Nelumbo nucifera leaves extract ameliorated scopolamine-induced cognition impairment via enhanced adult hippocampus neurogenesis.

In this presentation, we explored the molecular mechanisms of N. nucifera leaf water extracts (NLWEs) and polyphenol extract (NLPE) on scopolamine-induced cell apoptosis and cognition defects. The administration of NLWE and NLPE did not alter the body weight and serum biomarker rs and significantly ameliorated scopolamine-induced cognition impairment according to Y-maze test analysis. In mice, treatment with scopolamine disrupted normal histoarchitecture in the hippocampus, whereas the administration of NLWE and NLPE reversed the phenomenon. Western blot analysis revealed that scopolamine mitigated the expression of doublecortin (DCX), nestin, and NeuN, and cotreatment with NLWE or NLPE significantly recovered the expression of these proteins. NLWE and NLPE upregulated DCX and NeuN expression in the hippocampus region, as evidenced by immunohistochemical staining analysis of scopolamine-treated mice. NLWE and NLPE obviously elevated brain-derived neurotrophic factor (BDNF) and enhanced its downstream proteins activity. NLWE and NLPE attenuated scopolamine-induced apoptosis by reducing Bax and increased Bcl-2 expression. In addition, scopolamine also triggered apoptosis in human neuroblastoma SH-SY5Y cells whereas co-treatment with NLWE or quercetin-3-glucuronide (Q3G) reversed the phenomenon. NLWE or Q3G enhanced Bcl-2 and reduced Bax expression in the presence of scopolamine in SH-SY5Y cells. NLWE or Q3G recovered the inhibitory effects of scopolamine on neurogenesis and BDNF signals in SH-SY5Y cells. Overall, our results revealed that N. nucifera leaf extracts and Q3G promoted adult hippocampus neurogenesis and prevented apoptosis to mitigate scopolamine-induced cognition dysfunction through the regulation of BDNF signaling pathway.

Metabolite profiling and screening of callus browning-related genes in lotus (Nelumbo nucifera).

Callus browning is a major drawback to lotus callus proliferation and regeneration. However, the underlying mechanism of its formation remains largely unknown. Herein, we aimed to explore the metabolic and molecular basis of lotus callus browning by combining histological staining, high-throughput metabolomics, and transcriptomic assays for lotus callus at three browning stages. Histological stained brown callus cross sections displayed severe cell death symptoms, accompanied by an obvious accumulation of polyphenols and lignified materials. Widely targeted metabolomics revealed extensively decreased accumulation of most detected flavonoids and benzylisoquinoline alkaloids (BIAs), as well as a few phenolic acids, amino acids and their derivatives in callus with browning symptoms. Conversely, the contents of most detected tannins were significantly increased. Subsequent comparative transcriptomics identified a set of differentially expressed genes (DEGs) associated with the biosynthesis and regulation of flavonoids and BIAs in lotus. Notably, callus browning was coupled with significantly up-regulated expression of two polyphenol oxidase (PPO) and 17 peroxidase (POD) encoding genes, while the expression of ethylene associated genes remained at marginal levels. These results suggest that lotus callus browning is primarily controlled at the level of metabolism, wherein the oxidation of flavonoids and BIAs is crucially decisive.

Overexpression of geranyl diphosphate synthase 1 (NnGGPPS1) from Nelumbo nucifera enhances carotenoid and chlorophyll content and biomass.

