Therapeutic effect of Hosta plantaginea (Lam.) Aschers flowers on acute pharyngitis through inhibition of multi-inflammatory pathways in rats.

BACKGROUND: Hosta plantaginea (Lam.) Aschers flower is a famous Mongolian folk medicine in China and has a therapeutic effect on acute pharyngitis (AP). However, the effect and potential mechanism of H. plantaginea flower on AP have not been fully elucidated. AIM OF THE STUDY: The present work aimed to evaluate the effects and mechanisms of the crude extract of H. plantaginea flowers (HP) and its four fractions of petroleum ether fraction (HPA), ethyl acetate fraction (HPB), n-butanol fraction (HPC), and water residue (HPD) against AP in rats. MATERIALS AND METHODS: A 15% ammonia-induced AP rat model in rats was established. Therapeutic effects of HP and HPA∼D in model rats were evaluated based on body weight, histopathological analysis, and inflammatory parameters, including tumor necrosis factor α (TNF-α), prostaglandin E2 (PGE2), interleukin 1ß (IL-1ß), and IL-6. The protein expression of nuclear factor kappa-B p65 (NF-κB p65), inhibitor of NF-κB alpha (IκBα), c-Jun N-terminal kinases (JNK), mitogen-activated protein kinase (MAPK) p38, extracellular signal-regulated kinase (Erk), just another kinase 1 (JAK1), signal transducer and activator of transcription 3 (STAT3), phosphoinositide 3-kinase (PI3K), and protein kinase B (Akt) were detected by a Western blotting assay. RESULTS: HP, HPB, and HPC treatments markedly alleviated AP in rats by increasing body weight and improving pathological damages in pharyngeal tissues. In addition, HP, HPB, and HPC treatments significantly inhibited inflammation, including decreasing the levels of TNF-α, PGE2, IL-1ß, and IL-6, and suppressing phosphorylated protein expression of p65, IκBα, JNK, p38, Erk, JAK1, STAT3, PI3K, and Akt in pharyngeal tissues of rats. CONCLUSION: Collectively, HP, HPB, and HPC can attenuate pharynx injury in rats by suppressing inflammation via inhibition of NF-κB, MAPKs, JAK-STAT, and PI3K-Akt pathways, which supports the traditional use of H. plantaginea flowers.

Extraction optimization and constituent analysis of total flavonoid from Hosta plantaginea (Lam.) Aschers flowers and its ameliorative effect on chronic prostatitis via inhibition of multiple inflammatory pathways in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Hosta plantaginea (Lam.) Aschers flowers (HPF) are well-known for their high flavonoid content, which contribute to their widely as traditional Chinese medicine for alleviating inflammation. Despite their recognized potential, information regarding the total flavonoid (TF) of HPF and its therapeutic application in treating chronic prostatitis (CP) remains unknown. AIM OF THE STUDY: We aimed to investigate the extraction optimization, constituent analysis, and alleviating effect of TF on CP as well as its potential mechanism. MATERIALS AND METHODS: The optimized extraction of TF from HPF was explored using response surface methodology with a Box-Behnken design model. The major flavonoids in TF were identified based on UHPLC-MS approach. Efficacy of TF (25 and 100 mg/kg, p.o.) on CP was evaluated in prostate antigen emulsion-induced autoimmune CP rat model by measuring prostatic index, the levels of leukocytes and lecithin bodies, as well as histopathological examination. The protein expression contents were detected by western blotting. Additionally, the antioxidant (DPPH and ABTS) and anti-inflammatory (cyclooxygenase 2, COX-2 inhibitory) effects of TF were also evaluated in vitro. RESULTS: The optimized conditions for TF extraction were determined as 60% ethanol concentration, 30 mL/g liquid-to-solid ratio, 30 min extraction time, and 90 °C extraction temperature, and the extraction ratio is 65.98 ± 2.14%. A total of 15 major flavonoids in TF were characterized by comparison with reference standards. TF ameliorated the efficacy of CP in rats in a dose-independent manner, including reduced prostatic index and leukocytes levels, elevated lecithin body levels, ameliorated histopathological damage to prostate, and suppressed phosphorylated protein expressions of nuclear factor kappa-B (NF-κB) p65, inhibitor of NF-κB alpha (IκBα), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase (Erk), just another kinase 1 (JAK1), signal transducer and activator of transcription 3 (STAT3), phosphoinositide 3-kinase (PI3K) and protein kinase B (Akt). Simultaneously, the IC50 of TF to DPPH, ABTS radicals, and COX-2 were 2.02, 1.79, and 0.0838 mg/mL, respectively. CONCLUSIONS: We first demonstrated that TF from HPF represents a promising candidate to alleviate CP through suppression of NF-κB, MAPKs, JAK-STAT, and PI3K-Akt signaling pathways.

