Search Results (57)

Background/Objectives: Anthocyanins, water-soluble flavonoid pigments with critical roles in plant stress resistance, are not naturally accumulated in cultivated tomato (Solanum lycopersicum) due to an incomplete flavonoid metabolism pathway. In contrast, ‘Black Pearl’ tomato exhibits distinct peel color transitions (from indigo rose to deep purple–red) during ripening, making it an ideal model for investigating the regulatory mechanisms of anthocyanin synthesis. A comprehensive strategy was employed to elucidate these mechanisms, involving transcriptomic (Illumina HiSeq), metabolomic (UPLC-MS/MS), and functional analyses of the ‘Black Pearl’ tomato peel across four developmental stages: mature green (S1), coloring (S2), purple immature (S3), and fully ripened (S4). Results: Transcriptome profiling identified 597 core differentially expressed genes (DEGs) associated with anthocyanin accumulation. Temporal analysis indicated that structural genes and activators peaked at S3, whereas repressive MYBs, including SlMYB76 which peaked at S2, exhibited staged expression. In parallel, metabolomic analysis identified 36 metabolites, with cyanidin and pelargonidin derivatives being characterized as the principal pigments. Functionally, SlMYB76 was confirmed to be a negative regulator, as its transient overexpression reduced anthocyanin content and downregulated SlANS. Mechanistically, direct binding and repression of the SlANS promoter by SlMYB76 were confirmed through yeast one-hybrid and dual-luciferase assays. Furthermore, its physical interaction with the bHLH factor SlJAF13 in the nucleus was demonstrated by Y2H, BiFC, LCI, and Co-IP, supporting the formation of a repressive complex that co-regulates SlANS. Conclusions: A novel SlMYB76-SlJAF13-SlANS regulatory module controlling anthocyanin accumulation in the peel of ‘Black Pearl’ tomato was identified. This discovery enhances the current understanding of the tomato flavonoid regulatory network and provides strategic targets for the genetic improvement of fruit color and anthocyanin content through molecular breeding. Full article

Isoflavone synthase (IFS) is the key enzyme in isoflavonoid biosynthesis and has been functionally characterized in numerous plant species. Glycyrrhiza species, valued for their medicinal properties, accumulate flavonoids with significant physiological activities. Among these, isoflavones play crucial roles in plant growth, development and stress responses. However, the IFS gene family in Glycyrrhiza remains poorly understood. In this study, we identified 10, 9 and 9 IFS genes in G. uralensis, G. inflata and G. glabra, respectively. Phylogenetic analysis classified these genes into four distinct clades (Clade A–D). Further characterization included chromosomal localization, gene structure, conserved motifs, cis-acting elements and synteny analysis. Using yeast one-hybrid (Y1H) screening, dual-luciferase assays and an electrophoretic mobility shift assay (EMSA), these results revealed that auxin response factor 4 (GgARF4) directly binds to the isoflavone synthase 9 (GgIFS9) promoter and activates its expression. Following indole-3-acetic acid (IAA) treatment, RNA-seq revealed that in the differentially expressed genes (DEGs), the genes involved in isoflavonoid and flavonoid biosynthesis pathways were significantly enriched. The result of quantitative reverse transcription polymerase chain reaction (qRT-PCR) revealed that GgIFS9 was strongly induced by IAA. β-Glucuronidase (GUS) assays confirmed that IAA activates the expression of the GgIFS9 promoter in Nicotiana tabacum. Our findings reveal that, through GgARF4 and its downstream-activated gene GgIFS9, IAA may promote flavonoid synthesis in G. glabra. This study provides novel insights into the auxin-mediated regulation of secondary metabolism in Glycyrrhiza species. Full article

Aeromonas veronii is a major pathogen threatening freshwater aquaculture, yet the molecular mechanisms of Pelteobagrus vachellii’s immune response to this infection remain unclear. This study integrated histopathology, mRNA-seq and small RNA-seq to investigate P. vachellii’s response to A. veronii at 48 h post-challenge. Histopathologically, infection induced gill epithelial detachment, hepatocyte swelling with cytoplasmic vacuolation, and melanomacrophage centers (MMCs) in the mid-kidney (histological assessment of the head kidney was not feasible due to sampling limitations associated with its small size). Transcriptomic analysis identified 1210 differentially expressed genes (DEGs) in the head kidney (819 downregulated, 391 upregulated), significantly enriched in 11 immune pathways (e.g., NF-κB, Th17 cell differentiation, Complement and coagulation cascades), with key immune genes (e.g., IL-1β, TCRα, CCL4) upregulated. Gene Set Enrichment Analysis (GSEA) revealed activation of the proteasome, ribosome and oxidative phosphorylation pathways, and suppression of the autophagy-animal, FoxO and AMPK pathways. Small RNA-seq identified 544 known and 958 novel miRNAs in the head kidney, with 42 downregulated and 36 upregulated differentially expressed miRNAs (DE miRNAs). The miRNA-mRNA network showed that DE miRNAs (e.g., miR-101-y/z, miR-132-z, miR-3167-y) negatively regulated immune-related target genes (IL-1R1, IRF4, IκBα) in core immune pathways. Collectively, this study clarifies the pathological and miRNA-mRNA regulatory modules of P. vachellii head kidney against A. veronii infection, providing valuable information that enables the further analyses of the defense mechanisms of P. vachellii against A. veronii infection. Full article

