Arbuscular mycorrhizal fungi promote functional gene regulation of phosphorus cycling in rhizosphere microorganisms of Iris tectorum under Cr stress.

The mutualistic symbiotic system formed by clumping arbuscular mycorrhizal fungi (AMF) and plants can remediate heavy metal-contaminated soils. However, the specific mechanisms underlying the interaction between AMF and inter-root microbial communities, particularly their impact on organic phosphorus (P) cycling, remain unclear. This study investigated the gene regulation processes involved in inter-root soil phosphorus cycling in wetland plants, specifically Iris tectorum, following inoculation with AMF under varying concentrations of chromium (Cr) stress. Through macro-genome sequencing, we analyzed the composition and structure of the inter-root soil microbial community associated with Iris tectorum under greenhouse pot conditions. The results demonstrated significant changes in the diversity and composition of the inter-root soil microbial community following AMF inoculation, with Proteobacteria, Actinobacteria, Chloroflexi, Acidobacteria, and Bacteroidetes being the dominant taxa. Under Cr stress, species and gene co-occurrence network analysis revealed that AMF promoted the transformation process of organic phosphorus mineralization and facilitated inorganic phosphorus uptake. Additionally, network analysis of functional genes indicated strong aggregation of (pstS, pstA, pstC, TC.PIT, phoR, pp-gppA) genes, which collectively enhanced phosphorus uptake by plants. These findings shed light on the inter-root soil phosphorus cycling process during the co-remediation of Cr-contaminated soil by AMF-Iris tectorum symbiosis, providing valuable theoretical support for the application of AMF-wetland plant symbiosis systems to remediate heavy metal-contaminated soil.

Phytochemical characterization and biomedical potential of Iris kashmiriana flower extracts: a promising source of natural antioxidants and cytotoxic agents.

Iris kashmiriana belongs to the family Iridaceae and is an important endemic medicinal plant of Kashmir. The current study was designed to determine the phytoconstituents, antioxidant, and cytotoxic potential of ethyl acetate (IRK-ETH) and methanol (IRK-MTH) extracts of Iris kashmiriana flowers. IRK-MTH extract demonstrated maximum radical scavenging activity in DPPH, ABTS, and Superoxide anion radical antioxidant assays with IC50 values of 73.15 µg/ml, 79.05 µg/ml, and 86.52 µg/ml respectively. IRK-ETH and IRK-MTH extracts possessed phenolic (70.9 and 208.5 mgGAE/gdw) and flavonoid (487.7 and 40.55 mgRE/gdw) contents respectively. In MTT assay IRK-ETH demonstrated the highest cytotoxicity towards the MCF-7 cell line with a GI50 value of 49.13 µg/ml. Phase contrast and fluorescence microscopic studies in MCF-7 cells revealed that IRK-ETH extract caused condensation of chromatin, rounding of cells, and nuclear condensation in cells which shows the apoptotic potential of the extract. GCMS analysis for phytochemical characterization revealed the presence of 9 compounds in both extracts which have been reported to possess antibacterial, cytotoxic, and anti-oxidant activities. HPLC analysis confirmed the presence of different polyphenols in both extracts with IRK-MTH extract having maximum polyphenols like epicatechin, rutin, quercetin, vanillic acid, sinapic acid, caffeic acid, chlorogenic acid and ellagic acid. These findings suggest that the flowers of Iris kashmiriana possess very good antioxidant and cytotoxic potential owing to its rich phytoconstituents.

Gas Chromatography-Mass Spectrometry Analysis of Volatile Organic Compounds from Three Endemic Iris Taxa: Headspace Solid-Phase Microextraction vs. Hydrodistillation.

