Passion fruit is an important industrial crop with considerable economic value, yet its cultivation and postharvest quality are often limited by abiotic stresses. Expanding on previous work that identified 91 AP2/ERF transcription factors in passion fruit, this study reports three additional members—PeERF-54, PeERF-55, and PeERF-56—bringing the total to 94. We systematically investigated the roles of these PeAP2/ERF genes in abiotic stress adaptation and fruit maturation. Transcriptome profiling revealed that the majority of PeAP2/ERFs were differentially expressed under various abiotic stress conditions. During fruit pulp development, several members exhibited significantly higher expression at early (T1) and middle (T2) ripening stages compared to the late stage (T3). Notably, PeDREB-1 was strongly induced under cold stress. Meanwhile, transcriptomic data indicate that its expression levels decline progressively throughout fruit ripening, with expression levels at stage T1 being 1.82-fold and 5.49-fold higher than those at stages T2 and T3, respectively. Functional analyses in transgenic Saccharomyces cerevisiae and Arabidopsis thaliana demonstrated that PeDREB-1 enhances cold stress tolerance, supported by deeper staining and upregulation of stress-responsive genes. Moreover, transient overexpression and silencing assays in passion fruit revealed that PeDREB-1 delays ripening. Together, these results establish PeAP2/ERF transcription factors as key regulators of abiotic stress responses in passion fruit and identify PeDREB-1 as a dual-function regulator involved in both cold stress tolerance and the regulation of fruit ripening—an attribute of high relevance for industrial production and postharvest management.

The genome-wide analysis identified 55 PeSTs (Passiflora edulis sugar transporters), having significant variations in exon–intron structure, protein motifs, cis-regulatory elements, and syntenic relationships, which might reveal their functional diversity. Phylogenetic analysis classified these genes into eight subfamilies, where the STP subfamily showed significant expansion during the process of evolution. In molecular docking, the strongest binding affinities in Glu–PePLT4, Fru–PeSTP10, Gal–PePLT4, and Suc–PeTMT2 complexes might reveal their substrate specificity and transport kinetics. Following that, the molecular dynamics simulation supported the stability of the docked complexes. The presence of significant light, hormone, and stress regulatory cis-elements among the PeST promoters might reveal their hormone and stress regulatory functions. The highest homologous pairs (62) of PeSTs with M. domestica during synteny analysis among five plant genomes might guide their duplication pattern across the genomes. RNA-seq-based expression analysis, following real-time qRT-PCR mediated validation of upregulated expression of PeERD6L-1, PeSTP6, PeSTP8, PePLT1, and PeTMT5 genes under drought, salt, cold, and heat stresses, might guide their potential regulatory roles in tolerance to the respective stresses. Significant protein–protein and protein-TFs interactions in PeSTs might reveal their inter-regulatory roles in transporting sugars. These critical findings might guide the plant biologists in developing a synthetic biology-guided circuit enabled multiple abiotic stress-tolerant high-sugar passion fruit.

Passiflora germplasm, characterized by rich genetic diversity, play a pivotal role in the evolutionary history of tropical crops. With the expansion of commercial passion fruit cultivation, germplasm conservation faces multiple challenges. Establishing efficient in vitro preservation systems is crucial for protecting the crop’s genetic diversity, ensuring a stable supply of breeding materials, and supporting sustainable industry development. This study achieves dual breakthroughs in the field of Passiflora in vitro conservation. First, a unified induction protocol was developed, enabling synchronized regulation of axillary bud induction and clustered shoot proliferation across five genetically divergent Passiflora species including edible, ornamental, and medicinal types. This breakthrough overcomes the prevalent interspecific recalcitrance that plagues in vitro culture of multiple species. The optimal conditions for axillary bud induction were identified as MS medium supplemented with 1.5 mg/L 6-Benzylaminopurine (6-BA), 0.03 mg/L Zeatin (ZT), and 0.05 mg/L indole-3-butyric acid (IBA). A modified MS-based subculture medium containing 1.75 mg/L 6-BA with the same ZT/IBA ratio achieved efficient proliferation of shoots. Second, a high-efficiency soil substrate conservation system was established for long-term conservation. Through periodic MS nutrient supplementation, the survival rate of all species exceeded 67