As the traditional herb with pharmacological compounds in China, the key genes related with terpenoid biosynthesis are still unveiled in Nelumbo nucifera. Geranylgeranyl pyrophosphate synthase (GGPPS) is one of the key enzymes in terpenoids biosynthesis, synthesizing the common precursor of GGPP for downstream enzymes for generating various terpenoids. In this study, four NnGGPPS genes were isolated from N. nucifera. Sequence and phylogenetic analyses indicate that NnGGPPS1 and NnGGPPS2 belong to large subunit (LSU). Whereas NnGGPPS3 and NnGGPPS4 are classified as small subunit (SSU) of SSU Ⅱ and SSU I, respectively. Among four NnGGPPSs, only NnGGPPS1 and NnGGPPS2 can produce GGPP in bacterial pigment complementation assay. Combination analysis of subcellular localization and gene co-expression analysis (GCN) illustrates that NnGGPPS1 is the main transcript related with methylerythritol phosphate (MEP) pathway, abscisic acid (ABA) biosynthesis, carotenoid and chlorophyll biosynthesis and degradation. Overexpression of NnGGPPS1 improves the growth of transgenic tobacco, and increases carotenoids and chlorophyll contents. Moreover, NnGGPPS1 transgenic tobacco exhibits improved photosynthesis efficiency and ROS scavenging ability. The up-regulated expression of the key genes in MEP pathway, carotenoid biosynthesis and chlorophyll biosynthesis, result in the increase of metabolic flux in NnGGPPS1 transgenic lines. Furthermore, the elevated MEP-derived primary metabolites of carotenoid and chlorophyll was attributed to enhancement of plant biomass of NnGGPPS1 transgenic lines. Therefore, NnGGPPS1 plays a vital role in biosynthesis of carotenoid and chlorophyll.

Transcription factor NnMYB5 controls petal color by regulating GLUTATHIONE S-TRANSFERASE2 in Nelumbo nucifera.

Lotus (Nelumbo spp.) is an important aquatic ornamental genus in the family Nelumbonaceae comprising only 2 species: Nelumbo lutea with yellow flowers and Nelumbo nucifera with red or white flowers. The petal color variations between these 2 species have previously been associated with the potential activities of FLAVONOL SYNTHASE (FLS) and MYB5. However, the underlying genetic mechanisms of flower color divergence within the N. nucifera species remain unclear. Here, quantitative trait locus mapping led to the identification of MYB5, a candidate gene controlling petal color in N. nucifera. Genotyping of 213 natural lotus accessions revealed an 80 kb presence/absence variant (PAV) of the NnMYB5 gene that is associated with petal color variation. Transcriptome analysis, dual-luciferase, and yeast 1-hybrid assays showed that NnMYB5 could directly activate the anthocyanin transporter gene GLUTATHIONE S-TRANSFERASE2 (NnGST2). Heterologous expression of NnGST2 in Arabidopsis (Arabidopsis thaliana) and its overexpression in lotus petals induced anthocyanin accumulation. Deletion of the 80 kb PAV within NnMYB5 inactivated NnGST2 expression and blocked anthocyanin accumulation in white N. nucifera petals. In contrast, the anthocyanin deficiency of N. lutea occurred due to pseudogenized NlMYB5 alleles. Our results establish a regulatory link between NnMYB5 and NnGST2 in petal anthocyanin accumulation and demonstrate the independent mechanisms controlling flower coloration in Nelumbo.

Substrate specificity of polyphenol oxidase and its selectivity towards polyphenols: Unlocking the browning mechanism of fresh lotus root (Nelumbo nucifera Gaertn.).

Polyphenol oxidase (PPO) causes the browning of lotus roots (LR), negatively affecting their nutrition and shelf-life. This study aimed to explore the specific selectivity of PPO toward polyphenol substrates, thus unlocking the browning mechanism of fresh LR. Results showed that two highly homologous PPOs were identified in LR and exhibited the highest catalytic activity at 35 ℃ and pH 6.5. Furthermore, the substrate specificity study revealed (-)-epigallocatechin had the lowest Km among the polyphenols identified in LR, while (+)-catechin showed the highest Vmax. The molecular docking further clarified that (-)-epigallocatechin exhibited lower docking energy and formed more hydrogen bonds and Pi-Alkyl interactions with LR PPO than (+)-catechin, while (+)-catechin entered the active cavity of PPO more quickly due to its smaller structure, both of which enhance their affinity to PPO. Thus, (+)-catechin and (-)-epigallocatechin are the most specific substrates responsible for the browning mechanism of fresh LR.

