Ethylene induced AcNAC3 and AcNAC4 take part in ethylene synthesis through mediating AcACO1 during kiwifruit (Actinidia chinensis) ripening.

BACKGROUND: Ethylene plays a vital role in the ripening process of kiwifruit. A terrific amount of transcription factors (TFs) have been shown to regulate ethylene synthesis in various fruits. RESULTS: In this research, two new NAC TFs, named AcNAC3 and AcNAC4, were isolated from kiwifruit, which belonged to NAM subfamily. Bioinformatics analysis showed that both AcNAC3 and AcNAC4 were hydrophilic proteins with similar three-dimensional structures. The expression levels of AcNAC3, AcNAC4 and AcACO1 increased during kiwifruit ripening, as well as were induced by ethylene and repressed by 1-methylcyclopropene (1-MCP). Correlation analysis exhibited that ethylene production was positively correlated with the expression levels of AcNAC3, AcNAC4 and AcACO1. Moreover, both AcNAC3 and AcNAC4 acted as transcriptional activators and could bind to and activate AcACO1 promoter. CONCLUSION: All results unveiled that the ethylene-induced AcNAC3 and AcNAC4 were transcriptional activators, and might participate in kiwifruit ripening and ethylene biosynthesis through activating AcACO1, providing a new insight of ethylene synthetic regulation during kiwifruit ripening. © 2024 Society of Chemical Industry.

Two NAC transcription factors regulated fruit softening through activating xyloglucan endotransglucosylase/hydrolase genes during kiwifruit ripening.

Kiwifruit is a climacteric fruit that is prone to ripening and softening. Understanding molecular regulatory mechanism of kiwifruit softening, is helpful to ensure long-term storage of fruit. In the study, two NAC TFs and two XTH genes were isolated from kiwifruit. Phylogenetic tree showed that both AcNAC1 and AcNAC2 belonged to NAP subfamily, AcXTH1 belong to I subfamily, and AcXTH2 belong to III subfamily. Bioinformatics analysis predicted that AcNAC1 and AcNAC2 possessed similar three-dimensional structural, and belonged to hydrophilic proteins. AcXTH1 and AcXTH2 were hydrophilic proteins and contained signal peptides. AcXTH1 had a transmembrane structure, but AcXTH2 did not. qRT-PCR results showed that AcNAC1, AcNAC2, AcXTH1 and AcXTH2 were increased during kiwifruit ripening. Correlation analysis showed that kiwifruit softening was closely related to endotransglucosylase/hydrolase genes and NAC TFs, as well as there was also a close relationship between AcXTHs and AcNACs. Moreover, both AcNAC1 and AcNAC2 were transcriptional activators located in nucleus, which bound to and activated the promoters of AcXTH1 and AcXTH2. In shortly, we proved that the roles of NAC TFs in mediating fruit softening during kiwifruit ripening. Altogether, our results clarified that AcNAC1 and AcNAC2 were transcriptional activators, and took part in kiwifruit ripening and softening through activating endotransglucosylase/hydrolase genes, providing a new insight of fruit softening network in kiwifruit ripening.

Antimicrobial activity of gamma-poly (glutamic acid), a preservative coating for cherries.

We investigated the minimum inhibitory concentration (MIC), antibacterial activity, and preservation ability of four molar masses of γ-polyglutamic acid (PGA) against Escherichia coli, Bacillus subtilis, and yeast. The antibacterial mechanism was determined based on the cell structure, membrane permeability, and microscopic morphology of the microorganisms. We then measured the weight loss, decay rate, total acid, catalase activity, peroxidase activity, and malondialdehyde content toward the possible use of PGA as a preservative coating for cherries. When the molar mass was greater than 700 kDa, the MIC for Escherichia coli and Bacillus subtilis was less than 2.5 mg/mL. The mechanism of action of the four molar masses of PGA was different with respect to the three microbial species, but a higher molar mass of PGA corresponded to stronger inhibition against the microbes. PGA of 2000 kDa molar mass damaged the microbial cellular structure, resulting in excretion of alkaline phosphatase, but PGA of 1.5 kDa molar mass affected the membrane permeability and the amount of soluble sugar. Scanning electron microscopy indicated the inhibitory effect of PGA. The antibacterial mechanism of PGA was related to the molar mass of PGA and the microbial membrane structure. Compared with the control, a PGA coating effectively inhibit the spoilage rate, delay the ripening, and prolong the shelf life of cherries.

Preparation and Properties of Pea Starch/ε-Polylysine Composite Films.