Research on passion fruit traits primarily focuses on abiotic stress due to its detrimental impact on the industry. Transcription factors (TFs) mitigate abiotic stress by participating in various biological processes, among which heat shock factors (HSFs) play a pivotal role in responding to both biotic and abiotic stresses and conferring stress tolerance. This study identified 15 PeHSF family members with complete sequences using a high-quality genome of passion fruit. A systematic analysis of PeHSFs across the genome was conducted through bioinformatics and transcriptome sequencing. Transcriptomic data revealed higher expression levels of most PeHSFs in fruit pulp at stages T1 and T2 compared with T3, demonstrating the family's responsiveness to diverse abiotic stresses. Subsequent subcellular localization confirmed nuclear localization of the selected gene PeHSF-2. Heterologous expression of PeHSF-2 in the INVSc1 yeast strain and Arabidopsis thaliana significantly enhanced tolerance to drought, salt, cold, and heat stresses. Furthermore, PeHSF-2 over-expression up-regulated stress-responsive genes (P5CS1, SOS1, HSP70, and CBF2), and interacted with PeSIP2-2. This study lays the groundwork for further investigation into the regulatory mechanisms of PeHSFs under abiotic stress conditions.

Natural Antisense Transcripts (NATs) are a kind of complex regulatory RNAs that play crucial roles in gene expression and regulation. However, the NATs in Cannabis Sativa L., a widely economic and medicinal plant rich in cannabinoids remain unknown. In this study, we comprehensively predicted C. sativa NATs genome-wide using strand-specific RNA sequencing (ssRNA-Seq) data, and validated the expression profiles by strand-specific quantitative reverse transcription PCR (ssRT-qPCR). Consequently, a total of 307 NATs were predicted in C. sativa, including 104 cis- and 203 trans- NATs. Functional enrichment analysis demonstrated the potential involvement of the C. sativa NATs in DNA polymerase activity, RNA-DNA hybrid ribonuclease activity, and nucleic acid binding. Finally, 18 cis- and 376 trans- NAT-ST pairs were predicted to produce 621 cis- and 5,679 trans- small interfering RNA (nat-siRNAs), respectively. These nat-siRNAs were potentially involved in the biosynthesis of cannabinoids and cellulose. All these results will shed light on the regulation of NATs and nat-siRNAs in C. sativa.

Non-small cell lung cancer (NSCLC) is a malignancy with poor outcomes. METTL3 has been demonstrated to facilitate aerobic glycolysis for malignancies. We aimed to explore how METTL3 regulates aerobic glycolysis in NSCLC. METTL3 and PIK3CB expressions in NSCLC tissues, and the associations between METTL3 and AKT, as well as METTL3 and mTOR were analyzed by public databases. After NSCLC cells with METTL3 knockdown or overexpression were successfully established, the malignant behaviors of the cells were evaluated. Subsequently, cellular aerobic glycolysis was detected by measuring glucose consumption, lactate production, ATP/ADP ratio and the expressions of GLUT1, HK2, PKM2 and LDHA proteins. Furthermore, PI3K, AKT and mTOR phosphorylation of the cells was detected. Finally, METTL3 silenced cells were exposed to 740Y-P (a PI3K activator) and 2-Deoxy-d-glucose (a glycolysis inhibitor) to further investigate the mechanisms of METTL3 on NSCLC. METTL3 and PI3KCB were highly expressed, and METTL3 expression positively correlated with AKT and mTOR expressions in NSCLC clinical samples. METTL3 knockdown suppressed NSCLC cellular viability, migration and invasion, but induced apoptosis. Furthermore, METTL3 knockdown decreased above aerobic glycolysis-related indicators, and suppressed PI3K, AKT and mTOR phosphorylation. Conversely, METTL3 overexpression produced opposite effects. Notably, the suppression of METTL3 silencing on NSCLC cellular viability, migration and aerobic glycolysis were reserved by 740Y-P, and the roles of 740Y-P on METTL3-silenced cells were further counteracted by 2-Deoxy-d-glucose. METTL3 may facilitate the aerobic glycolysis of NSCLC cells by activating the PI3K/AKT/mTOR pathway, indicating that the METTL3-mediated PI3K/AKT/mTOR axis is an underlying therapeutic target for NSCLC.

