The Influence of Emodin Succinyl Ethyl Ester on Non-alcoholic Steatohepatitis Induced by a Diet High in Fructose, Cholesterol, and Fat in Mice.

Nonalcoholic steatohepatitis (NASH) is a subtype of nonalcoholic fatty liver disease (NAFLD) characterized by hepatic steatosis and evidence of hepatocyte injury (ballooning) and inflammation, with or without liver fibrosis. In this study, after 12 weeks of induction, the mice were treated with emodin succinyl ethyl ester (ESEE) for four weeks at doses of 10/30/90 mg/kg/d. The blood analysis of experimental endpoints showed that ESEE exhibited significant therapeutic effects on the progression of disorders of glycolipid metabolism and the induced liver injury in the model animals. Histopathological diagnosis of the liver and total triglyceride measurements revealed that ESEE had a significant therapeutic effect on the histopathological features of nonalcoholic fatty liver disease/hepatitis, such as cellular steatosis and activation of intrahepatic inflammation. Additionally, ESEE was able to improve hepatocyte fat deposition, steatosis, and the course of intrahepatic inflammatory activity. Furthermore, it showed some inhibitory effect on liver fibrosis in the model animals. In summary, this study confirms the therapeutic effects of ESEE on the NAFLD/NASH model in C57BL/6J mice induced by a high-fat, high cholesterol, and fructose diet. These effects were observed through improvements in liver function, inhibition of fibrosis, and inflammatory responses. Changes in blood glucose levels, blood lipid metabolism, liver histopathological staining, liver fibrosis staining, and related pathological scores further supported the therapeutic effects of ESEE. Therefore, this study has important implications for the exploration of novel drugs for nonalcoholic fatty liver disease.

Assessment of the impact of climate change on endangered conifer tree species by considering climate and soil dual suitability and interspecific competition.

Climate change results in the habitat loss of many conifer tree species and jeopardizes species biodiversity and forest ecological functions. Delineating suitable habitats for tree species via climate niche model (CNM) is widely used to predict the impact of climate change and develop conservation and management strategies. However, the robustness of CNM is broadly debated as it usually does not consider soil and competition factors. Here we developed a new approach to combine soil variables with CNM and evaluate interspecific competition potential in the niche overlapping areas. We used an endangered conifer species - Chamaecyparis formosensis (red cypress) - as a case study to predict the impact of climate change. We developed a novel approach to integrate the climate niche model and soil niche model predictions and considered interspecific competition to predict the impacts of climate change on tree species. Our results show that the suitable habitat for red cypress would decrease significantly in the future with an additional threat from the competition of an oak tree species. Our approach and results may represent significant implications in making conservation strategies and evaluating the impacts of climate change, and providing the direction of the refinement of the ecological niche model.

GbFLSa overexpression negatively regulates proanthocyanin biosynthesis.

Flavonoids are important secondary metabolites with extensive pharmacological functions. Ginkgo biloba L. (ginkgo) has attracted extensive attention because of its high flavonoid medicinal value. However, little is understood about ginkgo flavonol biosynthesis. Herein, we cloned the full-length gingko GbFLSa gene (1314 bp), which encodes a 363 amino acid protein that has a typical 2-oxoglutarate (2OG)-Fe(II) oxygenase region. Recombinant GbFLSa protein with a molecular mass of 41 kDa was expressed in Escherichia coli BL21(DE3). The protein was localized to the cytoplasm. Moreover, proanthocyanins, including catechin, epicatechin, epigallocatechin and gallocatechin, were significantly less abundant in transgenic poplar than in nontransgenic (CK) plants. In addition, dihydroflavonol 4-reductase, anthocyanidin synthase and leucoanthocyanidin reductase expression levels were significantly lower than those of their CK counterparts. GbFLSa thus encodes a functional protein that might negatively regulate proanthocyanin biosynthesis. This study helps elucidate the role of GbFLSa in plant metabolism and the potential molecular mechanism of flavonoid biosynthesis.

Predicting suitable habitats of Melia azedarach L. in China using data mining.

