo nucifera Gaertn.) root slices

This study investigated the effects of 1 g L−1 ethylene immersion treatment for 1 min on the storage quality, antioxidant capacity and ethylene signal transduction pathway of fresh-cut lotus roots. Phenylalanine ammonia-lyase and polyphenol oxidase activities were increased in ethylene treatment slices, and total phenolic content was increased. The activities of peroxidase, catalase and superoxide dismutase were increased after ethylene treatment, as well as the expression of related genes (NnPOD, NnSOD, NnCAT) was upregulated. The rate of superoxide anion radical and hydroxyl radical production in ethylene treatment fresh-cut lotus roots was inhibited, the hydrogen peroxide content was reduced, and the degree of membrane lipid peroxidation was decreased, which delayed browning during the early storage. However, ethylene treatment inhibited microbial growth and improved 1,1-Diphenyl-2-picrylhydrazyl scavenging capacity only during the early storage, promoted respiration and accelerated the decline of hardness and soluble solids content throughout the storage period. At the molecular level, ethylene treatment overall upregulated the transcription of NnmetK, NnACO, NnETR, NnEIN3, and NnERF genes and promoted ethylene biosynthesis and signal transduction. This finding may be related to the senescence of fresh-cut lotus roots during late storage. These results suggest that ethylene treatment improved the antioxidant system, reduced the accumulation of reactive oxygen species, and effectively delayed the early browning of fresh-cut lotus roots, while promoting late senescence.

Comparative transcriptome analysis to reveal key ethylene genes involved in a Lonicera macranthoides mutant.

Lonicera macranthoides Hand.-Mazz. is an important medicinal plant. Xianglei-type (XL) L. macranthoides was formed after many years of cultivation by researchers on the basis of the natural mutant. The corolla of L. macranthoides XL remains unexpanded and its flowering period is nearly three times longer than that of wild-type (WT) plants. However, the molecular mechanism behind this desirable trait remains a mystery. To understand the floral phenotype differences between L. macranthoides and L. macranthoides XL at the molecular level. Transcriptome analysis was performed on L. macranthoides XL and WT. One DEG was cloned by RT-PCR amplification and selected for qRT-PCR analysis. Transcriptome analysis showed that there were 5603 differentially expressed genes (DEGs) in XL vs. WT. Enrichment analysis of DEGs showed that pathways related to plant hormone signal transduction were significantly enriched. We identified 23 key genes in ethylene biosynthesis and signal transduction pathways. The most abundant were the ethylene biosynthesis DEGs. In addition, the open reading frames (ORFs) of WT and XL ETR2 were successfully cloned and named LM-ETR2 (GenBank: MW334978) and LM-XL-ETR2 (GenBank: MW334978), respectively. qRT-PCR at different flowering stages suggesting that ETR2 acts in the whole stage of flower development of WT and XL. This study provides new insight into the molecular mechanism that regulates the development of special traits in the flowers of L. macranthoides XL. The plant hormone ethylene plays an important role in flower development and flowering duration prolongation in L. macranthoides. The ethylene synthesis gene could be more responsible for the flower phenotype of XL. The genes identified here can be used for breeding and improvement of other flowering plants after functional verification.

Integrated transcriptomic and metabolic analyses provide insights into the maintenance of embryogenic potential and the biosynthesis of phenolic acids and flavonoids involving transcription factors in Larix kaempferi (Lamb.) Carr.