Pathway elucidation and microbial synthesis of proaporphine and bis-benzylisoquinoline alkaloids from sacred lotus (Nelumbo nucifera).

Sacred lotus (Nelumbo nucifera) has been utilized as a food, medicine, and spiritual symbol for nearly 3000 years. The medicinal properties of lotus are largely attributed to its unique profile of benzylisoquinoline alkaloids (BIAs), which includes potential anti-cancer, anti-malarial and anti-arrhythmic compounds. BIA biosynthesis in sacred lotus differs markedly from that of opium poppy and other members of the Ranunculales, most notably in an abundance of BIAs possessing the (R)-stereochemical configuration and the absence of reticuline, a major branchpoint intermediate in most BIA producers. Owing to these unique metabolic features and the pharmacological potential of lotus, we set out to elucidate the BIA biosynthesis network in N. nucifera. Here we show that lotus CYP80G (NnCYP80G) and a superior ortholog from Peruvian nutmeg (Laurelia sempervirens; LsCYP80G) stereospecifically convert (R)-N-methylcoclaurine to the proaporphine alkaloid glaziovine, which is subsequently methylated to pronuciferine, the presumed precursor to nuciferine. While sacred lotus employs a dedicated (R)-route to aporphine alkaloids from (R)-norcoclaurine, we implemented an artificial stereochemical inversion approach to flip the stereochemistry of the core BIA pathway. Exploiting the unique substrate specificity of dehydroreticuline synthase from common poppy (Papaver rhoeas) and pairing it with dehydroreticuline reductase enabled de novo synthesis of (R)-N-methylcoclaurine from (S)-norcoclaurine and its subsequent conversion to pronuciferine. We leveraged our stereochemical inversion approach to also elucidate the role of NnCYP80A in sacred lotus metabolism, which we show catalyzes the stereospecific formation of the bis-BIA nelumboferine. Screening our collection of 66 plant O-methyltransferases enabled conversion of nelumboferine to liensinine, a potential anti-cancer bis-BIA from sacred lotus. Our work highlights the unique benzylisoquinoline metabolism of N. nucifera and enables the targeted overproduction of potential lotus pharmaceuticals using engineered microbial systems.

Proteomic and metabolomic analyses showing the differentially accumulation of NnUFGT2 is involved in the petal red-white bicolor pigmentation in lotus (Nelumbo nucifera).

Bicolor flower lotus is rare with high ornamental value. During the long history of breeding and artificial selection, a very famous lotus cultivar 'Da Sajin' with red and white picotee bicolor petals were obtained. In order to reveal the mechanism underlying the formation of its picotee bicolor pattern in the petal, an integrative metabolomics and proteomics analyses were conducted between red and white parts of its petals. The results showed that the defect of anthocyanidin 3-O-glucosyltransferases (UFGTs) accumulation resulted in the failure of the glycosylation of anthocyanidin, the last step of anthocyanin biosynthesis in white part of the petals. And proteomic data and biochemical analysis showed that the defect of UFGTs accumulation is not related to their transcription, but because of their degradation. Function of one differentially accumulated NnUFGT were proven being involved in anthocyanin biosynthesis through both in-vitro enzyme assay and in-vivo transgenic analyses. This regulation on the protein accumulation of structural genes in anthocyanin biosynthesis was not explored in any other plants, and hence supposed to be a novel mechanism for the formation of picotee bicolor pattern flower. The results not only provide some new insights into the understanding of lotus flower coloration, but also might assist the breeding of flower lotus.

Integration of metabolomics and transcriptomics analyses investigates the accumulation of secondary metabolites in maturing seed plumule of sacred lotus (Nelumbo nucifera).