Melia azedarach L. is an important economic tree widely distributed in tropical and subtropical regions of China and some other countries. However, it is unclear how the species' suitable habitat will respond to future climate changes. We aimed to select the most accurate one among seven data mining models to predict the current and future suitable habitats for M. azedarach in China. These models include: maximum entropy (MaxEnt), support vector machine (SVM), generalized linear model (GLM), random forest (RF), naive bayesian model (NBM), extreme gradient boosting (XGBoost), and gradient boosting machine (GBM). A total of 906 M. azedarach locations were identified, and sixteen climate predictors were used for model building. The models' validity was assessed using three measures (Area Under the Curves (AUC), kappa, and overall accuracy (OA)). We found that the RF provided the most outstanding performance in prediction power and generalization capacity. The top climate factors affecting the species' suitable habitats were mean coldest month temperature (MCMT), followed by the number of frost-free days (NFFD), degree-days above 18 °C (DD > 18), temperature difference between MWMT and MCMT, or continentality (TD), mean annual precipitation (MAP), and degree-days below 18 °C (DD < 18). We projected that future suitable habitat of this species would increase under both the RCP4.5 and RCP8.5 scenarios for the 2011-2040 (2020s), 2041-2070 (2050s), and 2071-2100 (2080s). Our findings are expected to assist in better understanding the impact of climate change on the species and provide scientific basis for its planting and conservation.

Full-Length Transcriptome Sequencing and Comparative Transcriptomic Analyses Provide Comprehensive Insight Into Molecular Mechanisms of Cellulose and Lignin Biosynthesis in Cunninghamia lanceolata.

Cunninghamia lanceolata is an essential timber species that provide 20%-30% raw materials for China's timber industry. Although a few transcriptomes have been published in C. lanceolata, full-length mRNA transcripts and regulatory mechanisms behind the cellulose and lignin biosynthesis have not been thoroughly investigated. Here, PacBio Iso-seq and RNA-seq analyses were adapted to identify the full-length and differentially expressed transcripts along a developmental gradient from apex to base of C. lanceolata shoots. A total of 48,846 high-quality full-length transcripts were obtained, of which 88.0% are completed transcriptome based on benchmarking universal single-copy orthologs (BUSCO) assessment. Along stem developmental gradient, 18,714 differentially expressed genes (DEGs) were detected. Further, 28 and 125 DEGs were identified as enzyme-coding genes of cellulose and lignin biosynthesis, respectively. Moreover, 57 transcription factors (TFs), including MYB and NAC, were identified to be involved in the regulatory network of cellulose and lignin biosynthesis through weighted gene co-expression network analysis (WGCNA). These TFs are composed of a comparable regulatory network of secondary cell wall formation in angiosperms, revealing a similar mechanism may exist in gymnosperms. Further, through qRT-PCR, we also investigated eight specific TFs involved in compression wood formation. Our findings provide a comprehensive and valuable source for molecular genetics breeding of C. lanceolata and will be beneficial for molecular-assisted selection.

Using landscape genomics to delineate seed and breeding zones for lodgepole pine.

Seed and breeding zones traditionally are delineated based on local adaptation of phenotypic traits associated with climate variables, an approach requiring long-term field experiments. In this study, we applied a landscape genomics approach to delineate seed and breeding zones for lodgepole pine. We used a gradient forest (GF) model to select environment-associated single nucleotide polymorphisms (SNPs) using three SNP datasets (full, neutral and candidate) and 20 climate variables for 1906 lodgepole pine (Pinus contorta) individuals in British Columbia and Alberta, Canada. The two GF models built with the full (28 954) and candidate (982) SNPs were compared. The GF models identified winter-related climate as major climatic factors driving genomic patterns of lodgepole pine's local adaptation. Based on the genomic gradients predicted by the full and candidate GF models, lodgepole pine distribution range in British Columbia and Alberta was delineated into six seed and breeding zones. Our approach is a novel and effective alternative to traditional common garden approaches for delineating seed and breeding zone, and could be applied to tree species lacking data from provenance trials or common garden experiments.

Adaptation to climate change through seasonal migration revealed by climatic versus demographic niche models.