Somatic embryogenesis (SE) techniques have been established for micropropagation or basic research related to plant development in many conifer species. The frequent occurrence of non-embryogenic callus (NEC) during SE has impose constraints on the application of somatic embryogenesis SE in Larix kaempferi (Lamb.) Carr, but the potential regulatory mechanisms are poorly understood. In this study, integrated transcriptomic and metabolomic analyses were performed in embryogenic callus (EC) and NEC originating from a single immature zygotic embryo to better decipher the key molecular and metabolic mechanisms required for embryogenic potential maintenance. The results showed that a total of 13,842 differentially expressed genes (DEGs) were found in EC and NEC, among which many were enriched in plant hormone signal transduction, starch and sucrose metabolism, phenylpropanoid biosynthesis, flavonoid biosynthesis, and the biosynthesis of amino acids pathways. Metabolite profiling showed that 441 differentially accumulated metabolites (DAMs) were identified in EC and NEC. Both EC and NEC had vigorous primary metabolic activities, while most secondary metabolites were upregulated in NEC. Many totipotency-related transcription factor (TF) genes such as BBMs, WUSs, and LEC1 showed higher expression levels in EC compared with NEC, which may result in the higher accumulation of indole 3-acetic acid (IAA) in EC. NEC was characterized by upregulation of genes and metabolites associated with stress responses, such as DEGs involved in jasmonic acid (JA) and ethylene (ETH) biosynthesis and signal transduction pathways, and DEGs and DAMs related to phenylpropanoid and flavonoid biosynthesis. We predicted and analyzed TFs that could target several key co-expressed structural DEGs including two C4H genes, two CcoAOMT genes and three HCT genes involved in phenylpropanoid and flavonoid biosynthesis. Based on the targeted relationship and the co-expression network, two ERFs (Lk23436 and Lk458687), one MYB (Lk34626) and one C2C2-dof (Lk37167) may play an important role in regulating phenolic acid and flavonoid biosynthesis by transcriptionally regulating the expression of these structural genes. This study shows an approach involving integrated transcriptomic and metabolic analyses to obtain insights into molecular events underlying embryogenic potential maintenance and the biosynthesis mechanisms of key metabolites involving TF regulation, which provides valuable information for the improvement of SE efficiency in L. kaempferi.

Digital gene expression profiling of the transcriptional response to Sclerotinia sclerotiorum and its antagonistic bacterium Bacillus amyloliquefaciens in soybean.

Soybean Sclerotinia stem rot caused by Sclerotinia sclerotiorum is a common disease in soybean, and effective biological control is urgently needed. We have previously confirmed that Bacillus amyloliquefaciens can effectively antagonize S. sclerotiorum in a plate competition experiment and a soybean seedling inoculation experiment. In this study, the mechanisms underlying plant death caused by S. sclerotiorum and soybean resistance to S. sclerotiorum induced by B. amyloliquefaciens were evaluated. The stems of potted soybean seedlings were inoculated with S. sclerotiorum (Gm-Ss), B. amyloliquefaciens (Gm-Ba), and their combination (Gm-Ba-Ss), using scratch treatments as a control, followed by dual RNA sequencing and bioinformatics analyses. Global gene expression levels in the Gm-Ss treatment were much lower than those in the Gm-Ba, Gm-Ba-Ss, and Gm groups, suggesting that S. sclerotiorum strongly inhibited global gene expression in soybean. In a pairwise comparison of Gm-Ss vs. Gm, 19983 differentially expressed genes (DEGs) were identified. Down-regulated DEGs were involved in various KEGG pathways, including ko01110 (biosynthesis of secondary metabolites), ko01100 (metabolic pathways), ko01120 (microbial metabolism in diverse environments), ko00500 (starch and sucrose metabolism), and ko04075 (plant hormone signal transmission), suggesting that S. sclerotiorum inoculation had a serious negative effect on soybean metabolism. In Gm-Ba vs. Gm, 13091 DEGs were identified, and these DEGs were significantly enriched in ko03010 (ribosome) and ko03008 (ribosome biogenesis in eucaryotes). Our results suggest that B. amyloliquefaciens increases the expression of genes encoding the ribosomal subunit, promotes cell wall biogenesis, and induces systemic resistance. S. sclerotiorum strongly inhibited metabolism in soybean, inhibited the synthesis of the cytoskeleton, and induced the up-regulation of programmed death and senescence-related genes via an ethylene signal transduction pathway. These results improve our understanding of S. sclerotiorum-induced plant death and soybean resistance to S. sclerotiorum induced by B. amyloliquefaciens and may contribute to the improvement of strategies to avoid yield losses.

The molecular mechanism on suppression of climacteric fruit ripening with postharvest wax coating treatment via transcriptome.