The composite films comprising pea starch (St) and ε-polylysine (PL) as the matrix and glycerol and sodium alginate as the plasticizers were investigated. The rheological properties, mechanical properties, Fourier transformed infrared spectroscopy, water vapor permeability (WVP), oil permeability, microstructure, thermogravimetry (TGA), and antimicrobial properties of the composite films were analyzed. The properties of the composite films with different mass ratios of St/PL varied significantly. First, the five film solutions were different pseudoplastic fluids. Additionally, as the mass ratio of PL increased, the tensile strength of the blends decreased from 9.49 to 0.14 MPa, the fracture elongation increased from 38.41 to 174.03%, the WVP increased, and the oil resistance decreased substantially. The films with a broad range of St/PL ratios were highly soluble; however, the solubility of the film with a St/PL ratio of 2:8 was reduced. Lastly, the inhibition of E. coli, B.subtilis, and yeast by the films increased with increasing mass ratios of PL, and the inhibition of B.subtilis was the strongest.

Two papaya MYB proteins function in fruit ripening by regulating some genes involved in cell-wall degradation and carotenoid biosynthesis.

BACKGROUND: MYB transcription factors (TFs) are common in plants and play important functions in growth and development, including fruit development and ripening. However, the role of MYB proteins in papaya ripening (fruit ripening and carotenoid biosynthesis) remains unclear. RESULTS: Two MYB genes were cloned from papaya pulp. They were named CpMYB1 (MYB44-like) and CpMYB2, and belong to the S22 subgroup of the R2R3-MYB family. Their expression levels decreased during fruit ripening. Subcellular localization analysis showed that both CpMYB1 and CpMYB2 were nuclear proteins, indicating that they might function in the nucleus. Moreover, CpMYB1 and CpMYB2 could bind to the promoters of cell-wall degradation genes (CpPME1, CpPME2, and CpPG5) and carotenoid biosynthesis genes (CpPDS2, CpPDS4, and CpCHY-b). Further research found that both CpMYB1 and CpMYB2 were transcriptional repressors, and they could suppress the activities of the promoters of CpPME1, CpPME2, CpPG5, CpPDS2, CpPDS4, and CpCHY-b. CONCLUSION: These results indicated that MYB TFs CpMYB1 and CpMYB2 might have a function in papaya fruit softening and carotenoid accumulation by regulating cell-wall degradation and carotenoid biosynthesis related genes, which provide a new view about the role of MYB TFs in fruit ripening. © 2020 Society of Chemical Industry.

Quality Evaluation of Corydalis yanhusuo by High-Performance Liquid Chromatography Fingerprinting Coupled with Multicomponent Quantitative Analysis.

Corydalis Rhizoma is the tuber of Corydalis yanhusuo W. T. Wang, which has been long used in traditional Chinese medicine. Herein, the quality of C. yanhusuo samples collected from 23 regions of three provinces in China is evaluated through high-performance liquid chromatography fingerprinting coupled with similarity, hierarchical clustering, and principal component analyses. Sample similarities are evaluated according to the State Food and Drug Administration requirements by selection of 18 characteristic chromatographic fingerprint peaks and are found to vary between 0.455 and 0.999. Moreover, common patterns of a typical local variety of C. yanhusuo sourced in the Panan County are established. The obtained results show that the combination of quantitative analysis and chromatographic fingerprint analysis can be readily utilized for quality control purposes, offering a comprehensive strategy for quality evaluation of C. yanhusuo and related products.

Histone Deacetylase CpHDA3 Is Functionally Associated with CpERF9 in Suppression of CpPME1/2 and CpPG5 Genes during Papaya Fruit Ripening.

Histone deacetylase (HDAC) performs important functions in plant growth and development, including fruit ripening. As a complex biological process, fruit ripening involves the histone acetylation modification of ripening-associated genes. Histone deacetylase genes (HDACs) have been well studied in Arabidopsis and rice, but the biological functions of HDACs in papaya are poorly understood. In the present work, three CpHDACs, belonging to the RPD3/HDA1 subfamily, were identified from papaya and named as CpHDA1, CpHDA2, and CpHDA3. CpHDA1 and CpHDA2 were induced by propylene, while CpHDA3 was propylene-repressed. Moreover, CpHDA3 protein could physically interact with CpERF9 and enhance the transcriptional repression activities of CpERF9 to downstream genes CpPME1, CpPME2 and CpPG5. Histone acetylation levels of CpPME1 and CpPG5 were increased during fruit ripening. Taken together, these results suggested that CpERF9 recruits CpHDA3 to form a histone deacetylase repressor complex to mediate pectin methylesterase and polygalacturonase genes expression during papaya fruit ripening and softening.

Cold-inducible MaC2H2s are associated with cold stress response of banana fruit via regulating MaICE1.