Association Analysis of Transcriptome and Targeted Metabolites Identifies Key Genes Involved in Iris germanica Anthocyanin Biosynthesis.

The anthocyanin biosynthetic pathway is the main pathway regulating floral coloration in Iris germanica, a well-known ornamental plant. We investigated the transcriptome profiles and targeted metabolites to elucidate the relationship between genes and metabolites in anthocyanin biosynthesis in the bitone flower cultivar 'Clarence', which has a deep blue outer perianth and nearly white inner perianth. In this study, delphinidin-, pelargonidin-, and cyanidin-based anthocyanins were detected in the flowers. The content of delphinidin-based anthocyanins increased with the development of the flower. At full bloom (stage 3), delphinidin-based anthocyanins accounted for most of the total anthocyanin metabolites, whereas the content of pelargonidin- and cyanidin-based anthocyanins was relatively low. Based on functional annotations, a number of novel genes in the anthocyanin pathway were identified, which included early biosynthetic genes IgCHS, IgCHI, and IgF3H and late biosynthetic genes Ig F3'5'H, IgANS, and IgDFR. The expression of key structural genes encoding enzymes, such as IgF3H, Ig F3'5'H, IgANS, and IgDFR, was significantly upregulated in the outer perianth compared to the inner perianth. In addition, most structural genes exhibited their highest expression at the half-color stage rather than at the full-bloom stage, which indicates that these genes function ahead of anthocyanins synthesis. Moreover, transcription factors (TFs) of plant R2R3-myeloblastosis (R2R3-MYB) related to the regulation of anthocyanin biosynthesis were identified. Among 56 R2R3-MYB genes, 2 members belonged to subgroup 4, with them regulating the expression of late biosynthetic genes in the anthocyanin biosynthetic pathway, and 4 members belonged to subgroup 7, with them regulating the expression of early biosynthetic genes in the anthocyanin biosynthetic pathway. Quantitative real-time PCR (qRT-PCR) analysis was used to validate the data of RNA sequencing (RNA-Seq). The relative expression profiles of most candidate genes were consistent with the FPKM of RNA-seq. This study identified the key structural genes encoding enzymes and TFs that affect anthocyanin biosynthesis, which provides a basis and reference for the regulation of plant anthocyanin biosynthesis in I. germanica.

PeCIN8 expression correlates with flower size and resistance to yellow leaf disease in Phalaenopsis orchids.