Iris taxa are sources of valuable essential oils obtained from aged rhizomes used by various industries, including pharmacy, cosmetic, perfume, and food industry, in which irones are the most important aroma components. In this study, volatile organic compounds (VOCs) obtained from dried rhizomes of three endemics from Croatia, Iris pseudopallida, I. illyrica, and I. adriatica, were studied. The VOCs were isolated by three different methods: headspace solid-phase microextraction (HS-SPME) using divinylbenzene/carboxene/polydimethylsiloxane (DVB/CAR/PDMS) fiber or polydimethylsiloxane/divinylbenzene (PDMS/DVB) fiber, and hydrodistillation (HD). The samples were analyzed by gas chromatography-mass spectrometry (GC-MS). In five out of six samples, the main compounds detected by HS-SPME were perilla aldehyde, butan-2,3-diol, acetic acid, 2-phenylethanol, benzyl alcohol, hexanal, and nonanal, while 6-methylhept-5-en-2-one, trans-caryophyllene, and ethanol were common for all studied samples. The former VOCs were absent from the oldest, irone-rich I. pseudopallida sample, mainly characterized by cis-α-irone (43.74-45.76%). When using HD, its content was reduced (24.70%), while docosane prevailed (45.79%). HD yielded predominantly fatty acids, including myristic, common for all studied taxa (4.20-97.01%), and linoleic (40.69%) and palmitic (35.48%) as the major VOCs of I. adriatica EO. The performed GC-MS analyses of EOs, in combination with HS-SPME/GC-MS, proved to be useful for gaining a better insight into Iris VOCs.

The effects of polypropylene microplastics on the removal of nitrogen and phosphorus from water by Acorus calamus, Iris tectorum and functional microorganisms.

Polypropylene microplastics (PP-MPs), an emerging pollutant, adversely affect the ability of aquatic plants to restore water bodies, thereby compromising the functionality and integrity of wetland ecosystems. This study examines the effects of microplastic stress on the nitrogen and phosphorus removal capacities of Acorus calamus and Iris tectorum, as well as on functional microorganisms within the aquatic system. The findings indicate that under PP-MP stress, the nitrogen and phosphorus absorption capabilities of both plants were diminished. Additionally, there was a significant reduction in the metabolic enzyme activities related to nitrogen and phosphorus in the plants, alongside a notable decrease in leaf nitrogen content. PP-MPs hinder the nutrient uptake of plants, affecting their growth and indirectly reducing their ability to utilize nitrogen and phosphorus. Specifically, in the 10 mg L-1 treatment group, A. calamus and I. tectorum showed reductions in leaf nitrogen content by 23.1% and 31.0%, respectively, and by 14.8% and 27.7% in the 200 mg L-1 treatment group. Furthermore, I. tectorum had higher leaf nitrogen levels than A. calamus. Using fluorescent tagging, the distribution of PP-MPs was traced in the roots, stems, and leaves of the plants, revealing significant growth impairment in both species. This included a considerable decline in photosynthetic pigment synthesis, enhanced oxidative stress responses, and increased lipid peroxidation in cell membranes. PP-MP exposure also significantly reduced the abundance of functional microorganisms involved in denitrification and phosphorus removal at the genus level in aquatic systems. Ecological function predictions revealed a notable decrease in nitrogen cycling functions such as nitrogen respiration and nitrite denitrification among water microorganisms in both treatment groups, with a higher ecological risk potential in the A. calamus treatment group. This study provides new insights into the potential stress mechanisms of PP-MPs on aquatic plants involved in water body remediation and their impacts on wetland ecosystems.

Spatiotemporal dynamics of reactive oxygen species and its effect on beta-blockers' degradation in aquatic plants' rhizosphere.

The pathway for pollutant degradation involving reactive oxygen species (ROS) in the rhizosphere is poorly understood. Herein, a rootchip system was developed to pinpoint the ROS hotspot along the root tip of Iris tectorum. Through mass balance analysis and quenching experiment, we revealed that ROS contributed significantly to rhizodegradation for beta-blockers, ranging from 22.18 % for betaxolol to 83.83 % for atenolol. The identification of degradation products implicated ROS as an important agent to degrade atenolol into less toxic transformation products during phytoremediation. Moreover, an active production of ROS in rhizosphere was identified by mesocosm experiment. Across three root-associated regions aquatic plants inhabiting the rhizosphere accumulated the highest •OH of ∼1200 nM after 3 consecutive days, followed by rhizoplane (∼230 nM) and bulk environment (∼60 nM). ROS production patterns were driven by rhizosphere chemistry (Fe and humic substances) and microbiome variations in different rhizocompartments. These findings not only deepen understanding of ROS production in aquatic plants rhizosphere but also shed light on advancing phytoremediation strategies.

Five new flavonoids and their pharmacological activities from Iris tenuifolia Pall.