KEY MESSAGE: MaC2H2s are involved in cold stress response of banana fruit via repressing the transcription of MaICE1. Although C2H2 zinc finger proteins have been found to be involved in banana fruit ripening through transcriptional controlling of ethylene biosynthetic genes, their involvement in cold stress of banana remains elusive. In this study, another C2H2-ZFP gene from banana fruit was identified, which was named as MaC2H2-3. Gene expression analysis revealed that MaC2H2-1, MaC2H2-2 and MaC2H2-3 were cold inducible in the peel of banana during low temperature storage. MaC2H2-3 functions as a transcriptional repressor and localizes predominantly in nucleus. Particularly, promoters of MaC2H2-2 and MaC2H2-3 were noticeably activated by cold as well, further indicating the potential roles of C2H2 in cold stress of banana. Moreover, MaC2H2-2 and MaC2H2-3 significantly repressed the transcription of MaICE1, a key component in cold signaling pathway. Overall, these findings suggest that MaC2H2s may take part in controlling cold stress of banana through suppressing the transcription of MaICE1, providing new insight of the regulatory basis of C2H2 in cold stress.

Papaya CpEIN3a and CpNAC2 Co-operatively Regulate Carotenoid Biosynthesis-Related Genes CpPDS2/4, CpLCY-e and CpCHY-b During Fruit Ripening.

Papaya is an important tropical fruit with a rich source of carotenoids. The ripening of papaya is a physiological and metabolic process with remarkable changes including accumulation of carotenoids, which depends primarily on the action of ethylene. Ethylene response is mediated by a transcriptional cascade involving the transcription factor families of EIN3/EILs and ERFs. Although ERF members have been reported to control carotenoid production in Arabidopsis and tomato, whether EIN3/EILs are also involved in carotenoid biosynthesis during fruit ripening remains unclear. In this work, two EIN3 genes from papaya fruit, namely CpEIN3a and CpEIN3b, were studied, of which CpEIN3a was increased during fruit ripening, concomitant with the increase of transcripts of carotenoid biosynthesis-related genes including CpPDS2/4, CpZDS, CpLCY-e and CpCHY-b, and carotenoid content. Electrophoretic mobility shift assays (EMSAs) and transient expression analyses revealed that CpEIN3a was able to bind to the promoters of CpPDS4 and CpCHY-b, and promoted their transcription. Protein-protein interaction assays indicated that CpEIN3a physically interacted with another transcription factor CpNAC2, which acted as a transcriptional activator of CpPDS2/4, CpZDS, CpLCY-e and CpCHY-b by directly binding to their promoters. More importantly, the transcriptional activation abilities of CpPDS2/4, CpLCY-e and CpCHY-b were more pronounced following their interaction. Collectively, our findings suggest that CpEIN3a interacts with CpNAC2 and, individually or co-operatively, activates the transcription of a subset of carotenoid biosynthesis-related genes, providing new insights into the regulatory networks of carotenoid biosynthesis during papaya fruit ripening.

Papaya CpERF9 acts as a transcriptional repressor of cell-wall-modifying genes CpPME1/2 and CpPG5 involved in fruit ripening.

KEY MESSAGE: CpERF9 controls papaya fruit ripening through transcriptional repression of cell-wall-modifying genes CpPME1/2 and CpPG5 by directly binding to their promoters. Papaya fruit ripening is an intricate and highly coordinated developmental process which is controlled by the action of ethylene and expression of numerous ethylene-responsive genes. Ethylene response factors (ERFs) representing the last regulators of ethylene-signaling pathway determine the specificities of ethylene response. However, knowledge concerning the transcriptional controlling mechanism of ERF-mediated papaya fruit ripening is limited. In the present work, a gene-encoding AP2/ERF protein with two ERF-associated amphiphilic repression (EAR) motifs, named CpERF9, was characterized from papaya fruit. CpERF9 was found to localize in nucleus, and possess transcriptional repression ability. CpERF9 expression steadily decreased during papaya fruit ripening, while several genes encoding pectin methylesterases (PMEs) and polygalacturonases (PGs), such as CpPME1/2 and CpPG5, were gradually increased, paralleling the decline of fruit firmness. Electrophoretic mobility shift assay (EMSA) demonstrated a specific binding of CpERF9 to promoters of CpPME1/2 and CpPG5, via the GCC-box motif. Transient expression of CpERF9 in tobacco repressed CpPME1/2 and CpPG5 promoter activities, which was depended on two EAR motifs of CpERF9 protein. Taken together, these findings suggest that papaya CpERF9 may act as a transcriptional repressor of several cell-wall modifying genes, such as CpPME1/2 and CpPG5, via directly binding to their promoters.

The Banana Transcriptional Repressor MaDEAR1 Negatively Regulates Cell Wall-Modifying Genes Involved in Fruit Ripening.