BACKGROUND: The orchid industry has seen a recent surge in export values due to the floral morphology and versatile applications of orchids in various markets for medicinal, food additive, and cosmetic usages. However, plant-related diseases, including the yellow leaf disease caused by Fusarium solani, have caused significant losses in the production value of Phalaenopsis (up to 30%). RESULTS: In this study, 203 Phalaenopsis cultivars were collected from 10 local orchid nurseries, and their disease severity index and correlation with flower size were evaluated. Larger flowers had weaker resistance to yellow leaf disease, and smaller flowers had stronger resistance. For the genetic relationship of disease resistance to flower size, the genetic background of all cultivars was assessed using OrchidWiz Orchid Database Software and principal component analysis. In addition, we identified the orthologous genes of BraTCP4, namely PeIN6, PeCIN7, and PeCIN8, which are involved in resistance to pathogens, and analyzed their gene expression. The expression of PeCIN8 was significantly higher in the most resistant cultivars (A7403, A11294, and A2945) relative to the most susceptible cultivars (A10670, A6390, and A10746). CONCLUSIONS: We identified a correlation between flower size and resistance to yellow leaf disease in Phalaenopsis orchids. The expression of PeCIN8 may regulate the two traits in the disease-resistant cultivars. These findings can be applied to Phalaenopsis breeding programs to develop resistant cultivars against yellow leaf disease.

INDETERMINATE1-mediated expression of FT family genes is required for proper timing of flowering in Brachypodium distachyon.

The transition to flowering is a major developmental switch in plants. In many temperate grasses, perception of indicators of seasonal change, such as changing day-length and temperature, leads to expression of FLOWERING LOCUS T1 (FT1) and FT-Like (FTL) genes that are essential for promoting the transition to flowering. However, little is known about the upstream regulators of FT1 and FTL genes in temperate grasses. Here, we characterize the monocot-specific gene INDETERMINATE1 (BdID1) in Brachypodium distachyon and demonstrate that BdID1 is a regulator of FT family genes. Mutations in ID1 impact the ability of the short-day (SD) vernalization, cold vernalization, and long-day (LD) photoperiod pathways to induce certain FTL genes. BdID1 is required for upregulation of FTL9 (FT-LIKE9) expression by the SD vernalization pathway, and overexpression of FTL9 in an id1 background can partially restore the delayed flowering phenotype of id1. We show that BdID1 binds in vitro to the promoter region of FTL genes suggesting that ID1 directly activates FTL expression. Transcriptome analysis shows that BdID1 is required for FT1, FT2, FTL12, and FTL13 expression under inductive LD photoperiods, indicating that BdID1 is a regulator of the FT gene family. Moreover, overexpression of FT1 in the id1 background results in rapid flowering similar to overexpressing FT1 in the wild type, demonstrating that BdID1 is upstream of FT family genes. Interestingly, ID1 negatively regulates a previously uncharacterized FTL gene, FTL4, and we show that FTL4 is a repressor of flowering. Thus, BdID1 is critical for proper timing of flowering in temperate grasses.

Effects of Magnoliae Flos on Atopic Dermatitis-Like Inflammation Evaluated via Extracellular Signal-regulated Kinase or Signal Transducers and Activators of Transcription 1/3 Signalling Pathways.

Atopic dermatitis is a chronic inflammatory skin  disease. Skin is the largest organ and plays a pivotal role in protecting the body. Not only does the skin act as a physical barrier against the external environment, but it also has its own immune system. Atopic dermatitis is caused by prolonged excessive inflammatory responses that worsen under imbalanced cutaneous immune system skin conditions. Although the prevalence and burden of atopic dermatitis is increasing, the standard therapeutic agents remain unclear due to  the complicated pathophysiology of the condition. The objective of this study is to examine the use of Magnoliae flos, the dried flower bud of Magnolia biondii or  related plants. The effects and underlying mechanism of  action of aqueous extract of the buds of Magnoliae flos (MF) were evaluated. Immortalized human keratinocytes (HaCaT) stimulated with tumour necrosis factor-α and interferon-γ mixture and NC/Nga mice stimulated with 2,4-dinitrochlorobenzene were used as atopic dermatitis models, in vitro and in vivo, respectively. The effects of MF were determined by measuring the suppression of pro-inflammatory signalling pathways, such as extracellular signal-regulated kinase or signal transducers and activators of transcription 1/3 and restoring skin barrier molecules. In conclusion, MF is a potential therapeutic alternative for the treatment of atopic dermatitis through repressing inflammatory pathways.