Five new components including two new isoflavones, 5, 7, 2', 3'-tetrahydroxy-6-methoxyisoflavone (1), 5, 7, 2', 3'-tetrahydroxy-8-methoxyisoflavone (2), one flavonol 3, 5, 3', 4'-tetrahydroxy-7, 2'-dimethoxyflavonol (3), one flavanone (2S)-5, 7, 3'-trihydroxy-2'-methoxyflavanone (4), and one flavanonol (2R, 3R)-3, 5, 3', 4'-tetrahydroxy-7, 2'-dimethoxyflavanonol (5), along with nine known flavonoids (6-14) were isolated from under ground parts of Iris tenuifolia Pall. Their structures were elucidated by NMR and HRESIMS data and by comparison of CD spectra with compounds having similar structure. The separated compounds were evaluated for in vitro antioxidant activities by DPPH and ABTS. The α-glucosidase inhibitory activity of the compounds were evaluated with the pNPG method, the results indicated flavonoids were potential inhibitors of α-glucosidase. Moreover, in vitro anti-oxidative assay using flow cytometry indicated that compounds 1-5 showed strong oxidation resistance ability on C8D1A cells without affecting the cell viability.

Effects of cadmium and zinc interactions on the physiological biochemistry and enrichment characteristics of Iris pseudacorus.

Compound pollution with cadmium (Cd) and zinc (Zn) is common in nature. The effects of compounded Cd and Zn on the growth and development of Iris pseudacorus in the environment and the plant's potential to remediate heavy metals in the environment remain unclear. In this study, the effects of single and combined Cd and Zn stress on I. pseudacorus growth and the enrichment of heavy metals in I. pseudacorus seedlings were investigated. The results showed that under Cd (160 µM) and Zn (800 µM) stress, plant growth was significantly inhibited and photosynthetic performance was affected. Cd+Zn200 (160 µM + 200 µM) reduced the levels of malondialdehyde, hydrogen peroxide, and non-protein thiols by 31.29%, 53.20%, and 13.29%, respectively, in the aboveground tissues compared with levels in the single Cd treatment. However, Cd+Zn800 (160 µM + 800 µM) had no effect. Cd and Zn800 inhibited the absorption of mineral elements, while Zn200 had little effect on plants. Compared with that for Cd treatment alone, Cd + Zn200 and Cd+Zn800 reduced the Cd content in aboveground tissues by 54.15% and 49.92%, respectively, but had no significant effect on Cd in the root system. Zn significantly reduced the Cd content in subcellular components and limited the content and proportion of Cd extracted using water and ethanol. These results suggest that a low supply of Zn reduces Cd accumulation in aboveground tissues by promoting antioxidant substances and heavy metal chelating agents, thus protecting the photosynthetic systems. The addition of Zn also reduced the mobility and bioavailability of Cd to alleviate its toxicity in I. pseudacorus.

Complete plastid genome of Iris orchioides and comparative analysis with 19 Iris plastomes.

Iris is a cosmopolitan genus comprising approximately 280 species distributed throughout the Northern Hemisphere. Although Iris is the most diverse group in the Iridaceae, the number of taxa is debatable owing to various taxonomic issues. Plastid genomes have been widely used for phylogenetic research in plants; however, only limited number of plastid DNA markers are available for phylogenetic study of the Iris. To understand the genomic features of plastids within the genus, including its structural and genetic variation, we newly sequenced and analyzed the complete plastid genome of I. orchioides and compared it with those of 19 other Iris taxa. Potential plastid markers for phylogenetic research were identified by computing the sequence divergence and phylogenetic informativeness. We then tested the utility of the markers with the phylogenies inferred from the markers and whole-plastome data. The average size of the plastid genome was 152,926 bp, and the overall genomic content and organization were nearly identical among the 20 Iris taxa, except for minor variations in the inverted repeats. We identified 10 highly informative regions (matK, ndhF, rpoC2, ycf1, ycf2, rps15-ycf, rpoB-trnC, petA-psbJ, ndhG-ndhI and psbK-trnQ) and inferred a phylogeny from each region individually, as well as from their concatenated data. Remarkably, the phylogeny reconstructed from the concatenated data comprising three selected regions (rpoC2, ycf1 and ycf2) exhibited the highest congruence with the phylogeny derived from the entire plastome dataset. The result suggests that this subset of data could serve as a viable alternative to the complete plastome data, especially for molecular diagnoses among closely related Iris taxa, and at a lower cost.

Biological effects and phytochemical study of the underground part of Iris scariosa Willd. ex Link extract: A new source of bioactive constituents.