Bisphenol A (BPA) replacement chemicals are used in products like food packaging, plastic piping, and sportswear. While they can be toxic, their neurotoxicity is less understood. The aim of this study was to treat differentiated human SH-SY5Y cells with Bisphenol S (BPS) and Bisphenol F (BPF) to investigate mechanisms of toxicity. BPS reduced cell viability (>50 µM at 48 h) more than BPF (>200 µM at 48 h), with concentration- and time-dependent effects. Both induced caspase 3/7 activity at 250 µM after 48 h, though no changes were observed in levels of reactive oxygen species nor mitochondrial ATPase activity. RNA-seq analysis at 0.1 nM revealed distinct transcriptional networks: BPS altered IL15R, causing NF-kB/NFATC activation, and triggered NF-kB signaling through CD8, while BPF affected TLR9 and activated NF-kB targets through TNF. Pathway analysis showed that genes involved in neuroinflammation, protein folding, microglial function, and motor neuron regulation were disrupted, demonstrating that BPS and BPF, even at low, environmentally relevant concentrations, significantly alter gene expression in pathways linked to neuroinflammation, immune signaling, and neurodegenerative diseases. BPS primarily affected ribosomal and immune-related networks, while BPF disrupted oxidative phosphorylation and protein-folding pathways. These alterations suggest mechanisms for long-term neurological effects, highlighting the need for comprehensive evaluations of BPA alternatives. Full article

Plant heat shock proteins (Hsps) are from a diverse and ancient protein family, with small Hsps of ~20 kDa molecular weight classified as Hsp20s. As a key transcription factor in the jasmonic acid (JA) pathway, myelocytomatosis protein 2 (MYC2) plays a vital role in stamen development. In this study, we identified six genes with significantly altered expression levels using previous RNA-Seq data from PhMYC2a-overexpressing and methyl jasmonate (MeJA)-treated petunia. Interestingly, five of these are Hsp20 family members (PhHsp16.0A, PhHsp16.1, PhHsp16.8, PhHsp21.9, and PhHsp40.8). Yeast one-hybrid (Y1H) and dual-luciferase assays demonstrated that PhMYC2a directly binds their promoters, indicating a collective effect. Thus, a genome-wide analysis was conducted and a total of 38 genes encoding Hsp20s were identified in the reference genome of Petunia axillaris. Phylogenetic analysis revealed that 38 members of Hsp20s were irregularly distributed on 34 chromosome scaffolds and separated into 13 subfamilies, with only PaHsp16.0A and 16.1, among the five selected Hsp20s, being in the same Cytosol IV (CIV) subfamily. Conserved motif analysis suggested that the PaHsp20 gene family members may have a high degree of conservation. The promoter sequence analysis suggested that the promoter regions of PaHsp20 genes contained multiple light- and hormone-related cis-regulatory elements. Subsequently, spatiotemporal expression patterns, analyzed by qRT-PCR, showed that PhHsp16.0A and PhHsp16.1 had relatively high expression levels in flowers, with similar expression patterns at various stages of flower bud and anther development. Furthermore, virus-induced gene silencing (VIGS) of PhHsp16.0A and PhHsp16.1 resulted in significantly reduced pollen fertility, indicating their regulation in the process of flower development and echoing the role of PhMYC2a. This study highlights the pivotal role of Hsp20s in MYC2a-mediated regulatory mechanisms during petunia pollen development. Full article