Predicting the geographic range of species and their response to climatic variation and change are entwined goals in conservation and evolutionary ecology. Species distribution models (SDMs) are foundational in this effort and used to visualize the geographic range of species as the spatial representation of its realized niche. SDMs are also used to forecast range shifts under climate change, but often in the absence of empirical evidence that climate limits population growth. We explored the influence of climate on demography, seasonal migration, and the extent of the geographic range in song sparrows (Melospiza melodia), a species thought to display marked local adaptation to regional climate. To do so, we developed SDMs to predict the demographic and climate niches of migratory and resident song sparrows across our study area in western North America from California to Alaska, using 48 years of demographic data from a focal population in British Columbia and 1.2 million continental-scale citizen science observations. Spatial agreement of our demographic and climate niche models in the region of our focal population was strong (76%), supporting the hypothesis that demographic performance and the occurrence of seasonal migration varied predictably with climatic conditions. In contrast, agreement at the northern (58%) and southern (40%) extents of our study area was lower, as expected if the factors limiting population growth vary regionally. Our results support the hypothesis that local climate drives spatial variation in the occurrence of seasonal migration in song sparrows by limiting the fitness of year-round residents, and suggest that climate warming has favored range expansions and facilitated an upward shift in elevational range song sparrows that forgo seasonal migration. Our work highlights the potential role of seasonal migration in climate adaptation and limits on the reliability of climate niche models not validated with demographic data.

A comprehensive annotation dataset of intact LTR retrotransposons of 300 plant genomes.

LTR retrotransposons (LTR-RTs) are ubiquitous and represent the dominant repeat element in plant genomes, playing important roles in functional variation, genome plasticity and evolution. With the advent of new sequencing technologies, a growing number of whole-genome sequences have been made publicly available, making it possible to carry out systematic analyses of LTR-RTs. However, a comprehensive and unified annotation of LTR-RTs in plant groups is still lacking. Here, we constructed a plant intact LTR-RTs dataset, which is designed to classify and annotate intact LTR-RTs with a standardized procedure. The dataset currently comprises a total of 2,593,685 intact LTR-RTs from genomes of 300 plant species representing 93 families of 46 orders. The dataset is accompanied by sequence, diverse structural and functional annotation, age determination and classification information associated with the LTR-RTs. This dataset will contribute valuable resources for investigating the evolutionary dynamics and functional implications of LTR-RTs in plant genomes.

Metabolome and Transcriptome Analyses Reveal the Regulatory Mechanisms of Photosynthesis in Developing Ginkgo biloba Leaves.

Ginkgo (Ginkgo biloba L.) is a deciduous tree species with high timber, medicinal, ecological, ornamental, and scientific values, and is widely cultivated worldwide. However, for such an important tree species, the regulatory mechanisms involved in the photosynthesis of developing leaves remain largely unknown. Here, we observed variations in light response curves (LRCs) and photosynthetic parameters (photosynthetic capacity (Pnmax) and dark respiration rate (Rd)) of leaves across different developmental stages. We found the divergence in the abundance of compounds (such as 3-phospho-d-glyceroyl phosphate, sedoheptulose-1,7-bisphosphate, and malate) involved in photosynthetic carbon metabolism. Additionally, a co-expression network was constructed to reveal 242 correlations between transcription factors (TFs) and photosynthesis-related genes (p < 0.05, |r| > 0.8). We found that the genes involved in the photosynthetic light reaction pathway were regulated by multiple TFs, such as bHLH, WRKY, ARF, IDD, and TFIIIA. Our analysis allowed the identification of candidate genes that most likely regulate photosynthesis, primary carbon metabolism, and plant development and as such, provide a theoretical basis for improving the photosynthetic capacity and yield of ginkgo trees.

Temporospatial Flavonoids Metabolism Variation in Ginkgo biloba Leaves.

Ginkgo (Ginkgo biloba L.) is a high-value medicinal tree species characterized by its flavonoids beneficial effects that are abundant in leaves. We performed a temporospatial comprehensive transcriptome and metabolome dynamics analyses of clonally propagated Ginkgo plants at four developmental stages (time: May to August) across three different environments (space) to unravel leaves flavonoids biosynthesis variation. Principal component analysis revealed clear gene expression separation across samples from different environments and leaf-developmental stages. We found that flavonoid-related metabolism was more active in the early stage of leaf development, and the content of total flavonoid glycosides and the expression of some genes in flavonoid biosynthesis pathway peaked in May. We also constructed a co-expression regulation network and identified eight GbMYBs and combining with other TF genes (3 GbERFs, 1 GbbHLH, and 1 GbTrihelix) positively regulated the expression of multiple structural genes in the flavonoid biosynthesis pathway. We found that part of these GbTFs (Gb_11316, Gb_32143, and Gb_00128) expressions was negatively correlated with mean minimum temperature and mean relative humidity, while positively correlated with sunshine duration. This study increased our understanding of the molecular mechanisms of flavonoids biosynthesis in Ginkgo leaves and provided insight into the proper production and management of Ginkgo commercial plantations.