The expected toxicity and resistance of chemotherapeutic agents necessitate and encourage for the use of natural chemotherapeutic sources of plant origin in the clinical stage of cancer therapy. Plants of the genus Iris (Iridaceae) used by local populations for the treatment of cancer, bacterial and viral infections. In this study, an ethanol extract of rhizomes of I. scariosa was prepared and tested for the cytotoxicity using the MTT assay. The extract exhibited the most potent cytotoxicity against the breast cancer cell line MCF7 (IC50 = 9.28 ± 0.49 µg/ml, selectively index ˃5), and induced apoptosis in MCF7 lines. Notably, the extract significantly inhibited the colony formation of MCF7 and HepG2 cancer cells at a concentration range from 10.6 to 85.0 µg/ml, including non-toxic concentrations for HepG2 cells. The ethanol extract was analyzed by HPLC, revealed the identification of 5 secondary metabolites (quercetin, rutin, myricetin, apigenin, artemisetin), the content of which was shown to reach around 15% of the extract. The petroleum ether (PE) part of the extract (yield 2.62%) was analyzed by GC-MS. The composition of tert-butyl methyl ether (TBME) part of the extract (yield 23.72%) was studied. Total of 15 individual compounds: two benzophenones, eight isoflavones, four flavones and a (2R)-flavanone were isolated. The pentamethoxyflavone artemisetin and flavanone pinocembrin were isolated for the first from Iris sp. The readily available isoflavones from the TBME part of extract (irilone, iriflogenin, irigenin and tectorigenin) may serve as new leads for the discovery of anticancer drugs.

Metagenomic analysis revealed N-metabolizing microbial response of Iris tectorum to Cr stress after colonization by arbuscular mycorrhizal fungi.

Arbuscular mycorrhizal fungi (AMF) and plant growth-promoting bacteria enhance plant tolerance to abiotic stress and promote plant growth in contaminated soil. However, the interaction mechanism between rhizosphere microbial communities under chromium (Cr) stress remains unclear. This study conducted a greenhouse pot experiment and metagenomics analysis to reveal the comprehensive effects of the interaction between AMF (Rhizophagus intraradices) and nitrogen-N metabolizing plant growth promoters on the growth of Iris tectorum. The results showed that AMF significantly increased the biomass and nutrient levels of I. tectorum in contaminated soil and decreased the content of Cr in the soil. Metagenomics analysis revealed that the structure and composition of the rhizosphere microbial community involved in nitrogen metabolism changed significantly after inoculation with AMF under Cr stress. Functional genes related to soil nitrogen mineralization (gltB, gltD, gdhA, ureC, and glnA), nitrate reduction to ammonium (nirB, nrfA, and nasA), and soil nitrogen assimilation (NRT, nrtA, and nrtC) were up-regulated in the N-metabolizing microbial community. In contrast, the abundance of functional genes involved in denitrification (nirK and narI) was down-regulated. In addition, the inoculation of AMF regulates the synergies between the N-metabolic rhizosphere microbial communities and enhances the complexity and stability of the rhizosphere ecological network. This study provides a basis for improving plant tolerance to heavy metal stress by regulating the functional abundance of N-metabolizing plant growth-promoting bacteria through AMF inoculation. It helps to understand the potential mechanism of wetland plant remediation of Cr-contaminated soil.

Mechanism of Iris sibirica and aeration combination on promoting the water purification performance of constructed wetland under low temperature.

Temperature is an important factor affecting the water purification performance of constructed wetland (CW). In the previous study, the combined measures of Iris sibirica and aeration at the bottom of the first quarter filtration chamber could improve the pollutant removal capacity of CW at low temperature. However, the mechanism between the combined measures of Iris sibirica and aeration on enhancing the performance of domestic sewage treatment is unclear. Our study aims to provide scientific validation for the combined measure through monitoring the concentrations of dissolved oxygen (DO), chemical oxygen demand (CODCr), ammonia nitrogen (NH4+-N), and total nitrogen (TN) along the water flow pathway of the CW and measuring the superoxide dismutase (SOD) activities of the plants and the abundance of nitrogen cycle-related microbial functional genes in the substrates of CW to explore the mechanism of combined measures promoting the removal efficiency of the CW under low-temperature stress. Results showed that aerating at the bottom of the first quarter filtration chamber increased DO concentration in the front part of the CW, which benefited the aerobic removal of pollutants and the activities of microorganisms, and the removal CODCr and NH4+-N occurred mainly in the front part of the CW. SOD activities showed that I. sibirica had better resistance to low temperature than Canna indica did. The combined measures of I. sibirica and aeration activated the activities of microorganisms, increased the abundance of the denitrification process genes along the water flow pathway and formed a clear nitrification-denitrification zone in the CW, thus promoted the nitrogen removal efficiency at low temperature. Therefore, this study confirmed the feasibility of the combined measures from a mechanistic perspective.