Obstructive sleep apnea syndrome (OSAS) is characterized by cycles of decreased blood oxygen saturation followed by reoxygenation due to transient apnea. Cognitive dysfunction is a complication of OSAS, but its mechanisms remain unclear. Eight-week-old C57BL/6J mice were exposed to intermittent hypoxia (IH) to model OSAS, and cognitive function and hippocampal gene expression were analyzed. Three groups were maintained for 28 days: an IH group (oxygen alternating between 10 and 21% in 2 min cycles, 8 h/day), sustained hypoxia group (SH) (10% oxygen, 8 h/day), and control group (21% oxygen). Behavioral tests and RNA sequencing (RNA-seq) analysis were performed. While Y-maze test results showed no differences, the IH group demonstrated impaired memory and learning in passive avoidance tests compared to control and SH groups. RNA-seq revealed coordinated suppression of mitochondrial function genes and oxidative stress response pathways, specifically in the IH group. RT-qPCR showed decreased Lars2, Hmcn1, and Vstm2l expression in the IH group. Pathway analysis showed the suppression of the KEAP1-NFE2L2 antioxidant pathway in the IH group vs. the SH group. Our findings demonstrate that IH induces cognitive dysfunction through suppression of the KEAP1-NFE2L2 antioxidant pathway and downregulation of mitochondrial genes (Lars2, Vstm2l), leading to oxidative stress and mitochondrial dysfunction. These findings advance our understanding of the molecular basis underlying OSAS-related cognitive impairment. Full article

Phalaenopsis pulcherrima are known for their captivating floral morphology and diverse colors, demonstrate exceptional resilience to adverse environmental conditions, and exhibit significant potential for hybrid breeding. However, current research on flower coloration is still limited. The data from this study indicates that variations in anthocyanin levels are the primary determinants of the difference between white and purple colors. Through RNA-seq, we identified 469 genes that were differentially expressed. Furthermore, our bioinformatics exploration uncovered two potential transcription factors, PpMYB1 and PpbHLH1, which play regulatory roles in anthocyanin accumulation. Y2H assays demonstrated that these two TFs could form heterodimers and interact with each other. Afterwards, transient expression assays were conducted for the first time in P. pulcherrima flowers, revealing that overexpression of PpMYB1 alone or in combination with PpbHLH1 resulted in purple petal pigmentation. Overexpressing PpMYB1 in tobacco resulted in more purple-colored corollas, stamens, pistils, and pods compared to control plants. Y1H and dual-luciferase assays provided further evidence that PpMYB1 and PpbHLH1 interact with the promoters of the structural genes PpF3H, PpDFR, and PpANS in the anthocyanin biosynthesis pathway, thereby driving their robust expression. This study not only enhances our understanding of the molecular mechanisms underlying anthocyanin synthesis but also holds significant practical implications for advancing plant hybrid breeding and genetic engineering applications in flower color regulation. Full article

Gaseous chlorine dioxide (ClO₂) is a potent antimicrobial agent used to control microbial contamination in food and water. This study evaluates the bactericidal activity of gaseous ClO₂ released from a sodium chlorite (NaClO₂) pad against Campylobacter jejuni. Exposure to a low concentration (0.4 mg/L) of dissolved ClO₂ for 2 h resulted in a >93% reduction of C. jejuni, highlighting the bacterium’s extreme sensitivity to gaseous ClO₂. To elucidate the molecular mechanism of ClO₂-induced bactericidal action, transcriptomic analysis was conducted using RNA sequencing (RNA-seq). The results indicate that C. jejuni responds to ClO₂-induced oxidative stress by upregulating genes involved in reactive oxygen species (ROS) detoxification (sodB, ahpC, katA, msrP, and trxB), iron transport (ceuBCD, cfbpABC, and chuBCD), phosphate transport (pstSCAB), and DNA repair (rdgB and mutY). Reverse transcription-quantitative PCR (RT-qPCR) validated the increased expression of oxidative stress response genes but not general stress response genes (spoT, dnaK, and groES). These findings provide insights into the antimicrobial mechanism of ClO₂, demonstrating that oxidative damage to essential cellular components results in bacterial cell death. Full article