Wax coating is an important means to maintain fruit quality and extend fruit shelf life, especially for climacteric fruits, such as apples (Malus domestica). Here, we found that wax coating could inhibit ethylene production, chlorophyll degradation, and carotenoid synthesis, but the molecular mechanism remains unclear. The regulatory mechanism of wax coating on apple fruit ripening was determined by subjecting wax-treated apple fruits to transcriptome analysis. RNA-seq revealed that 1,137 and 1,398 genes were upregulated and downregulated, respectively. These differentially expressed genes (DEGs) were shown to be related to plant hormones, such as ethylene, auxin, abscisic acid, and gibberellin, as well as genes involved in chlorophyll degradation and carotenoid biosynthesis. Moreover, we found that some genes related to the wax synthesis process also showed differential expression after the wax coating treatment. Among the DEGs obtained from RNA-seq analysis, 15 were validated by quantitative RT-PCR, confirming the results from RNA-seq analysis. RNA-seq and qRT-PCR of pear (Pyrus ussuriensis) showed similar changes after wax treatment. Our data suggest that wax coating treatment inhibits fruit ripening through ethylene synthesis and signal transduction, chlorophyll metabolism, and carotenoid synthesis pathways and that waxing inhibits endogenous wax production. These results provide new insights into the inhibition of fruit ripening by wax coating.

Physiological, Transcriptomic and Metabolomic Analyses of Overwintering Cryptomeria fortunei Needles

Low temperatures affect plant growth, development, and geographical distribution. Cryptomeriafortunei (Chinese cedar) is a major industrial tree species used for timber manufacturing in southern China. However, its popularization and application in northern China are limited due to its poor low-temperature resistance (approximately −6 °C), and its overwintering mechanism remains unclear. Here, we performed physiological, metabolomic, and transcriptomic analyses of overwintering C.fortunei needles at three stages: before winter, during winter, and in early spring. Physiological analyses showed that electrolyte leakage, H2O2, malondialdehyde (MDA), soluble sugar, and protein contents increased, while the maximum quantum yield of photosystem II (PSII) (Fv/Fm), effective quantum yield of PSII (YII), and chlorophyll content decreased as overwintering progressed. Metabolomic and transcriptomic analyses revealed that downregulated gibberellin (GA), salicylic acid (SA), cytokinin (CTK), and auxin signal transduction and upregulated abscisic acid (ABA), ethylene, brassinosteroids (BR), and jasmonic acid (JA) signal transduction pathways promoted the winter acclimation of C.fortunei, while the opposite expression pattern promoted the transition from dormancy to growth. In addition, upregulated genes/metabolites involved in phenylpropanoid and flavonoid biosynthesis, starch and sucrose metabolism, cold-related protein and fatty acid desaturases, and downregulated photosynthesis-related pathways promoted winter acclimation, while five (WRKY, AP2/ERF, NAC, MYB, and bHLH) and three (AP2/ERF, MYB, and bHLH) transcription factors were associated with winter acclimation and early spring transition from dormancy to growth, respectively. In summary, we report the first transcriptome of overwintering C.fortunei, providing a foundation for the cultivation of and research on overwintering varieties.

Comparative Transcriptome Analysis of Deep-Rooting and Shallow-Rooting Potato (Solanum tuberosum L.) Genotypes under Drought Stress.

The selection and breeding of deep rooting and drought-tolerant varieties has become a promising approach for improving the yield and adaptability of potato (Solanum tuberosum L.) in arid and semiarid areas. Therefore, the discovery of root-development-related genes and drought tolerance signaling pathways in potato is important. In this study, we used deep-rooting (C119) and shallow-rooting (C16) potato genotypes, with different levels of drought tolerance, to achieve this objective. Both genotypes were treated with 150 mM mannitol for 0 h (T0), 2 h (T2), 6 h (T6), 12 h (T12), and 24 h (T24), and their root tissues were subjected to comparative transcriptome analysis. A total of 531, 1571, 1247, and 3540 differentially expressed genes (DEGs) in C16 and 1531, 1108, 674, and 4850 DEGs in C119 were identified in T2 vs. T0, T6 vs. T2, T12 vs. T6, and T24 vs. T12 comparisons, respectively. Gene expression analysis indicated that a delay in the onset of drought-induced transcriptional changes in C16 compared with C119. Functional enrichment analysis revealed genotype-specific biological processes involved in drought stress tolerance. The metabolic pathways of plant hormone transduction and MAPK signaling were heavily involved in the resistance of C16 and C119 to drought, while abscisic acid (ABA), ethylene, and salicylic acid signal transduction pathways likely played more important roles in C119 stress responses. Furthermore, genes involved in root cell elongation and division showed differential expression between the two genotypes under drought stress. Overall, this study provides important information for the marker-assisted selection and breeding of drought-tolerant potato genotypes.