Transcriptome-Wide Identification and Expression Analysis of bHLH Family Genes in Iris domestica under Drought and Cu Stress.

The role of bHLH transcription factors in plant response to abiotic stress and regulation of flavonoid metabolism is well documented. However, to date, the bHLH transcription factor family in Iris domestica remains unreported, impeding further research on flavonoid metabolism in this plant. To address this knowledge gap, we employed bioinformatics to identify 39 IdbHLH genes and characterised their phylogenetic relationships and gene expression patterns under both drought and copper stress conditions. Our evolutionary tree analysis classified the 39 IdbHLHs into 17 subfamilies. Expression pattern analysis revealed that different IdbHLH transcription factors had distinct expression trends in various organs, suggesting that they might be involved in diverse biological processes. We found that IdbHLH36 was highly expressed in all organs (Transcripts Per Million (TPM) > 10), while only 12 IdbHLH genes in the rhizome and four in the root were significantly upregulated under drought stress. Of these, four genes (IdbHLH05, -37, -38, -39) were co-upregulated in both the rhizome and root, indicating their potential role in drought resistance. With regards to copper stress, we found that only 12 genes were upregulated. Further co-expression analysis revealed that most bHLH genes were significantly correlated with key enzyme genes involved in isoflavone biosynthesis. Thereinto, IdbHLH06 showed a significant positive correlation with IdC4H1 and Id4CL1 (p < 0.05). Furthermore, a transient expression assay confirmed that the IdbHLH06 protein was localised in the nucleus. Our findings provide new insights into the molecular basis and regulatory mechanisms of bHLH transcription factors in isoflavone biosynthesis in I. domestica.

Selective extraction of plant bioactive compounds with deep eutectic solvents: Iris sibirica L. as example.

INTRODUCTION: Deep eutectic solvents (DESs) are promising extractants with tuneable properties. However, there is a lack of reports about the influence of the nature of the original DES on obtaining the metabolomic profile of a plant. OBJECTIVE: The aim of this study is to investigate the possibility of obtaining Iris sibirica L. chromatographical profiles with DESs based on various hydrogen bond donors and acceptors as extraction solvents. METHODOLOGY: DESs were prepared by mixing choline chloride or tetrabutylammonium bromide with various hydrogen bond donors and investigated for the extraction of bioactive substances from biotechnological raw materials of I. sibirica L. The obtained extracts were analysed by HPLC with diode array detector (DAD) and Q-MS. RESULTS: Chromatographic profiles for I. sibirica L. extracts by eight choline chloride DESs and six tetrabutylammonium DESs have been obtained. It has been found that selective recovery of bioactive substances can be achieved by varying the composition of DESs. Eleven phenolic compounds were identified in I. sibirica L. using HPLC-MS. Phase separation was observed with acetonitrile for four DESs. New flavonoid derivatives have been found in DES extracts compared with methanol extracts. CONCLUSION: The results showed the possibility of DES usage for extraction without water addition. Selectivity of DESs varies depending on the chemical composition of hydrogen bond donors and acceptors. Choline chloride is a more suitable hydrogen bond acceptor for the flavonoid extraction. Choline chloride-lactic acid (1:1) DES has demonstrated a metabolic profile that was the closest to the methanol one and enhanced the extraction up to 2.6-fold.

Flower color polymorphism of a wild Iris on the Qinghai-Tibet plateau.

BACKGROUND: Flower color plays a crucial role in attracting pollinators and facilitating environmental adaptation. Investigating the causes of flower color polymorphism and understanding their potential effects on both ecology and genetics can enhance our understanding of flower color polymorphism in wild plant. RESULTS: In this study, we examined the differences of potential male and female fitness between purple- and yellow- flower individuals in Iris potaninii on the Qinghai-Tibet Plateau, and screened key genes and positively selective genes involved in flower color change. Our results showed that yellow flower exhibited a higher pollen-to-ovule ratio. Yellow flowers were derived from purple flowers due to the loss of anthocyanins, and F3H could be an essential gene affecting flower color variation though expression regulation and sequence polymorphism in this species. Furthermore, our findings suggest that genes positively selected in yellow-flowered I. potaninii might be involved in nucleotide excision repair and plant-pathogen interactions. CONCLUSIONS: These results suggest that F3H induces the flower color variation of Iris potaninii, and the subsequent ecological and additive positive selection on yellow flowers may further enhance plant adaptations to alpine environments.