Ethylene plays an essential role in many biological processes including fruit ripening via modulation of ethylene signaling pathway. Ethylene Response Factors (ERFs) are key transcription factors (TFs) involved in ethylene perception and are divided into AP2, RAV, ERF, and DREB sub-families. Although a number of studies have implicated the involvement of DREB sub-family genes in stress responses, little is known about their roles in fruit ripening. In this study, we identified a DREB TF with a EAR motif, designated as MaDEAR1, which is a nucleus-localized transcriptional repressor. Expression analysis indicated that MaDEAR1 expression was repressed by ethylene, with reduced levels of histone H3 and H4 acetylation at its regulatory regions during fruit ripening. In addition, MaDEAR1 promoter activity was also suppressed in response to ethylene treatment. More importantly, MaDEAR1 directly binds to the DRE/CRT motifs in promoters of several cell wall-modifying genes including MaEXP1/3, MaPG1, MaXTH10, MaPL3, and MaPME3 associated with fruit softening during ripening and represses their activities. These data suggest that MaDEAR1 acts as a transcriptional repressor of cell wall-modifying genes, and may be negatively involved in ethylene-mediated ripening of banana fruit. Our findings provide new insights into the involvement of DREB TFs in the regulation of fruit ripening.

The Papaya Transcription Factor CpNAC1 Modulates Carotenoid Biosynthesis through Activating Phytoene Desaturase Genes CpPDS2/4 during Fruit Ripening.

Papaya fruits accumulate carotenoids during fruit ripening. Although many papaya carotenoid biosynthesis pathway genes have been identified, the transcriptional regulators of these genes have not been characterized. In this study, a NAC transcription factor, designated as CpNAC1, was characterized from papaya fruit. CpNAC1 was localized exclusively in nucleus and possessed transcriptional activation activity. Expression of carotenoid biosynthesis genes phytoene desaturases (CpPDSs) and CpNAC1 was increased during fruit ripening and by propylene treatment, which correlates well with the elevated carotenoid content in papaya. The gel mobility shift assays and transient expression analyses demonstrated that CpNAC1 directly binds to the NAC binding site (NACBS) motifs in CpPDS2/4 promoters and activates them. Collectively, these data suggest that CpNAC1 may act as a positive regulator of carotenoid biosynthesis during papaya fruit ripening possibly via transcriptional activation of CpPDSs such as CpPDS2/4.

Banana Transcription Factor MaERF11 Recruits Histone Deacetylase MaHDA1 and Represses the Expression of MaACO1 and Expansins during Fruit Ripening.

Phytohormone ethylene controls diverse developmental and physiological processes such as fruit ripening via modulation of ethylene signaling pathway. Our previous study identified that ETHYLENE RESPONSE FACTOR11 (MaERF11), a transcription factor in the ethylene signaling pathway, negatively regulates the ripening of banana, but the mechanism for the MaERF11-mediated transcriptional regulation remains largely unknown. Here we showed that MaERF11 has intrinsic transcriptional repression activity in planta. Electrophoretic mobility shift assay and chromatin immunoprecipitation analyses demonstrated that MaERF11 binds to promoters of three ripening-related Expansin genes, MaEXP2, MaEXP7 and MaEXP8, as well as an ethylene biosynthetic gene MaACO1, via the GCC-box motif. Furthermore, expression patterns of MaACO1, MaEXP2, MaEXP7, and MaEXP8 genes are correlated with the changes of histone H3 and H4 acetylation level during fruit ripening. Moreover, we found that MaERF11 physically interacts with a histone deacetylase, MaHDA1, which has histone deacetylase activity, and the interaction significantly strengthens the MaERF11-mediated transcriptional repression of MaACO1 and Expansins Taken together, these findings suggest that MaERF11 may recruit MaHDA1 to its target genes and repress their expression via histone deacetylation.

The dimer, trimer and 1,2,4-trithiolane of adamantanethione.

The molecules of dispiro[1,3-dithietane-2,2':4,2"-diadamantane], C(20)H(28)S(2), have crystallographic C(i) symmetry, as well as local D(2h) symmetry, and a planar 1,3-dithietane ring. The molecules of trispiro[1,3,5-trithiane-2,2':4,2":6,2"'-triadamantane], C(30)H(42)S(3), have approximate C(2) symmetry and the 1,3,5-trithiane ring has a twist-boat conformation. The C-S-C bond angles within the ring are about 8 degrees larger than observed in most related 1,3,5-trithiane structures. In dispiro[1,2,4-trithiolane-3,2':5,2"-diadamantane], C(20)H(28)S(3), the molecules have local C(2) symmetry and the 1,2,4-trithiolane ring has a half-chair conformation.