Lotus seed plumule (LP) is rich in a variety of antioxidant and anti-inflammatory secondary metabolites, making it a traditional food and medicine widely used in China. Physiological and histological evidences indicated that LP mainly accumulated metabolites in 15-24 days after pollination (DAP) during their development. To systematically investigate the dynamic accumulation of major secondary metabolites, the UPLC-HRMS-based widely targeted metabolomics analyses were performed on maturing LP at 15, 18, 21, and 24 DAP. In total, 767 metabolites were identified, including many secondary metabolites, e.g., 27 % flavonoids and 8 % alkaloids. Among them, 591 were identified as differentially accumulated metabolites (DAMs). The majority of secondary metabolites showed great accumulation after 18 DAP even at the late stage of LP maturation, such as hesperidin, neohesperidin, orobol, serotonin, and lotus special O-nornuciferine, endowing mature LP with effective pharmaceutical properties. The paralleled transcriptomic analysis identified 11,019 differentially expressed genes (DEGs). Based on the comprehensive data, several systematical metabolic regulation maps were established for different secondary metabolites, and 18 DAP was found as a switching point for LP maturing from active primary metabolism to massive secondary metabolites deposition. This study provides valuable information for understanding the mechanism of secondary metabolite accumulation in maturing LP and facilitates its pharmaceutical application.

TMT-based quantitative proteomic and physiological analyses on lotus plumule of artificially aged seed in long-living sacred lotus Nelumbo nucifera.

Seed longevity is important for the maintenance of seed nutritional quality, vigor, and germination potential during storage. Sacred lotus is known as one of the longest living seeds in the world and their ability to maintain longevity has been widely investigated. In this study, a suitable controlled deterioration treatment (CDT) method was first established to evaluate the vigor loss of lotus plumule (LP), and then the Tandem Mass Tags (TMT)-based proteomic analysis was performed on LP from the CDT-treated seed to quantitatively and qualitatively analyze the protein profile dynamic. In total, 4002 proteins were successfully quantified, of them, 558 differently accumulated proteins (DAPs) were identified. Protein processing and RNA-related proteins were found more easily to be affected by CDT, which may directly result in seed vigor loss. Meanwhile, CDT resulted in remarkable up-regulation of numerous proteins related to antioxidation, photosynthesis, RNA and DNA stability, starch and sucrose mobilization, and cell membrane and wall stability, which potentially played key roles in maintaining the lotus seed vigor under CDT. Histological and physiological analyses were also performed to verify some proteome results. This study provided both fundamental data and new insights to further uncover the secret of lotus seed longevity. SIGNIFICANCE: Seed aging affects the seed quality and can result in direct economic losses. The exceptional longevity of sacred lotus seed has attracted extensive attention. In this study, an optimized CDT method was used to mimic the natural aging process of sacred lotus seed, and based on TMT-based quantitative proteomic analysis on the LP profile of CDT-treated seeds, a series of differentially accumulation of specific proteins (DEPs) were revealed related to CDT resistance. Correspondingly, the physiological state and histological structure of the LP along with the CDT were detected to verify the proteome data. This study provided comprehensive information for the molecular basis of lotus seed aging analysis and facilitate to screen seed longevity related proteins for other plant species.

Total alkaloids from the seed embryo of Nelumbo nucifera Gaertn. improve cognitive impairment in APP/PS1 mice and protect Aß-damaged PC12 cells.

The seed embryo of Nelumbo nucifera Gaertn. is a famous traditional Chinese medicine and food which is considered conducive to the prevention of Alzheimer's disease (AD). In this study, the effect and mechanism of TASENN (total alkaloids from the seed embryo of Nelumbo nucifera Gaertn.) on AD mice and amyloid-ß (Aß) injured PC12 cells were evaluated. HPLC-UV analysis showed that the extracted TASENN (purity = 95.6%) mainly contains Liensinine, Isoliensinine, and Neferine (purity was 23.01, 28.02, and 44.57%, respectively). In vivo, oral treatment with TASENN (50 mg/kg/day for 28 days) improved the learning and memory functions of APP/PS1 transgenic mice, ameliorated the histopathological changes of cortical and hippocampal neurons, and inhibited neuronal apoptosis. We found that TASENN reduced the phosphorylation of Tau and the formation of neurofibrillary tangles (NFTs) in APP/PS1 mouse brain. Moreover, TASENN down-regulated the expression of APP and BACE1, ameliorated Aß deposition, and inhibited microglial proliferation and aggregation. The elevated protein expression of CaM and p-CaMKII in APP/PS1 mouse brain was also reduced by TASENN. In vitro, TASENN inhibited the apoptosis of PC12 cells injured by Aß25-35 and increased the cell viability. Aß25-35-induced increase of cytosolic free Ca2+ level and high expression of CaM, p-CaMKII, and p-Tau were decreased by TASENN. Our findings indicate that TASENN has a potential therapeutic effect on AD mice and a protective effect on PC12 cells. The anti-AD activity of TASENN may be closely related to its negative regulation of the CaM pathway.