Identification of candidate genes and clarification of the maintenance of the green pericarp of weedy rice grains.

The weedy rice (Oryza sativa f. spontanea) pericarp has diverse colors (e.g., purple, red, light-red, and white). However, research on pericarp colors has focused on red and purple, but not green. Unlike many other common weedy rice resources, LM8 has a green pericarp at maturity. In this study, the coloration of the LM8 pericarp was evaluated at the cellular and genetic levels. First, an examination of their ultrastructure indicated that LM8 chloroplasts were normal regarding plastid development and they contained many plastoglobules from the early immature stage to maturity. Analyses of transcriptome profiles and differentially expressed genes revealed that most chlorophyll (Chl) degradation-related genes in LM8 were expressed at lower levels than Chl a/b cycle-related genes in mature pericarps, suggesting that the green LM8 pericarp was associated with inhibited Chl degradation in intact chloroplasts. Second, the F2 generation derived from a cross between LM8 (green pericarp) and SLG (white pericarp) had a pericarp color segregation ratio of 9:3:4 (green:brown:white). The bulked segregant analysis of the F2 populations resulted in the identification of 12 known genes in the chromosome 3 and 4 hotspot regions as candidate genes related to Chl metabolism in the rice pericarp. The RNA-seq and sqRT-PCR assays indicated that the expression of the Chl a/b cycle-related structural gene DVR (encoding divinyl reductase) was sharply up-regulated. Moreover, genes encoding magnesium-chelatase subunit D and the light-harvesting Chl a/b-binding protein were transcriptionally active in the fully ripened dry pericarp. Regarding the ethylene signal transduction pathway, the CTR (encoding an ethylene-responsive protein kinase) and ERF (encoding an ethylene-responsive factor) genes expression profiles were determined. The findings of this study highlight the regulatory roles of Chl biosynthesis- and degradation-related genes influencing Chl accumulation during the maturation of the LM8 pericarp.

Transcriptomics integrated with metabolomics reveals underlying mechanisms of cold-induced flesh bleeding in plum (cv. Friar) fruit during storage

An integrative analysis of the metabolomics and transcriptomics was conducted to reveal the underlying mechanisms responsible for the flesh bleeding of ‘Friar’ plum (Prunus salicina Lindl.) fruit subjected to cold storage. Results from quasi-targeted metabolomics showed that 115 differentially accumulated metabolites were detected and cyanidin derivatives were identified as the main components in red flesh. The accumulation of cyanidin-3-O-glucoside was further quantitatively proved using UPLC-MS. A weighted gene co-correlation network analysis was employed to obtain genes positively associated with flesh bleeding according to the comprehensive transcriptomic profiling, which was verified by Pearson’s correlation coefficient analysis. 21 differentially expressed genes were identified in the biosynthesis of anthocyanins, and 79 overlap up-regulated genes of transcription factors were identified as candidate regulators contributing to flesh bleeding in the fruit in response to cold stress. The up-regulation of genes involving in the MAPK cascade signal system participated in the ethylene signal transduction pathway under cold stress, which stimulates transcription factors and ultimately leads to the activation of structural genes associated with the biosynthesis of anthocyanins. The results of qRT-PCR verified the reliability of transcriptome data. These results would provide a new insight on the mechanism in flesh bleeding in plums during cold storage.