Metabolomic and transcriptomic analyses of mutant yellow leaves provide insights into pigment synthesis and metabolism in Ginkgo biloba.

BACKGROUND: Ginkgo (Ginkgo biloba L.) is an excellent landscape species. Its yellow-green leaf mutants are ideal materials for research on pigment synthesis, but the regulatory mechanism of leaf coloration in these ginkgo mutants remains unclear. RESULTS: We compared the metabolomes and transcriptomes of green and mutant yellow leaves of ginkgo over the same period in this study. The results showed that the chlorophyll content of normal green leaves was significantly higher than that of mutant yellow leaves of ginkgo. We obtained 931.52M clean reads from different color leaves of ginkgo. A total of 283 substances in the metabolic profiles were finally detected, including 50 significantly differentially expressed metabolites (DEMs). We identified these DEMs and 1361 differentially expressed genes (DEGs), with 37, 4, 3 and 13 DEGs involved in the photosynthesis, chlorophyll, carotenoid, and flavonoid biosynthesis pathways, respectively. Moreover, integrative analysis of the metabolomes and transcriptomes revealed that the flavonoid pathway contained the upregulated DEM (-)-epicatechin. Fourteen DEGs from the photosynthesis pathway were positively or negatively correlated with the DEMs. CONCLUSIONS: Our findings suggest a complex metabolic network in mutant yellow leaves. This study will provide a basis for studies of leaf color variation and regulation.

ClimateEU, scale-free climate normals, historical time series, and future projections for Europe.

Interpolated climate data have become essential for regional or local climate change impact assessments and the development of climate change adaptation strategies. Here, we contribute an accessible, comprehensive database of interpolated climate data for Europe that includes monthly, annual, decadal, and 30-year normal climate data for the last 119 years (1901 to 2019) as well as multi-model CMIP5 climate change projections for the 21st century. The database also includes variables relevant for ecological research and infrastructure planning, comprising more than 20,000 climate grids that can be queried with a provided ClimateEU software package. In addition, 1 km and 2.5 km resolution gridded data generated by the software are available for download. The quality of ClimateEU estimates was evaluated against weather station data for a representative subset of climate variables. Dynamic environmental lapse rate algorithms employed by the software to generate scale-free climate variables for specific locations lead to improvements of 10 to 50% in accuracy compared to gridded data. We conclude with a discussion of applications and limitations of this database.

Overexpression of GbF3'5'H1 Provides a Potential to Improve the Content of Epicatechin and Gallocatechin.

The flavonoids in Ginkgo biloba L. (ginkgo) have important medicinal uses due to their antioxidant, antitumor, and blood circulation-promoting effects. However, the genetic mechanisms underlying flavonoid biosynthesis in ginkgo remain elusive. Flavonoid 3', 5'-hydroxylase (F3'5'H) is an important enzyme in flavonoid synthesis. We detected a novel differentially expressed GbF3'5'H1 gene homologous to the F3'5'H enzyme involved in the flavonoid synthesis pathway through transcriptome sequencing. In this study, we characterized this gene, performed an expression analysis, and heterologously overexpressed GbF3'5'H1 in Populus. Our results showed that GbF3'5'H1 is abundant in the leaf and highly expressed during April. We also found four metabolites closely related to flavonoid biosynthesis. Importantly, the contents of 4',5-dihydroxy-7-glucosyloxyflavanone, epicatechin, and gallocatechin were significantly higher in transgenic plants than in nontransgenic plants. Our findings revealed that the GbF3'5'H1 gene functions in the biosynthesis of flavonoid-related metabolites, suggesting that GbF3'5'H1 represents a prime candidate for future studies (e.g., gene-editing) aiming to optimize ginkgo flavonoid production, especially that of flavan-3-ols.

Landscape genomics predicts climate change-related genetic offset for the widespread Platycladus orientalis (Cupressaceae).

Understanding and quantifying populations' adaptive genetic variation and their response to climate change are critical to reforestation's seed source selection, forest management decisions, and gene conservation. Landscape genomics combined with geographic and environmental information provide an opportunity to interrogate forest populations' genome-wide variation for understanding the extent to which evolutionary forces shape past and contemporary populations' genetic structure, and identify those populations that may be most at risk under future climate change. Here, we used genotyping by sequencing to generate over 11,000 high-quality variants from Platycladus orientalis range-wide collection to evaluate its diversity and to predict genetic offset under future climate scenarios. Platycladus orientalis is a widespread conifer in China with significant ecological, timber, and medicinal values. We found population structure and evidences of isolation by environment, indicative of adaptation to local conditions. Gradient forest modeling identified temperature-related variables as the most important environmental factors influencing genetic variation and predicted areas with higher risk under future climate change. This study provides an important reference for forest resource management and conservation for P. orientalis.