Citri Reticulatae Pericarpium(CRP) and Polygalae Radix (PR) are traditional Chinese medicinal herbs and dietary supplements, and they are commonly included in most prescriptions. Fungal contamination in CRP and PR has long been a concern for the public. Thus, in this study, the fungal diversity and abundance in CRP and PR were investigated via high-throughput sequencing technology. Analysis was conducted to determine the differences in dominant fungal microbiomes between mouldy and nonmouldy samples, and the correlation between the total number of fungi and fungal diversity in medicinal herbs at different altitudes and latitudes with other research was explored. Results show the fungal contamination of all 12 samples. At the phylum level, Ascomycota prevailed the most in CRP and PR with relative abundances of 56.96%–99.26% and 23.28%–76.06%, respectively. The dominant genera in CRP comprised Xeromyces (2.57%–53.21%), Aspergillus (0.18%–23.04%), Cystofilobasidium (0.032%–28.03) and Xerochrysium (0.31%–34.41%). The dominant genera in PR included Wallemia (0.27%–56.20%), Aspergillus (2.03%–48.06%), Xeromyces (0.20%–48.28%) and Xerochrysium (0.093%–43.95%). Two potential toxigenic fungi were detected: Alternaria alternata and Aspergillus versicolor. In addition, combined with the results of previous studies, the findings reveal the possible relation of diversity and abundance of fungal species to altitude and latitude: As the altitude and latitude decrease, the diversity and abundance of fungal species may increase. In conclusion, for the first time, this study focused on altitude and latitude as indicators and integrated multiple research findings to explore the correlation between the total amount and diversity of fungal species in the two edible herbs. Results reveal the fungal contamination of the surfaces of CRP and PR and provide a theoretical basis for the prevention and control of their fungal contamination.

Passiflora edulis, also known as passion fruit, is celebrated for its rich nutritional content, distinctive flavour, and significant medicinal benefits. At present, viral diseases pose a major challenge to the passion fruit industry, affecting both the production and quality of the fruit. These diseases impede the sustainable and healthy growth of the passion fruit sector. In recent years, with the expansion of P. edulis cultivation areas, virus mutations, and advances in virus detection technology, an increasing number of virus species infecting P. edulis have been discovered. To date, more than 40 different virus species have been identified; however, there are different strains within the same virus. This poses a challenge for the control and prevention of P. edulis virus disease. Therefore, this review discusses the different types of viruses and their characteristics, modes of transmission, and effects on the growth of the passion fruit plant, as well as the mechanisms of virus generation and preventive measures, with the hope that these discussions will provide a comprehensive understanding of and countermeasures for viruses in passion fruit.

Background Invertases (INVs) are key enzymes in sugar metabolism, cleaving sucrose into glucose and fructose and playing an important role in plant development and the stress response, however, the INV gene family in passion fruit has not been systematically reported. Results In this study, a total of 16 PeINV genes were identified from the passion fruit genome and named according to their subcellular location and chromosome position. These include six cell wall invertase (CWINV) genes, two vacuolar invertase (VINV) genes, and eight neutral/alkaline invertase (N/AINV) genes. The gene structures, phylogenetic tree, and cis-acting elements of PeINV gene family were predicted using bioinformatics methods. Results showed that the upstream promoter region of the PeINV genes contained various response elements; particularly, PeVINV2, PeN/AINV3, PeN/AINV5, PeN/AINV6, PeN/AINV7, and PeN/AINV8 had more response elements. Additionally, the expression profiles of PeINV genes under different abiotic stresses (drought, salt, cold temperature, and high temperature) indicated that PeCWINV5, PeCWINV6, PeVINV1, PeVINV2, PeN/AINV2, PeN/AINV3, PeN/AINV6, and PeN/AINV7 responded significantly to these abiotic stresses, which was consistent with cis-acting element prediction results. Sucrose, glucose, and fructose are main soluble components in passion fruit pulp. The contents of total soluble sugar, hexoses, and sweetness index increased significantly at early stages during fruit ripening. Transcriptome data showed that with an increase in fruit development and maturity, the expression levels of PeCWINV2, PeCWINV5, and PeN/AINV3 exhibited an up-regulated trend, especially for PeCWINV5 which showed highest abundance, this correlated with the accumulation of soluble sugar and sweetness index. Transient overexpression results demonstrated that the contents of fructose, glucose and sucrose increased in the pulp of PeCWINV5 overexpressing fruit. It is speculated that this cell wall invertase gene, PeCWINV5, may play an important role in sucrose unloading and hexose accumulation. Conclusion In this study, we systematically identified INV genes in passion fruit for the first time and further investigated their physicochemical properties, evolution, and expression patterns. Furthermore, we screened out a key candidate gene involved in hexose accumulation. This study lays a foundation for further study on INV genes and will be beneficial on the genetic improvement of passion fruit breeding.