Background/Objectives: Pasteurella multocida commonly colonizes the bovine respiratory tract and can occasionally cause intramammary infections. Here, eight P. multocida isolates from clinical cases of bovine mastitis were investigated for their molecular characteristics as well as phenotypic and genotypic antimicrobial resistance (AMR) properties. Methods: The isolates originated from quarter milk samples obtained in Germany for diagnostic purposes. Antimicrobial susceptibility testing (AST) by broth microdilution was performed according to the Clinical and Laboratory Standards Institute. Closed whole-genome sequences were generated by hybrid assembly of Illumina MiSeq short-reads and Oxford Nanopore MinION long-reads, followed by consecutive sequence analysis. Results: The P. multocida isolates belonged either to capsular:lipopolysaccharide type A:3 (n = 7) or A:6 (n = 1), and multi-locus sequence types 1 (n = 7) or 7 (n = 1). Seven isolates carried AMR genes, such as mef(C), mph(G), strA, strB, aphA1, aadA31, tet(H), tet(Y), floR, catA3, and sul2, as part of an integrative and conjugative element (ICE). These mobile genetic elements, 58,382–78,401 bp in size, were highly similar to the ICEs Tn7406 or Tn7407 that have been previously described in bovine Mannheimia haemolytica and P. multocida, respectively. Moreover, the isolates showed elevated minimal inhibitory concentrations corresponding to the identified AMR determinants. Conclusions: Molecular typing and ICE organization suggest the bovine respiratory tract as reservoir of the investigated mastitis-associated P. multocida. Horizontal cross-genus transfer of multidrug-resistance-mediating ICEs seems to occur under in vivo conditions among different pathogens from cattle in Germany, which underlines the importance of pathogen identification followed by AST for successful bovine mastitis therapy. Full article

Pepper (Capsicum spp.) is an important global vegetable and spice, with fruit color being a key determinant of its commercial quality. However, the regulatory mechanisms underlying pepper fruit color are still not fully understood. This study focuses on the MADS-RIPENING INHIBITOR (MADS-RIN), a MADS-box transcription factor that regulates various aspects of fruit ripening, including pigmentation. We identified CaRIN, a homolog of tomato’s SlRIN, whose expression is closely associated with fruit ripening in pepper. Silencing CaRIN through virus-induced gene silencing (VIGS) resulted in increased chlorophyll and chlorophyll a content, reduced carotenoid accumulation, and uneven fruit coloration. Integrative analysis of the RNA-seq and DAP-seq data identified 77 target genes regulated by CaRIN, which was involved in processes such as chlorophyll metabolism and plant hormone signaling. Yeast one-hybrid (Y1H) and dual-luciferase (LUC) assays demonstrated that CaRIN directly bound to the promoter of CaLhcb-P4, repressing its expression. Downregulation of CaLhcb-P4 in pepper fruits via VIGS accelerated chlorophyll degradation. Additionally, CaRIN indirectly regulated multiple genes associated with chlorophyll and carotenoid metabolism, sugar transport, and cell wall degradation. These findings provide novel insights into the regulatory mechanisms of chlorophyll degradation during pepper fruit ripening, offering a foundation for further research and potential genetic improvement strategies. Full article

Background/Objectives: Apis cerana development is described as comprising four stages: embryo, larva, pupa, and adult. There are significant differences between workers and drones in terms of physiological functions and social roles, and the formation of the organ primordia occurs during the embryonic stage. Therefore, the objective of this study is to investigate the differential expression of and alternative splicing of genes in worker and drone embryos and to explain their unique developmental patterns. Methods: Long-read sequencing (PacBio Iso-Seq) and short-read sequencing (Illumina RNA-Seq) were used to investigate worker and drone embryo gene expression differences in A. cerana across five developmental points (12, 24, 36, 48, and 60 h). Results: The study identified 59,254 common isoforms, with 5744 and 5106 isoforms specific to worker and drone embryos, respectively. Additionally, a new transcript of the csd gene was identified. The number of differentially expressed genes (3391) and differential splicing events (470 genes) peaked at the 24-h embryonic stage. Differential splicing events of csd, dsx, and Y-y were observed in the worker and drone embryos. Conclusions: The gene expression results indicated that the 24-h embryonic point is a critical period for the expression of genes related to developmental and behavioral differences between workers and drones. The findings provide a theoretical basis for future research on the developmental differences between workers and drones. Full article

The bHLH (basic Helix–Loop–Helix) transcription factor serves as pivotal controller in plant growth and development. In a previous study, the overexpression of SlUPA-like in Solanum lycopersicum L. Ailsa Craig (AC⁺⁺) altered the JA (Jasmonic acid) response and endogenous GA (Gibberellic acid) content. However, the detailed regulation mechanism was not fully explored. In the present research, we found that the overexpression of SlUPA-like influenced the accumulation of GA, JA and BR (Brassinolide). RNA-Seq data illustrated that the expression levels of genes related to these plant hormones were significantly affected. Additionally, the interaction of SlUPA-like with SlMYB21, SlMYC2 and SlDELLA was characterized by employing Y2H (Yeast Two-Hybrid) and BiFC (Bimolecular Fluorescence Complementation) assay. Furthermore, Dual-LUC (Dual-Luciferase) assay and EMSA (Electrophoretic Mobility Shift Assay) identified that SlUPA-like directly targeted the E-box motif in the promoter of SlGID2 and activated the transcription of SlGID2. These results shed light on the potential role of SlUPA-like in mediating crosstalk among multiple plant hormones and established a robust theoretical framework for further unraveling the functions of SlUPA-like transcription factors in the context of plant growth and hormone signal transduction. Full article