Correlation between secondary metabolites of Iris confusa Sealy and Iris pseudacorus L. and their newly explored antiprotozoal potentials.

BACKGROUND: In the last few decades, the use of plant extracts and their phytochemicals as candidates for the management of parasitic diseases has increased tremendously. Irises are aromatic and medicinal plants that have long been employed in the treatment of different infectious diseases by traditional healers in many cultures. This study aims to explore the potential of three common Iris species (I. confusa Sealy, I. pseudacorus L. and I. germanica L.) against infectious diseases. Their in vitro antiprotozoal potency against Plasmodium falciparum, Trypanosoma brucei brucei, T. b. rhodesiense, T. cruzi and Leishmania infantum beside their cytotoxicity on MRC-5 fibroblasts and primary peritoneal murine macrophages were examined. METHODS: The secondary metabolites of the tested extracts were characterized by UPLC-HRMS/MS and Pearsons correlation was used to correlate them with the antiprotozoal activity. RESULTS: Overall, the non-polar fractions (NPF) showed a significant antiprotozoal activity (score: sc 2 to 5) in contrast to the polar fractions (PF). I. confusa NPF was the most active extract against P. falciparum [IC50 of 1.08 µg/mL, selectivity index (S.I. 26.11) and sc 5] and L. infantum (IC50 of 12.7 µg/mL, S.I. 2.22 and sc 2). I. pseudacorus NPF was the most potent fraction against T. b. rhodesiense (IC50 of 8.17 µg/mL, S.I. 3.67 and sc 3). Monogalactosyldiacylglycerol glycolipid (18:3/18:3), triaceylglycerol (18:2/18:2/18:3), oleic acid, and triterpenoid irridals (spirioiridoconfal C and iso-iridobelamal A) were the top positively correlated metabolites with antiplasmodium and antileishmanial activities of I. confusa NPF. Tumulosic acid, ceramide sphingolipids, corosolic, maslinic, moreollic acids, pheophytin a, triaceylglycerols, mono- and digalactosyldiacylglycerols, phosphatidylglycerol (22:6/18:3), phosphatidylcholines (18:1/18:2), and triterpenoid irridal iso-iridobelamal A, were highly correlated to I. pseudacorus NPF anti- T. b. rhodesiense activity. The ADME study revealed proper drug likeness properties for certain highly corelated secondary metabolites. CONCLUSION: This study is the sole map correlating I. confusa and I. pseudacorus secondary metabolites to their newly explored antiprotozoal activity.

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.

WRKY Transcriptional Factor IlWRKY70 from Iris laevigata Enhances Drought and Salinity Tolerances in Nicotiana tabacum.

Drought and high salinity greatly affect plant growth and development. WRKY transcription factors play a key role in plant tolerance to abiotic stress, but the functions of WRKYs in the ornamental monocotyledon Iris laevigata remain largely unexplored. In this study, we cloned IlWRKY70 and found that it is a Group III WRKY localized in the nucleus. The expression of IlWRKY70 was induced by NaCl and PEG-6000, which reached peaks (4.38 and 5.65 times) after 3 h and 1 h, respectively. The exogenous overexpression of IlWRKY70 in N. tabacum significantly improved the resistance under NaCl and drought treatments, as evidenced by higher germination rates, longer root lengths, and increased fresh weights compared to those of control plants. In addition, transgenic seedlings showed significantly reduced wilting, higher photosynthetic performance, higher Fv/Fm and chlorophyll content, and lower stomatal conductance. Moreover, transgenic lines showed higher antioxidant enzymatic activities, lower reactive oxygen species (ROS), and lower malondialdehyde contents. Accordingly, we also found higher expressions of antioxidant defense genes, including SOD, CAT, and POD, in transgenic lines compared to controls under salt and drought stresses. Thus, IlWRKY70 enhances the abilities of salt and drought tolerances in plants, at least partially, via ROS regulation and can be used for breeding I. laevigata possessing enhanced salt and drought resistances.

Identification of characteristic aroma compounds for spicy in Iris lactea var. chinensis.