WRKY transcription factors (TFs) are a superfamily of regulators involved in plant responses to pathogens and abiotic stress. Passion fruit is famous for its unique flavor and nutrient-rich juice, but its growth is limited by environmental factors and pathogens. In this study, 55 WRKY genes were identified from the Passiflora edulis genome. The structure and evolutionary characteristics of PeWRKYs were analyzed using a bioinformatics approach. PeWRKYs were classified into seven subgroups (I, IIa, IIb, IIc, IId, IIe, III) according to their homologs in Arabidopsis thaliana. Group IIa PeWRKY48 gene was highly up-regulated under cold stress by RNA expression analysis, and transgenic PeWRKY48 in yeast and Arabidopsis showed resistance exposure to cold, salt, and drought stress. Metabolome and transcriptome co-expression analysis of two different disease resistance genotypes of P. edulis identified PeWRKY30 as a key TF co-expressed with flavonoid accumulation in yellow fruit P. edulis, which may contribute to biotic or abiotic resistance. The qRT-PCR verified the expression of key genes in different tissues of P. edulis and in different species of Passiflora. This study provides a set of WRKY candidate genes that will facilitate the genetic improvement of disease and abiotic tolerance in passion fruit.

The HD-ZIP (homeodomain-leucine zipper) genes hold significant importance in transcriptional regulation, especially in plant development and responses to abiotic stresses. However, a comprehensive study targeting HD-ZIP family members in passion fruit has been absent. In our current research, 34 HD-ZIP family members (PeHBs) were identified by bioinformatics analysis. Transcriptome analysis revealed that PeHBs exhibited distinct expression patterns when subjected to the four different abiotic stresses, and significant differential expression of PeHBs was also found among the three developmental stages of the fruit and between the purple and yellow genotype passion fruit leaves. An integrated metabolome and transcriptome analysis further revealed that the HD-ZIP III class gene PeHB31 (homologous to ATHB8), was co-upexpressed with lignans in yellow fruit P. edulis (commonly used as a resistance rootstock) when compared to purple fruit P. edulis. The transformation of Arabidopsis and yeast with the PeHB31 gene showed an enhancement in their capacity to withstand drought conditions. Notably, the transgenic Arabidopsis plants exhibited an increase in lignin content within the vascular tissues of their stems. This research lays the groundwork for future studies on the control mechanisms of lignin biosynthesis by HD-ZIP genes (especially HD-ZIP classes III and I) involved in drought tolerance.

Genome visualization tools are important for exploring genomic features and their interactions. Currently, visualization of the plant mitochondrial genomes (mitogenome) depends on those tools designed originally for animal mitogenomes and plant plastomes. These tools cannot faithfully present features unique to the plant mitogenomes, such as non-linear exon arrangement for genes, prevalence of functional non-coding features, and complex chromosomal architectures. To address these challenges, a software package plant mitochondrial genome map (PMGmap), was developed using Python programming language. PMGmap can draw genes at exon levels, draw cis- and trans-splicing gene maps, draw non-coding features, draw repetitive sequences, scale the genic regions using a scaling the genic regions on the genome (SGM) algorithm, and draw multiple chromosomes simultaneously. We compared PMGmap with other leading tools on 405 plant mitogenomes and found that PMGmap allowed the visualization of the above-mentioned features better than those tools. We believe PMGmap will become an invaluable tool for plant mitogenome research. The web and container versions and the source code of PMGmap can be accessed at http://www.1kmpg.cn/pmgmap.