Novel Insights into Floral Thermogenesis: In Vivo Analyses of Mitochondrial Dynamics in Nelumbo nucifera Flowers.

Animal-like thermogenic (TM) activities in flowers have been reported in several families of seed plants. While an association of mitochondria with floral thermogenesis has been described, how mitochondrial dynamics are involved in the regulation of floral thermogenesis is unclear. In this study, the morphological and functional dynamics of mitochondria in vivo were assessed in Nelumbo nucifera Gaertn. flowers during floral thermogenesis. The results showed that mitochondrial biogenesis increased considerably in N. nucifera flowers during thermogenesis, accompanied by notable morphological changes in the mitochondria, including long elliptical, rod-shaped, and dumbbell-shaped morphologies, as well as increased mitochondrial reactive oxygen species (ROS) levels in TM cells. An increase in the expression of alternative oxidase (AOX) during the thermogenesis of N. nucifera flowers was also observed. These observations suggested the rapid change in mitochondrial morphology and increased density during thermogenesis implied activation of mitochondrial fission, which combined with elevated levels of mitochondrial ROS trigger a substantial increase in AOX within the respiratory pathway of TM N. nucifera.

Identification of the NAC Transcription Factors and Their Function in ABA and Salinity Response in Nelumbo nucifera.

Nelumbo nucifera Gaertn. is an important perennial aquatic herb that has high ornamental, edible, medicinal, and economic value, being widely distributed and used in China. The NAC superfamily (NAM, ATAF1/2, CUC2) plays critical roles in plant growth, development, and response to abiotic and biotic stresses. Though there have been a few reports about NAC genes in lotus, systematic analysis is still relatively lacking. The present study aimed to characterize all the NAC genes in the lotus and obtain better insights on the NnNACs in response to salt stress by depending on ABA signaling. Here, 97 NAC genes were identified by searching the whole lotus genome based on the raw HMM models of the conserved NAM domain and NAC domain. They were characterized by bioinformatics analysis and divided into 18 subgroups based on the phylogenetic tree. Cis-element analysis demonstrated that NAC genes are responsive to biotic and abiotic stresses, light, low temperature, and plant hormones. Meanwhile, NAC genes had tissue expression specificity. qRT-PCR analysis indicated that NAC genes could be upregulated or downregulated by NaCl treatment, ABA, and fluoridone. In addition, NAC016, NAC025, and NAC070, whose encoding genes were significantly induced by NaCl and ABA, were located in the nucleus. Further analysis showed the three NAC proteins had transcriptional activation capabilities. The co-expression network analysis reflected that NAC proteins may form complexes with other proteins to play a role together. Our study provides a theoretical basis for further research to be conducted on the regulatory mechanisms of salinity resistance in the lotus.

Sleep-promoting activity of lotus (Nelumbo nucifera) rhizome water extract via GABAA receptors.