Temperature-responsive module of OfAP1 and OfLFY regulates floral transition and floral organ identity in Osmanthus fragrans.

The MADS-box transcription factor APETELA1 (AP1) is crucially important for reproductive developmental processes. The function of AP1 and the classic LFY-AP1 interaction in woody plants are not widely known. Here, the OfAP1-a gene from the continuously flowering plant Osmanthus fragrans 'Sijigui' was characterized, and its roles in regulating flowering time, petal number robustness and floral organ identity were determined using overexpression in Arabidopsis thaliana and Nicotiana tabacum. The expression of OfAP1-a was significantly induced by low ambient temperature and was upregulated with the floral transition process. Ectopic expression OfAP1-a revealed its classic function in flowering and flower ABC models. The expression of OfAP1-a is inhibited by LEAFY (OfLFY) through direct promoter binding, as confirmed by yeast one-hybrid and dual luciferase assays. Arabidopsis plants overexpressing OfAP1-a exhibited accelerated flowering and altered floral organ identities. Moreover, OfAP1-a-overexpressing plants displayed variable petal numbers. Likewise, the overexpression of OfLFY in Arabidopsis and Nicotiana altered petal number robustness and inflorescence architecture, partially by regulating native AP1 in transformed plants. Furthermore, we performed RNA-seq analysis of transgenic Nicotiana plants. DEGs were identified by transcriptome analysis, and we found that the expression of several floral homeotic genes was altered in both OfAP1-a and OfLFY-overexpressing transgenic lines. Our results suggest that OfAP1-a may play important roles during floral transition and development in response to ambient temperature. OfAP1-a functions as a petal number modulator and may directly activate a subset of flowers to regulate floral organ formation. OfAP1-a and OfLFY mutually regulate the expression of each other and coregulate genes that might be involved in these phenotypes related to flowering. The results provide valuable data for understanding the function of the LFY-AP1 module in the reproductive process and shaping floral structures in woody plants.

Functional Characterization of AP2/ERF Transcription Factors during Flower Development and Anthocyanin Biosynthesis Related Candidate Genes in Lycoris.

The APETALA2/ethylene-responsive transcription factor (AP2/ERF) family has been extensively investigated because of its significant involvement in plant development, growth, fruit ripening, metabolism, and plant stress responses. To date, there has been little investigation into how the AP2/ERF genes influence flower formation and anthocyanin biosynthesis in Lycoris. Herein, 80 putative LrAP2/ERF transcription factors (TFs) with complete open reading frames (ORFs) were retrieved from the Lycoris transcriptome sequence data, which could be divided into five subfamilies dependent on their complete protein sequences. Furthermore, our findings demonstrated that genes belonging to the same subfamily had structural similarities and conserved motifs. LrAP2/ERF genes were analyzed for playing an important role in plant growth, water deprivation, and flower formation by means of gene ontology (GO) enrichment analysis. The expression pattern of the LrAP2/ERF genes differed across tissues and might be important for Lycoris growth and flower development. In response to methyl jasmonate (MeJA) exposure and drought stress, the expression of each LrAP2/ERF gene varied across tissues and time. Moreover, a total of 20 anthocyanin components were characterized using ultra-performance liquid chromatography-electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS) analysis, and pelargonidin-3-O-glucoside-5-O-arabinoside was identified as the major anthocyanin aglycone responsible for the coloration of the red petals in Lycoris. In addition, we mapped the relationships between genes and metabolites and found that LrAP2/ERF16 is strongly linked to pelargonidin accumulation in Lycoris petals. These findings provide the basic conceptual groundwork for future research into the molecular underpinnings and regulation mechanisms of AP2/ERF TFs in anthocyanin accumulation and Lycoris floral development.

Integrated Metabolome and Transcriptome Analysis of Petal Anthocyanin Accumulation Mechanism in Gloriosa superba 'Rothschildiana' during Different Flower Development Stages.