Phytochrome-interacting factor (PIF) is a subfamily of the basic helix–loop–helix (bHLH) transcription factors (TFs) and plays key roles in plant responses to diverse biotic and abiotic stresses. In this work, a PIF gene named CaPIF7a was cloned and its role in the regulation of pepper’s resistance to Phytophthora capsici infection (PCI) was studied. The cloned CaPIF7a gene has a CDS length of 1383 bp, encodes a hydrophilic protein containing bHLH and APB characteristic domains, and subcellular localization results showed that CaPIF7a was located in the nucleus. Expression analysis showed that CaPIF7a gene has the highest expression level in leaf, and its expression was regulated under PCI and salicylic acid (SA) treatment. Silencing of CaPIF7a in pepper plants by virus-induced gene silencing (VIGS) reduces the resistance of pepper to PCI, with decreased expression of SA-responsive and SA-biosynthesis genes and obviously decreased SA content. DNA affinity purification sequencing (DAP-seq) was employed to identify the potential targets of CaPIF7a, and yeast one-hybrid (Y1H) verified that CaPIF7a could regulate the expression of CaHY5 by binding its promoter. These findings indicated that CaPIF7a might be a key modulator in plant immune response and presented a possible regulatory network of CaPIF7a in PCI. Full article

Sulfur dioxide (SO₂) is the most effective preservative for table grapes as it reduces the respiratory intensity of berries and inhibits mold growth. However, excessive SO₂ causes berry abscission during storage, resulting in an economic loss postharvest. In this study, grapes were exogenously treated with SO₂, SO₂ + 1.5% chitosan, SO₂ + 1.5% eugenol, and SO₂ + eugenol-loaded chitosan nanoparticles (SN). In comparison to SO₂ treatment, SN treatment reduced the berries’ abscission rate by 74% while maintaining the quality of the berries. Among the treatments, SN treatment most effectively inhibited berry abscission and maintained berry quality. RNA-sequencing (RNA-seq) revealed that SN treatment promoted the expression of genes related to cell wall metabolism. Among these genes, VlCOMT was detected as the central gene, playing a key role in mediating the effects of SN. Dual luciferase and yeast one-hybrid (Y1H) assays demonstrated that VlbZIP14 directly activated VlCOMT by binding to the G-box motif in the latter’s promoter, which then participated in lignin synthesis. Our results provide key insights into the molecular mechanisms underlying the SN-mediated inhibition of berry abscission and could be used to improve the commercial value of SO₂-treated postharvest table grapes. Full article

Catharanthus roseus (C. roseus) can produce over 150 types of monoterpenoid indole alkaloids (MIAs), including vinblastine and vincristine, which are currently the primary sources of these alkaloids. Exploring the complex regulatory mechanisms of C. roseus is significant for resolving MIA biosynthesis. The Lateral Organ Boundaries Domain (LBD) is a plant-specific transcription factor family that plays crucial roles in the physiological processes of plant growth, stress tolerance, and specialized metabolism. However, the LBD gene family has not been extensively characterized in C. roseus, and whether its members are involved in MIA biosynthesis is still being determined. A total of 34 C. roseus LBD (CrLBD) genes were identified. RNA-Seq data were investigated to examine the expression patterns of CrLBD genes in various tissues and methyl jasmonate (MeJA) treatments. The results revealed that the Class Ia member CrLBD4 is positively correlated with iridoid biosynthetic genes (p < 0.05, r ≥ 0.8); the Class IIb member CrLBD11 is negatively correlated with iridoid biosynthetic genes (p < 0.05, r ≤ −0.8). Further validation in leaves at different growth stages of C. roseus showed that CrLBD4 and CrLBD11 exhibited different potential expression trends with iridoid biosynthetic genes and the accumulation of vindoline and catharanthine. Yeast one-hybrid (Y1H) and subcellular localization assays demonstrated that CrLBD4 and CrLBD11 could bind to the “aattatTCCGGccgc” cis-element and localize to the nucleus. These findings suggest that CrLBD4 and CrLBD11 may be potential candidates for regulating MIA biosynthesis in C. roseus. In this study, we systematically analyzed the CrLBD gene family and provided insights into the roles of certain CrLBDs in the MIA biosynthesis of C. roseus. Full article