CONTEXT: The sleep-promoting activity of Nelumbo nucifera Gaertn. (Nymphaeaceae) alkaloids in leaves or seeds are well known. However, the sleep-promoting activity of the lotus rhizome (LE), which is used mainly as food, has not yet been evaluated. OBJECTIVE: We investigated the sleep-promoting activity of LE water extract. MATERIALS AND METHODS: Institute of Cancer Research (ICR) mice (n = 8) were subject to a pentobarbital-induced sleep test to assess changes in sleep latency and duration following the administration of LE (80-150 mg/kg). In addition, electroencephalography analysis was performed to determine the sleep quality after LE treatment as well as the sleep recovery effect of LE using a caffeine-induced insomnia SD rat model. Real-time PCR and western blot analysis were performed to investigate the expression of neurotransmitter receptors, and the GABAA receptor antagonists were used for receptor binding analysis. RESULTS: An oral administration of 150 mg/kg LE significantly increased sleep duration by 24% compared to the control. Furthermore, LE increased nonrapid eye movement (NREM) sleep by increasing theta and delta powers. In the insomnia model, LE increased sleep time by increasing NREM sleep. Moreover, treatment with picrotoxin and flumazenil decreased the sleep time by 33% and 23%, respectively, indicating an involvement of the GABAA receptor in the sleep-enhancing activity of LE. The expression of GABAA receptors and the concentration of GABA in the brain were increased by LE. DISCUSSION AND CONCLUSIONS: The results suggest that the sleep-promoting activity of LE was via the GABAA receptor. Collectively, these data show that LE may promote sleep.

Inhibition of Three Diabetes-Related Enzymes by Procyanidins from Lotus (Nelumbo nucifera Gaertn.) Seedpods.

The inhibitory effects of procyanidins from lotus (Nelumbo nucifera Gaertn.) seedpods on the activities of α-amylase, α-glucosidase and protein tyrosine phosphatase 1B (PTP1B), were studied and compared with those of (+)-catechin, (-)-epicatechin, epigallocatechin gallate (EGCG), procyanidin dimer B2 and trimer C1. The results showed that Lotus procyanidin extract (LPE) significantly inhibited α-amylase, α-glucosidase and PTP1B with IC50 values of 5.5, 1.0, and 0.33 µg/mL, respectively. The inhibition increased with the degree of polymerization and the existence of galloyl or gallocatechin units. Kinetic analysis showed that LPE inhibited α-glucosidase activity in a mixed competitive and noncompetitive mode. Fluorescence quenching revealed that α-glucosidase interacted with LPE or EGCG in an apparent static mode, or the model of "sphere of action". The apparent static (K) and bimolecular (kq) constants were 4375 M-1 and 4.375 × 1011 M-1 s-1, respectively, for LPE and 1195 M-1 and 1.195 × 1011 M-1 s-1, respectively, for EGCG. Molecular docking analysis provided further information on the interactions of (+)-catechin, (-)-epicatechin, EGCG, B2 and C1 with α-glucosidase. It is hypothesized that LPE may bind to multiple sites of the enzyme through hydrogen bonding and hydrophobic interactions, leading to conformational changes in the enzyme and thus inhibiting its activity. These findings first elucidate the inhibitory effect of LPE on diabetes-related enzymes and highlight the usefulness of LPE as a dietary supplement for the prophylaxis of diabetes.

Hepatoprotective Activity of Nelumbo nucifera Gaertn. Seedpod Extract Attenuated Acetaminophen-Induced Hepatotoxicity.

The aim is to investigate the effect of lotus (Nelumbo nucifera Gaertn.) seedpod extract (LSE) on acetaminophen (APAP)-induced hepatotoxicity. LSE is rich in polyphenols and has potent antioxidant capacity. APAP is a commonly used analgesic, while APAP overdose is the main reason for drug toxicity in the liver. Until now, there has been no in vitro test of LSE in drug-induced hepatotoxicity responses. LSEs were used to evaluate the effect on APAP-induced cytotoxicity, ROS level, apoptotic rate, and molecule mechanisms. The co-treatment of APAP and LSEs elevated the survival rate and decreased intracellular ROS levels on HepG2 cells. LSEs treatment could significantly reduce APAP-induced HepG2 apoptosis assessed by DAPI and Annexin V/PI. The further molecule mechanisms indicated that LSEs decreased Fas/FasL binding and reduced Bax and tBid to restore mitochondrial structure and subsequently suppress downstream apoptosis cascade activation. These declines in COX-2, NF-κB, and iNOS levels were observed in co-treatment APAP and LSEs, which indicated that LSEs could ameliorate APAP-induced inflammation. LSE protected APAP-induced apoptosis by preventing extrinsic, intrinsic, and JNK-mediated pathways. In addition, the restoration of mitochondria and inflammatory suppression in LSEs treatments indicated that LSEs could decrease oxidative stress induced by toxic APAP. Therefore, LSE could be a novel therapeutic option for an antidote against overdose of APAP.