Flower color is a key ornamental trait in plants. The petals of Gloriosa superba 'Rothschildiana' petals undergo a color transformation from yellow to red during their development, but the molecular mechanism of this process remains unexplored. This study examines the anthocyanin profiles and gene expression patterns of 'Rothschildiana' petals across four developmental stages: bud (S1), initial opening (S2), half opening (S3), and full opening stage (S4). A total of 59 anthocyanins were identified with significant increases in cyanidin-3,5-O-diglucoside, cyanidin-3-O-glucoside, pelargonidin-3-O-glucoside, and pelargonidin-3,5-O-diglucoside levels observed during petal maturation. Transcriptome analysis revealed 46 differentially expressed genes implicated in flavonoid and anthocyanin biosynthesis. Additionally, three gene modules were found to be associated with anthocyanin accumulation throughout flower development. Expression levels of genes associated with auxin, abscisic acid, brassinosteroid signaling, and transcription factors such as NACs and WRKYs underwent significant changes and exhibited strong correlations with several flavonoid and anthocyanin biosynthetic genes in these modules. These findings offer novel insights into the molecular underpinnings of flower color variation and lay the groundwork for the improvement of G. superba.

The Abundant and Unique Transcripts and Alternative Splicing of the Artificially Autododecaploid London Plane (Platanus × acerifolia).

Transcription and alternative splicing (AS) are now appreciated in plants, but few studies have examined the effects of changing ploidy on transcription and AS. In this study, we showed that artificially autododecaploid plants of London plane (Platanus × acerifolia (Aiton) Willd) had few flowers relative to their hexaploid progenitors. Transcriptome analysis based on full-length Oxford Nanopore Technologies (ONTs) and next-generation sequencing (NGS) revealed that the increased ploidy level in P. × acerifolia led to more transcribed isoforms, accompanied by an increase in the number of isoforms per gene. The functional enrichment of genes indicated that novel genes transcribed specifically in the dodecaploids may have been highly correlated with the ability to maintain genome stability. The dodecaploids showed a higher number of genes with upregulated differentially expressed genes (DEGs) compared with the hexaploid counterpart. The genome duplication of P. × acerifolia resulted mainly in the DEGs involved in basic biological pathways. It was noted that there was a greater abundance of alternative splicing (AS) events and AS genes in the dodecaploids compared with the hexaploids in P. × acerifolia. In addition, a significant difference between the structure and expression of AS events between the hexaploids and dodecaploids of Platanus was found. Of note, some DEGs and differentially spliced genes (DSGs) related to floral transition and flower development were consistent with the few flower traits in the dodecaploids of P. × acerifolia. Collectively, our findings explored the difference in transcription and AS regulation between the hexaploids and dodecaploids of P. × acerifolia and gained new insight into the molecular mechanisms underlying the few-flower phenotype of P. × acerifolia. These results contribute to uncovering the regulatory role of transcription and AS in polyploids and breeding few-flower germplasms.

Heterologous Expression of Platycodon grandiflorus PgF3'5'H Modifies Flower Color Pigmentation in Tobacco.

Flavonoid-3',5'-hydroxylase (F3'5'H) is the key enzyme for the biosynthesis of delphinidin-based anthocyanins, which are generally required for purple or blue flowers. Previously, we isolated a full-length cDNA of PgF3'5'H from Platycodon grandiflorus, which shared the highest homology with Campanula medium F3'5'H. In this study, PgF3'5'H was subcloned into a plant over-expression vector and transformed into tobacco via Agrobacterium tumefaciens to investigate its catalytic function. Positive transgenic tobacco T0 plants were obtained by hygromycin resistance screening and PCR detection. PgF3'5'H showed a higher expression level in all PgF3'5'H transgenic tobacco plants than in control plants. Under the drive of the cauliflower mosaic virus (CaMV) 35S promoter, the over-expressed PgF3'5'H produced dihydromyricetin (DHM) and some new anthocyanin pigments (including delphinidin, petunidin, peonidin, and malvidin derivatives), and increased dihydrokaempferol (DHK), taxifolin, tridactyl, cyanidin derivatives, and pelargonidin derivatives in PgF3'5'H transgenic tobacco plants by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) analysis, resulting in a dramatic color alteration from light pink to magenta. These results indicate that PgF3'5'H products have F3'5'H enzyme activity. In addition, PgF3'5'H transfer alters flavonoid pigment synthesis and accumulation in tobacco. Thus, PgF3'5'H may be considered a candidate gene for gene engineering to enhance anthocyanin accumulation and the molecular breeding project for blue flowers.