Characterization, expression, and functional analysis of the pathogenesis-related gene PtDIR11 in transgenic poplar

Lignins and lignans are important for plant resistance to pathogens. Dirigent (DIR) proteins control the regio- and stereo-selectivity of coniferyl alcohol in lignan and lignin biosynthesis. DIR genes have been implicated in defense-related responses in several plant species, but their role in poplar immunity is unclear. We cloned PtDIR11 from Populus trichocarpa; we found that overexpression of PtDIR11 in poplar improved the lignan biosynthesis and enhanced the resistance of poplar to Septotis populiperda. PtDIR11 has a typical DIR domain; it belongs to the DIR-b/d family and is expressed in the cell membrane. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis showed that PtDIR11 expression was highest in stems, followed by leaves and roots. Furthermore, PtDIR11 expression was induced by S. populiperda, salicylic acid (SA), jasmonate (JA), and ethylene (ET) stresses. The recombinant PtDIR11 protein inhibited the growth of S. populiperda in vitro. Overexpressing (OE) PtDIR11 in “Nanlin 895” poplar enhanced growth. The OE lines exhibited minimal changes in lignin content, but their total lignan and flavonoid contents were significantly greater than in the wild-type (WT) lines. Overexpression of PtDIR11 affected multiple biological pathways of poplar, such as phenylpropanoid biosynthesis. The methanol extracts of OE-PtDIR11 lines showed greater anti-S. populiperda activity than did lignin extracts from the WT lines. Furthermore, OE-PtDIR11 lines upregulated genes that were related to phenylpropanoid biosynthesis and genes associated with the JA and ET signal transduction pathways; it downregulated genes that were related to SA signal transduction compared with the WT line under S. populiperda stress. Therefore, the OE transgenic plants analysis revealed that PtDIR11 is a good candidate gene for breeding of disease resistant poplar.

Ethylene and polyamines form a negative feedback loop to regulate peach fruit ripening via the transcription factor PpeERF113 by regulating the expression of PpePAO1

Ethylene and polyamines (PAs) have antagonistic roles during fruit ripening. PAs affect both ethylene biosynthetic and signal transduction pathways, but the regulatory effects of ethylene on PA metabolism during fruit ripening have remained largely unknown. Here, we found that ethylene enhanced PA catabolism by regulating expression of a peach PA oxidase gene, PpePAO1. Knocking out PpePAO1 delayed peach fruit ripening, whereas PpePAO1 overexpression promoted tomato fruit ripening. Both in vitro and in vivo assays demonstrated that an ethylene response factor (ERF), PpeERF113, activated PpePAO1 by direct binding to the promoter. PA levels were significantly increased and ripening was delayed in PpeERF113 knockdown peach fruit. Correspondingly, PA levels were significantly decreased when PpeERF113 was overexpressed in peach. PpeERF113 was also found to be upregulated in PpePAO1 knockdowns, indicating that PA catabolism has a regulatory function in ethylene signal transduction. Together, our results suggest that ethylene accelerates peach fruit ripening in part by promoting PA catabolism through PpeERF113-mediated regulation of PpePAO1 expression. To our knowledge, this is the first report related to the regulatory mechanism of ethylene in PA catabolism during fruit ripening. This study contributes new understanding of the crosstalk between ethylene and PAs during fruit ripening.

Effect of Glycine Betaine Treatment on Aroma Production of ‘Nanguo’ Pears After Long-Term Cold Storage–Possible Involvement of Ethylene Synthesis and Signal Transduction Pathways

Effect of Glycine Betaine Treatment on Aroma Production of ‘Nanguo’ Pears After Long-Term Cold Storage–Possible Involvement of Ethylene Synthesis and Signal Transduction Pathways

Tamarix hispida ThEIL1 improves salt tolerance by adjusting osmotic potential and increasing reactive oxygen species scavenging capability