Comprehensive Analysis for GRF Transcription Factors in Sacred Lotus (Nelumbo nucifera).

Sacred lotus (Nelumbo nucifera) is an aquatic perennial plant with essential food, ornamental, and pharmacological value. Growth-regulating factor (GRF) is a transcription factor (TF) family that plays an important role in regulating the growth and development of plants. In this study, a comprehensive analysis of the GRF family in N. nucifera was performed, and its role in N. nucifera development was studied. A total of eight GRF genes were identified in the N. nucifera genome. Phylogenetic analysis divided the 38 GRF genes into six clades, while the NuGRFs only contained five clades. The analyses of gene structures, motifs, and cis-acting regulatory elements of the GRF gene family were performed. In addition, the chromosome location and collinearity were analyzed. The expression pattern based on transcriptomic data and real-time reverse transcription-quantitative PCR (qRT-PCR) revealed that the GRF genes were expressed in multiple organs and were abundant in actively growing tissues, and the expression levels decreased as the age of N. nucifera increased. Then, 3D structures of the NuGRF proteins were predicted by homology modeling. Finally, the subcellular localization of GRF1 was ascertained in the tobacco leaf through a vector. Therefore, this study provides a comprehensive overview of the GRF TF family in N. nucifera.

The Genome-Wide Identification of Long Non-Coding RNAs Involved in Floral Thermogenesis in Nelumbo nucifera Gaertn.

The sacred lotus (Nelumbo nucifera Gaertn.) can maintain a stable floral chamber temperature when blooming, despite ambient temperature fluctuations; however, the long non-coding RNAs (lncRNAs) involved in floral thermogenesis remain unclear. In the present study, we obtain comprehensive lncRNAs expression profiles from receptacles at five developmental stages by strand-specific RNA sequencing to reveal the lncRNAs regulatory mechanism of the floral thermogenesis of N. nucifera. A total of 22,693 transcripts were identified as lncRNAs, of which approximately 44.78% had stage-specific expression patterns. Subsequently, we identified 2579 differential expressed lncRNAs (DELs) regulating 2367 protein-coding genes mainly involved in receptacle development and reproductive process. Then, lncRNAs with floral thermogenesis identified by weighted gene co-expression network analysis (WGCNA) were mainly related to sulfur metabolism and mitochondrial electron transport chains. Meanwhile, 70 lncRNAs were predicted to act as endogenous target mimics (eTMs) for 29 miRNAs and participate in the regulation of 16 floral thermogenesis-related genes. Our dual luciferase reporter assays indicated that lncRNA LTCONS_00068702 acted as eTMs for miR164a_4 to regulate the expression of TrxL2 gene. These results deepen our understanding of the regulation mechanism of floral thermogenesis by lncRNAs and accumulate data for further research.

Simulated digestion and in vitro fermentation of a polysaccharide from lotus (Nelumbo nucifera Gaertn.) root residue by the human gut microbiota.