Protamine folds DNA into flowers and loop stacks.

DNA in sperm undergoes an extreme compaction to almost crystalline packing levels. To produce this dense packing, DNA is dramatically reorganized in minutes by protamine proteins. Protamines are positively charged proteins that coat negatively charged DNA and fold it into a series of toroids. The exact mechanism for forming these ∼50-kbp toroids is unknown. Our goal is to study toroid formation by starting at the "bottom" with folding of short lengths of DNA that form loops and working "up" to more folded structures that occur on longer length scales. We previously measured folding of 200-300 bp of DNA into a loop. Here, we look at folding of intermediate DNA lengths (L = 639-3003 bp) that are 2-10 loops long. We observe two folded structures besides loops that we hypothesize are early intermediates in the toroid formation pathway. At low protamine concentrations (∼0.2 µM), we see that the DNA folds into flowers (structures with multiple loops that are positioned so they look like the petals of a flower). Folding at these concentrations condenses the DNA to 25% of its original length, takes seconds, and is made up of many small bending steps. At higher protamine concentrations (≥2 µM), we observe a second folded structure-the loop stack-where loops are stacked vertically one on top of another. These results lead us to propose a two-step process for folding at this length scale: 1) protamine binds to DNA, bending it into loops and flowers, and 2) flowers collapse into loop stacks. These results highlight how protamine uses a bind-and-bend mechanism to rapidly fold DNA, which may be why protamine can fold the entire sperm genome in minutes.

Beauvericin Immunotoxicity Prevention by Gentiana lutea L. Flower In Vitro.

Beauvericin (BEA) is an emerging mycotoxin produced by some species of Fusarium genera that widely contaminates food and feed. Gentiana lutea is a protected medicinal plant known for its antioxidant and anti-inflammatory properties, which are attributed to its rich content of bioactive compounds. In order to evaluate the beneficial effects of G. lutea flower against BEA cytotoxicity, the aim of this study is to evaluate changes in protein expression after Jurkat cell exposure through a proteomics approach. To carry out the experiment, cells were exposed to intestinally digested G. lutea flower alone or in combination with the BEA standard (100 nM) over 7 days. Differentially expressed proteins were statistically evaluated (p < 0.05), revealing a total of 172 proteins with respect to the control in cells exposed to the BEA standard, 145 proteins for G. lutea alone, and 139 proteins when exposing the cells to the combined exposure. Bioinformatic analysis revealed processes implicated in mitochondria, ATP-related activity, and RNA binding. After careful analysis of differentially expressed proteins, it was evident that G. lutea attenuated, in most cases, the negative effects of BEA. Furthermore, it decreased the presence of major oncoproteins involved in the modulation of immune function.

Genome-wide identification and analysis of the evolution and expression pattern of the SBP gene family in two Chimonanthus species.