Iris lactea var. chinensis (Fisch.) Koidz has a unique floral fragrance that differs from that of other Iris spp.; however, its characteristic aroma composition remains unknown. This study aimed to identify the floral fragrance components of I. lactea var. chinensis during different flowering stages using headspace solid-phase microextraction in conjunction with gas chromatography mass spectrometry, electronic nose, and sensory evaluation. During the three flowering phases (bud stage, bloom stage, and decay stage), 70 volatile organic compounds (VOCs), including 13 aldehydes, 13 esters, 11 alcohols, 10 alkanes, 8 ketones, 7 terpenes, 7 benzenoids, and 1 nitrogenous compound, were identified. According to principal component analysis, the primary VOCs were (-)-pinene, ß-irone, methyl heptenone, phenylethanol, hexanol, and 2-pinene. A comparison of the differential VOCs across the different flowering stages using orthogonal partial least squares discriminant analysis and hierarchical clustering analysis revealed that 3-carene appeared only in the bud stage, whereas hexanol, ethyl caprate, ethyl caproate, linalool, (-)-pinene, and 2-pinene appeared or were present at significantly increased levels during the bloom stage. The phenylethanol, methyl heptenone, 3-methylheptane, and ß-irone reached a peak in the decay stage. The odor activity value and sensory evaluation suggested that "spicy" is the most typical odor of I. lactea var. chinensis, mainly due to 2-methoxy-3-sec-butylpyrazine, which is rare in floral fragrances.

Vitisin A-13-O-ß-D-glucoside and Vitisin A from Iris lactea inhibit lipogenesis and promote lipolysis via the PKA/HSL pathway during adipogenic transdifferentiation of C2C12 cells.

Oligostilbenes are a group of natural products derived from the polymerization of stilbene monomers. Despite the demonstration of their activities in regulating lipid metabolism, the function of oligostilbenes in the adipogenic transdifferentiation of multipotent myoblast cells remains unknown. Hence, the five oligostilbenes from Iris lactea were tested for their regulatory effects on adipogenic transdifferentiation of C2C12 myoblast cells. As a result, it was shown that Vitisin A-13-O-ß-D-glucoside (VitAOG), Vitisin A (VitA) and Hopeaphenol (Hop) can greatly inhibit the adipogenic transdifferentiation of C2C12 cells by reducing lipid accumulation and downregulating the expression of peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer binding protein beta (C/EBPß) and fatty acid binding proteins 4 (FABP4). In contrast, Vitisin D (VitD) and Isohopeaphenol (Isohop) promote adipogenic transdifferentiation of C2C12 cells by increasing lipid accumulation and upregulating the expression of adipogenesis and lipogenesis markers. Further research found that the lipolytic protein levels of adipocyte triglyceride lipase (ATGL) and phosphorylation of hormone-sensitive lipase (HSL) were elevated by VitAOG and VitA. Additionally, VitAOG and VitA maintain lipid homeostasis by improving mitochondrial function. Taken together, our study reveals an effect of oligostilbenes on lipid metabolism in C2C12 cells, and VitAOG and VitA can be regarded as potential candidates for the treatment of obesity and other disorders of lipid metabolism.

IlNRAMP5 is required for cadmium accumulation and the growth in Iris lactea under cadmium exposures.

Iris lactea is potentially applied for remediating Cd-contaminated soils due to the strong ability of Cd uptake and accumulation. However, its molecular mechanism underlying Cd uptake pathway remains unknown. Here, we report a member of NRAMP (Natural Resistance-Associated Macrophage Protein) family, IlNRAMP5, is involved in Cd/Mn uptake and the growth in I. lactea response to Cd. IlNRAMP5 was localized onto the plasma membrane, and was induced by Cd. It was expressed in the root cortex rather than the central vasculature, and in leaf vascular bundle and mesophyll cells. Heterologous expression in yeast showed that IlNRAMP5 could transport Cd and Mn, but not Fe. Knockdown of IlNRAMP5 triggered a significant reduction in Cd uptake, further diminishing the accumulation of Cd. In addition, silencing IlNRAMP5 disrupted Mn homeostasis by lowering Mn uptake and Mn allocation, accompanied by remarkably inhibiting photosynthesis under Cd conditions. Overall, the findings suggest that IlNRAMP5 plays versatile roles in Cd accumulation by mediating Cd uptake, and contributes to maintain the growth via modulating Mn homeostasis in I. lactea under Cd exposures. This would provide a mechanistic understanding Cd phytoremediation efficiency in planta.