Lotus root polysaccharide (LRP) is an active water-soluble polysaccharide with average molecular weight of 1.24 × 104. It was composed of (1 â†’ 4)-α-D-glucan backbone with α-D-glycopyranosyl moieties connected to C-6 positions of the glucose residues as side chains approximately every six residues. However, little information is available for its digestion and fermentation characteristics in vitro. The results showed that the levels of reducing sugars were increased slightly, and the molecular weight was also reduced slightly, in simulated gastric and small intestinal juices. During in vitro fermentation, the total sugar, reducing sugar and glucose contents decreased gradually with increasing fermentation time. The molecular of LRP was degraded and to metabolize into a variety the short-chain fatty acids (SCFAs) such as acetic, propionic, and butyric acids. Furthermore, LRP fermentation decreased the pH of the fermentation broth and increased its absorbance. Meanwhile, LRP modulated the gut microbiota by altering the Firmicutes/Bacteroidetes ratio and increasing the relative abundance of Bifidobacterium. The findings from this study showed that LRP could be developed as potential prebiotic to regulate the composition of gut microbiota, thereby promote the production of SCFAs.

Comparative analyses of American and Asian lotus genomes reveal insights into petal color, carpel thermogenesis and domestication.

Nelumbo lutea (American lotus), which differs from Nelumbo nucifera (Asian lotus) morphologically, is one of the two remaining species in the basal eudicot family Nelumbonaceae. Here, we assembled the 843-Mb genome of American lotus into eight pseudochromosomes containing 31 382 protein-coding genes. Comparative analyses revealed conserved synteny without large chromosomal rearrangements between the genomes of American and Asian lotus and identified 29 533 structural variants (SVs). Carotenoid and anthocyanin pigments determine the yellow and red petal colors of American and Asian lotus, respectively. The structural genes encoding enzymes of the carotenoid and anthocyanin biosynthesis pathways were conserved between two species but differed in expression. We detected SVs caused by repetitive sequence expansion or contraction among the anthocyanin biosynthesis regulatory MYB genes. Further transient overexpression of candidate NnMYB5 induced anthocyanin accumulation in lotus petals. Alternative oxidase (AOX), uncoupling proteins (UCPs), and sugar metabolism and transportation contributed to carpel thermogenesis. Carpels produce heat with sugars transported from leaves as the main substrates, because there was weak tonoplast sugar transporter (TST) activity, and with SWEETs were highly expressed during thermogenesis. Cell proliferation-related activities were particularly enhanced in the warmer carpels compared with stamens during the cold night before blooming, which suggested that thermogenesis plays an important role in flower protogyny. Population genomic analyses revealed deep divergence between American and Asian lotus, and independent domestication affecting seed, rhizome, and flower traits. Our findings provide a high-quality reference genome of American lotus for exploring the genetic divergence and variation between two species and revealed possible genomic bases for petal color, carpel thermogenesis and domestication in lotus.

Color fading in lotus (Nelumbo nucifera) petals is manipulated both by anthocyanin biosynthesis reduction and active degradation.

Flower color is a key trait that determines the ornamental quality of aquatic lotus (Nelumbo nucifera). Color fading significantly decreases the ornamental value of lotus flowers. However, the molecular mechanism underlying lotus petal discoloration remains largely unknown. Here, the anthocyanin content and global transcriptional profiling of lotus petals of cultivar 'Qiusanse' in four developmental stages were analyzed. Five anthocyanin components were detected, and the total anthocyanin content decreased as the petal color changed from red to nearly white. Moreover, the malondialdehyde (MDA) content and peroxidase (POD) activity increased during color fading. RNA-seq analysis revealed a total of 4,092 differentially expressed genes (DEGs) between petal developmental stages. Notably, oxidoreductase and hydrolase activity related genes were overrepresented in DEGs. The expression pattern of key anthocyanin biosynthesis genes including, CHS, F3H, ANS, UFGT, and transcription factor regulators, including MYBs, WRKYs and bHLHs were correlated with anthocyanin accumulation. Interestingly, DEGs associated with anthocyanin degradation and vacuolar pH regulation, including peroxidase, proton pumps regulators such as WRKY3 and MYB5-like, were significantly upregulated during the late stages of flowering. This study reveals for the first time the transcriptional dynamics during lotus petal discoloration. Our results suggest the involvement of anthocyanin biosynthesis repressors and degrading genes as well as pH regulators in controlling color fading of lotus petals. The study also provides valuable information and candidate genes for improving the lotus flower color.