BACKGROUND: Chimonanthus praecox and Chimonanthus salicifolius are closely related species that diverged approximately six million years ago. While both C. praecox and C. salicifolius could withstand brief periods of low temperatures of - 15 °C. Their flowering times are different, C. praecox blooms in early spring, whereas C. salicifolius blooms in autumn. The SBP-box (SQUAMOSA promoter-binding protein) is a plant-specific gene family that plays a crucial vital role in regulating plant flowering. Although extensively studied in various plants, the SBP gene family remains uncharacterized in Calycanthaceae. METHODS AND RESULTS: We conducted genome-wide identification of SBP genes in both C. praecox and C. salicifolius and comprehensively characterized the chromosomal localization, gene structure, conserved motifs, and domains of the identified SBP genes. In total, 15 and 18 SBP genes were identified in C. praecox and C. salicifolius, respectively. According to phylogenetic analysis, the SBP genes from Arabidopsis, C. praecox, and C. salicifolius were clustered into eight groups. Analysis of the gene structure and conserved protein motifs showed that SBP proteins of the same subfamily have similar motif structures. The expression patterns of SBP genes were analyzed using transcriptome data. The results revealed that more than half of the genes exhibited lower expression levels in leaves than in flowers, suggesting their potential involvement in the flower development process and may be linked to the winter and autumn flowering of C. praecox and C. salicifolius. CONCLUSION: Thirty-three SBPs were identified in C. praecox and C. salicifolius. The evolutionary characteristics and expression patterns were examined in this study. These results provide valuable information to elucidate the evolutionary relationships of the SBP family and help determine the functional characteristics of the SBP genes in subsequent studies.

Distinct accessory roles of Arabidopsis VEL proteins in Polycomb silencing.

Polycomb repressive complex 2 (PRC2) mediates epigenetic silencing of target genes in animals and plants. In Arabidopsis, PRC2 is required for the cold-induced epigenetic silencing of the FLC floral repressor locus to align flowering with spring. During this process, PRC2 relies on VEL accessory factors, including the constitutively expressed VRN5 and the cold-induced VIN3. The VEL proteins are physically associated with PRC2, but their individual functions remain unclear. Here, we show an intimate association between recombinant VRN5 and multiple components within a reconstituted PRC2, dependent on a compact conformation of VRN5 central domains. Key residues mediating this compact conformation are conserved among VRN5 orthologs across the plant kingdom. In contrast, VIN3 interacts with VAL1, a transcriptional repressor that binds directly to FLC These associations differentially affect their role in H3K27me deposition: Both proteins are required for H3K27me3, but only VRN5 is necessary for H3K27me2. Although originally defined as vernalization regulators, VIN3 and VRN5 coassociate with many targets in the Arabidopsis genome that are modified with H3K27me3. Our work therefore reveals the distinct accessory roles for VEL proteins in conferring cold-induced silencing on FLC, with broad relevance for PRC2 targets generally.

Integrated metabolic profiling and transcriptome analysis of Lonicera japonica flowers for chlorogenic acid, luteolin and endogenous hormone syntheses.

The active ingredients of many medicinal plants are the secondary metabolites associated with the growth period. Lonicera japonica Thunb. is an important traditional Chinese medicine, and the flower development stage is an important factor that influences the quality of medicinal ingredients. In this study, transcriptomics and metabolomics were performed to reveal the regulatory mechanism of secondary metabolites during flowering of L. japonica. The results showed that the content of chlorogenic acid (CGA) and luteolin gradually decreased from green bud stage (Sa) to white flower stage (Sc), especially from white flower bud stage (Sb) to Sc. Most of the genes encoding the crucial rate-limiting enzymes, including PAL, C4H, HCT, C3'H, F3'H and FNSII, were down-regulated in three comparisons. Correlation analysis identified some members of the MYB, AP2/ERF, bHLH and NAC transcription factor families that are closely related to CGA and luteolin biosynthesis. Furthermore, differentially expressed genes (DEGs) involved in hormone biosynthesis, signalling pathways and flowering process were analysed in three flower developmental stage.

The intragenic cis-elements mediate temperature response of RrKSN.