The ETHYLENE INSENSITIVE 3-LIKE (EIL) transcription factor (TF) family plays a key role in the ethylene signal transduction pathway. However, it is still unknown how EIL proteins mediate salt tolerance in Tamarix hispida. Here, we cloned and functionally characterized the TF ThEIL1 from T. hispida. ThEIL1 is a nuclear protein possessing transcriptional activation activity. In addition to binding to the PERE motif (GGATTCAA), a novel EIL protein-DNA interaction was identified, in which ThEIL1 binds to GCC-box (AGCCGCC). Gain- and loss-of-function analysis in T. hispida showed that ThEIL1 regulates the expression of trehalose-6-phosphate synthase (TPS), pyrroline-5-carboxylate synthetase (P5CS), and trehalose-6-phosphate phosphatase (TPP) to mediate the biosynthesis of proline and trehalose, respectively, thereby elevating osmotic potential. Additionally, ThEIL1 controls the activities of peroxidase (POD) and superoxide dismutase (SOD) by regulating the expression of genes encoding SODs and PODs, leading to increased reactive oxygen species scavenging capability. Moreover, overexpression of ThEIL1 in Arabidopsis significantly increased root length and fresh weight under salt treatment conditions. Furthermore, the physiological traits of T. hispida were consistent with those of Arabidopsis stably overexpressing ThEIL1. Thus, ThEIL1 serves as a TF that mediates salt tolerance by elevating osmotic potential and improving ROS scavenging capability.

The B’ζ subunit of protein phosphatase 2A negatively regulates ethylene signaling in Arabidopsis

Ethylene is a major plant hormone that regulates plant growth, development, and defense responses to biotic and abiotic stresses. The major pieces of the ethylene signaling pathway have been put together, although several details still need to be elucidated. For instance, the phosphorylation and dephosphorylation processes controlling the ethylene responses are poorly understood and need to be further explored. The type 2A protein phosphatase (PP2A) was suggested to play an important role in the regulation of ethylene biosynthesis, where the A1 subunit of PP2A was shown to be involved in the regulation of the rate-limiting enzyme of the ethylene biosynthetic pathway. However, whether other subunits of PP2A play roles in the ethylene signal transduction pathway is yet to be answered. In this study, we demonstrate that a B subunit, PP2A-B’ζ, positively regulates plant germination and seedling development, as a pp2a-b’ζ mutant is very sensitive to ethylene treatment. Furthermore, PP2A-B’ζ interacts with and stabilizes the kinase CTR1 (Constitutive Triple Response 1), a key enzyme in the ethylene signal transduction pathway, and like CTR1, PP2A-B’ζ negatively regulates ethylene signaling in plants.

Functional characterization of the chlorophyll b reductase gene NYC1 associated with chlorophyll degradation and photosynthesis in Zoysia japonica

Chlorophyll plays essential roles in photosynthesis and its degradation should be regulated precisely. Chlorophyll b reductase encoded by the gene NYC1 is responsible for catalyzing chlorophyll b degradation and maintaining the stability of photosystems. Although NYC1s have been characterized in many plants, their function in the photosynthetic machinery and the underlying regulatory mechanisms at the whole transcriptome level remain elusive. In this study, the ZjNYC1 gene was cloned from the important warm-season turfgrass Zoysia japonica. Analysis of the subcellular localization revealed that ZjNYC1 was localized in the chloroplasts, and results of the bimolecular fluorescence complementation (BiFC) assay indicated that ZjNYC1 could interact in vivo with the NYC1-like protein, ZjNOL. Overexpression of ZjNYC1 accelerated the degradation of chlorophyll, and promoted senescence with accumulation of ABA and reactive oxygen species (ROS) in Arabidopsis. Expression of ZjNYC1 rescued the stay-green phenotype of the Arabidopsis nyc1 mutant. An integrated analysis of chlorophyll fluorescence test (JIP-test) and RNA-sequencing revealed that ZjNYC1 negatively affected the efficiency of the PSII, PSI, and the intersystem electron transport chain. Furthermore, the Calvin cycle and photorespiration were suppressed, but ABA and ethylene synthesis and signal transduction pathways were activated in the transgenic lines. Collectively, these results demonstrated that ZjNYC1 plays important roles in accelerating chlorophyll degradation and senescence, and caused negative affect on the integrity and functionality of the photosystems.