Abiotic stress is the focus of research on passion fruit characters because of its damage to the industry. Basic helix-loop-helix (bHLH) is one of the Transcription factors (TFs) which can act in an anti-abiotic stress role through diverse biological processes. However, no systemic analysis of the passion fruit bHLH (PebHLH) family was reported. In this study, 117 PebHLH members were identified from the genome of passion fruit, related to plant stress resistance and development by prediction of protein interaction. Furthermore, the transcriptome sequencing results showed that the PebHLHs responded to different abiotic stresses. At different ripening stages of passion fruit, the expression level of most PebHLHs in the immature stage (T1) was higher than that in the mature stage (T2 and T3). Eight PebHLHs with differentially expressed under different stress treatments and different ripening stages were selected and verified by qRT-PCR. In this research, the expression of one member, PebHLH56, was induced under cold stress. Further, the promoter of PebHLH56 was fused to β-Galactosidase (GUS) to generate the expression vector that was transformed into Arabidopsis. It showed that PebHLH56 could significantly respond to cold stress. This study provided new insights into the regulatory functions of PebHLH genes during fruit maturity stages and abiotic stress, thereby improving the understanding of the characteristics and evolution of the PebHLH gene family.

The bZIP transcription factors are well-known transcriptional regulators that are essential for regulating resistance to biotic and abiotic stresses in plants. In this study, a total of 56 putative bZIP members were identified in passion fruit (Passiflora edulis). An integrative analysis was performed using bioinformatics. Transcriptome analysis revealed that most PebZIPs respond to drought, salt, cold and heat stress. By combining the transcriptome results of two different resistant genotypes, four representative members were finally selected for differential expression validation in different tissues and cultivars. Furthermore, transcriptome and metabolome association analysis revealed consistent expression trends of PeZIP20 and PeZIP21, with only one difference at 63aa, with different metabolites including flavonoids, lipids and amino acids. This work will contribute to further studies of the functions of bZIPs and their resistance properties, as well as to the development of novel germplasm.

Passion fruit (Passiflora edulis Sims) is a vine of the Passiflora genus in the Passifloraceae family. The extracted components include flavonoids and terpenoids, which have good anti-anxiety and anti-inflammatory effects in humans. In this study, we analyzed the transcriptomes of four tissues of the ‘Zixiang’ cultivar using RNA-Seq, which provided a dataset for functional gene mining. The de novo assembly of these reads generated 96 883 unigenes, among which 61 022 unigenes were annotated (62.99% yield). In addition to its edible value, another important application of passion fruit is its medicinal value. The flavonoids and terpenoids are mainly derivatives of luteolin, apigenin, cycloartane triterpenoid saponins and other active substances in leaf extracts. A series of candidate unigenes in the transcriptome data that are potentially involved in the flavonoid and terpenoid synthesis pathways were screened using homology-based BLAST and phylogenetic analysis. The results showed that the biosynthesis of triterpenoids in passion fruit comes from the branches of the mevalonate (MVA) and 2-C-methyl-D-erythritol 4-phosphate/1-deoxy-D-xylulose 5-phosphate (MEP/DOXP) pathways, which is different from the MVA pathway that is used in other fruit trees. Most of the candidate genes were found to be highly expressed in the leaves and/or flowers. Quantitative real-time PCR (qRT-PCR) verification was carried out and confirmed the reliability of the RNA-Seq data. Further amplification and functional analysis of these putative unigenes will provide additional insight into the biosynthesis of flavonoids and terpenoids in passion fruit.

Flavanone 3-hydroxylase (F3H) plays a crucial role in the biosynthesis of flavonoids. In the present study, one F3H gene (P_edulia040010337.g) from Passiflora edulis Sims, which has a coding sequence (CDS) of 1161 bp, encoding a protein consisting of 386 amino acid residues was cloned. The PeF3H protein contains a non-heme dioxygenase (DIOX-N superfamily) domain and a typical F3H protein functional domain (2OG-FeII-Oxy dioxygenase). Phylogenetic analysis revealed that the PeF3H protein shared high similarity with F3H proteins in Turnera subulata, Populus alba, and Populus tomentosa, with 88% identities of amino acid sequences. The PeF3H protein lacks a transmembrane structure, indicating it is likely to be expressed in the mitochondria. Additionally, 3D structure, protein and protein interaction, and KEGG pathway of PeF3H were anticipated based on homologous proteins. qRT-PCR analysis showed that PeF3H was highly expressed in leaves, followed by stems and roots. These studies have provided insights into the molecular mechanisms underlying flavonoid biosynthesis and predicted potential targets for genetic engineering to improve the nutritional and medicinal properties of passion fruit. Bangladesh J. Bot. 52(2): 613-623, 2023 (June) Special