Gene regulation via intragenic sequences is becoming more recognized in many eukaryotes. However, the intragenic sequences mediated gene expressions in response to environmental stimuli have been largely uncharacterized. Here, we showed that the first intron of RrKSN from the Rosa rugosa cultivar 'Purple branch' had a positive effect on RrKSN expression, and the effect depends on its position and orientation. Further analyses revealed that the four adjacent cis-elements (T)CGATT/AATCG(A) within the first intron were critical for the positive regulation, and the RrKSN promotion was significantly suppressed with mutations of these elements. These cis-elements were further evidenced as binding sites for RrARR1, the homologous of Arabidopsis type-B ARABIDOPSIS RESPONSE REGULATOR 1 (ARR1) transcription factor. The first intron-mediated RrKSN expression was enhanced with over-expressing of RrARR1, but abolished with RrARR1 silencing in rose seedlings. Moreover, the expression difference of RrKSN between 16°C and 28°C was eliminated along with RrARR1-silencing. Taken together, these results suggested both RrARR1 and its binding elements are required for the first intron-mediated RrKSN expression in response to varying temperatures. Therefore, our results reveal a unique intragenic regulation mechanism of gene expression by which plants perceive the signal of ambient temperature in rose.

Characterization Variation of the Differential Coloring Substances in Rapeseed Petals with Different Colors Using UPLC-HESI-MS/MS.

Rapeseed's (Brassica napus L.) colorful petals have important ornamental values. However, the mechanisms of regulating petals coloration in rapeseed are still unknown. In our study, we investigated the key differential coloring substances in nine rapeseed cultivars with different petal colors, and 543 metabolites were detected and characterized through UPLC-HESI-MS/MS. Among them, the kinds and contents of flavonols, flavones, and anthocyanidins were the main contributors to petals' coloration. Tamarixetin-, quercetin-, butin-, naringenin- and luteolin-derivates were the main pigment bases in white and yellow petals. Peonidin-3,5-O-diglucoside, peonidin-3-O-(6″-O-caffeoyl)glucoside, and quercetin-derivatives were the main coloring substances in pink petals. Acylated cyanidin derivatives might lead to a series of different purple petal colors. Glycosylated anthocyanins were responsible for the coloration of rapeseed red petals, and peonidin-3-O-glucoside and kaempferol-derivatives were mainly detected from the red petals. These results provide comprehensive insights into the difference in flavonoid metabolites in rapeseed petals with different colors and supply theoretical supports for the breeding of novel colorful rapeseed cultivars.

Cell Biological Analyses of Anther Morphogenesis and Pollen Viability in Arabidopsis and Rice.

Major advances have been made in our understanding of anther developmental processes in flowering plants through a combination of genetic studies, cell biological technologies, biochemical analyses, microarray and high-throughput sequencing-based approaches. In this chapter, we summarize widely used protocols for pollen viability staining, investigation of anther morphogenesis by scanning electron microscopy (SEM), light microscopy of semi-thin sections, ultrathin section-based transmission electron microscopy (TEM), TUNEL (terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate (dUTP) nick end labeling) assay for tapetum programmed cell death, and laser microdissection procedures to obtain specific cells or cell layers for transcriptome analysis.

The GUS Reporter System in Flower Development Studies.

The ß-glucuronidase (GUS) reporter gene system is an important technique with versatile uses in the study of flower development in a broad range of species. Transcriptional and translational GUS fusions are used to characterize gene and protein expression patterns, respectively, during reproductive development. Additionally, GUS reporters can be used to map cis-regulatory elements within promoter sequences and to investigate whether genes are regulated post-transcriptionally. Gene trap/enhancer trap GUS constructs can be used to identify novel genes involved in flower development and marker lines useful in mutant characterization. Flower development studies primarily have used the histochemical assay in which inflorescence tissue from transgenic plants containing GUS reporter genes are stained for GUS activity and examined as whole-mounts or subsequently embedded into wax and examined as tissue sections. In addition, quantitative GUS activity assays can be performed on either floral extracts or intact flowers using a fluorogenic GUS substrate. Another use of GUS reporters is as a screenable marker for plant transformation. A simplified histochemical GUS assay can be used to quickly identify transgenic tissues.