Dendrobium nobile Lindl., as an endangered medicinal plant within the genus Dendrobium, is widely distributed in southwestern China and has important ecological and economic value. There are a variety of metabolites with pharmacological activity in D. nobile. The alkaloids and polysaccharides contained within D. nobile are very important active components, which mainly have antiviral, anti-tumor, and immunity improvement effects. However, the changes in the compounds and functional genes of D. nobile induced by methyl jasmonate (MeJA) are not clearly understood. In this study, the metabolome and transcriptome of D. nobile were analyzed after exposure to MeJA. A total of 377 differential metabolites were obtained through data analysis, of which 15 were related to polysaccharide pathways and 35 were related to terpenoids and alkaloids pathways. Additionally, the transcriptome sequencing results identified 3256 differentially expressed genes that were discovered in 11 groups. Compared with the control group, 1346 unigenes were differentially expressed in the samples treated with MeJA for 14 days (TF14). Moreover, the expression levels of differentially expressed genes were also significant at different growth and development stages. According to GO and KEGG annotations, 189 and 99 candidate genes were identified as being involved in terpenoid biosynthesis and polysaccharide biosynthesis, respectively. In addition, the co-expression analysis indicated that 238 and 313 transcription factors (TFs) may contribute to the regulation of terpenoid and polysaccharide biosynthesis, respectively. Through a heat map analysis, fourteen terpenoid synthetase genes, twenty-three cytochrome P450 oxidase genes, eight methyltransferase genes, and six aminotransferase genes were identified that may be related to dendrobine biosynthesis. Among them, one sesquiterpene synthase gene was found to be highly expressed after the treatment with MeJA and was positively correlated with the content of dendrobine. This study provides important and valuable metabolomics and transcriptomic information for the further understanding of D. nobile at the metabolic and molecular levels and provides candidate genes and possible intermediate compounds for the dendrobine biosynthesis pathway, which lays a certain foundation for further research on and application of Dendrobium.

Eight secondary metabolites including three new sesquiterpenoids, named dendronobin (1), dendrobilin L (2) and 1,4-trans-12-hydroxycalamenene-12-O-β-D-glucoside (3), together with five known compounds, dendrobine (4), nobiline (5), dihydroxyconiferyl alcohol (6), 5-methoxy-(+)-isolariciresinol (7), lyoniresinol (8) were isolated from the stems of Dendrobium nobile Lindl. Their structures were determined by extensive spectral analysis using HR-ESI-MS and NMR spectra (1H, 13C, 1H-1H COSY, HSQC, HMBC, and NOESY), and the absolute configurations of compounds 1-3 were determined by electronic circular dichroism (ECD) and theoretical calculations. Among these metabolites (1-8), dendronobin L (2) possessed a unique carbon skeleton. A potential biosynthetic pathway from dendronobin L (2) to dendrobine was suggested. Nobiline (5), dihydroxyconiferyl alcohol (6) and 5-methoxy-(+)-isolariciresinol (7) showed weak antioxidant activity.

The NAC (NAM, ATAF and CUC) gene family plays an important role in plant development and abiotic stress response. However, up to now, the identification and research of the NAC (PeNAC) family members of passion fruit are still lacking. In this study, 25 PeNACs were identified from the passion fruit genome, and their functions under abiotic stress and at different fruit-ripening stages were analyzed. Furthermore, we analyzed the transcriptome sequencing results of PeNACs under four various abiotic stresses (drought, salt, cold and high temperature) and three different fruit-ripening stages, and verified the expression results of some genes by qRT-PCR. Additionally, tissue-specific analysis showed that most PeNACs were mainly expressed in flowers. In particular, PeNAC-19 was induced by four various abiotic stresses. At present, low temperatures have seriously endangered the development of passion fruit cultivation. Therefore, PeNAC-19 was transformed into tobacco, yeast and Arabidopsis to study their function of resisting low temperature. The results show that PeNAC-19 responded to cold stress significantly in tobacco and Arabidopsis, and could improve the low temperature tolerance of yeast. This study not only improved the understanding of the PeNAC gene family characteristics and evolution, but also provided new insights into the regulation of the PeNAC gene at different stages of fruit maturation and abiotic stresses.