Transcriptome-wide identification and profiling of miRNAs in a stress-tolerant conifer Sabina chinensis.

miRNAs are important regulatory components involving in many biological processes, including plant development, vegetative and reproductive growth, and stress response. However, identification and characterization of miRNAs still remain limited for conifer species. In this study, with deep sequencing, we obtained 1,314,450 unique reads with 18-30 nt length from a stress-tolerant conifer, Sabina chinensis. We identified 37 conserved and 103 novel miRNAs, their unique characteristics were further analyzed, and 10 randomly selected were validated by qRT-PCR. Through miRNA target predictions and annotations, we found miRNA may have several targets as well a target could be regulated by several miRNAs, and a total of 2,397 mRNAs were predicted to be targets of the 140 miRNAs. These targets included not only important transcription factors such as auxin response factors, but also indispensable non-transcriptional factor proteins. Pathway-based analysis showed that S. chinensis miRNAs are involved in 172 metabolic pathways, of which 3 were discovered in adaptation-related pathways, indicating their possible relevance to the species' stress-tolerance characteristics. This study is expected to lay the foundation for exploring the regulative roles of miRNAs in development, growth, and response to environmental stresses of S. chinensis.

Evaluating genomic data for management of local adaptation in a changing climate: A lodgepole pine case study.

We evaluate genomic data, relative to phenotypic and climatic data, as a basis for assisted gene flow and genetic conservation. Using a seedling common garden trial of 281 lodgepole pine (Pinus contorta) populations from across western Canada, we compare genomic data to phenotypic and climatic data to assess their effectiveness in characterizing the climatic drivers and spatial scale of local adaptation in this species. We find that phenotype-associated loci are equivalent or slightly superior to climate data for describing local adaptation in seedling traits, but that climate data are superior to genomic data that have not been selected for phenotypic associations. We also find agreement between the climate variables associated with genomic variation and with 20-year heights from a long-term provenance trial, suggesting that genomic data may be a viable option for identifying climatic drivers of local adaptation where phenotypic data are unavailable. Genetic clines associated with the experimental traits occur at broad spatial scales, suggesting that standing variation of adaptive alleles for this and similar species does not require management at scales finer than those indicated by phenotypic data. This study demonstrates that genomic data are most useful when paired with phenotypic data, but can also fill some of the traditional roles of phenotypic data in management of species for which phenotypic trials are not feasible.

Overexpression of the GbF3'H1 Gene Enhanced the Epigallocatechin, Gallocatechin, and Catechin Contents in Transgenic Populus.

Ginkgo biloba L. leaves are a flavonoid resource for the pharmaceutical industry. The flavonoid 3'-hydroxylase (F3'H) is a key enzyme in the flavonoid biosynthesis pathway. However, the role of F3'H in flavonoid biosynthesis and metabolism is unclear. In this study, we characterized and functionally analyzed the ginkgo F3'H gene GbF3'H1 that encodes a protein of 520 amino acids. Expression profiling showed that GbF3'H1 was highly expressed in the leaves of ginkgo in September. Subcellular localization showed that GbF3'H1 occurred predominately in the cytoplasm. Transgenic poplars overexpressing GbF3'H1 had more red pigmentation in leaves than did wild-type (WT) plants. Furthermore, the concentrations of epigallocatechin, gallocatechin, and catechin in the downstream products synthesized by flavonoids were significantly higher in the transgenic plants than in the WT plants. These results indicate that the overexpression of GbF3'H1 enhances flavonoid production in transgenic plants and provides new insights into flavonoid biosynthesis and metabolism.

Regulation of flavonoid metabolism in ginkgo leaves in response to different day-night temperature combinations.

Flavonoids are the most important secondary metabolites in ginkgo (Ginkgo biloba L.) leaves that determine its medicinal quality. Studies have suggested that secondary metabolism is strongly affected by temperature in other plant species, but little is known about ginkgo. In this study, we investigated the effects of different day-night temperature combinations (15/10, 25/20, and 35/30 °C (day/night)) on key enzyme activity, growth regulator concentrations, and flavonoid accumulation in ginkgo leaves. We found that phenylalanine ammonia-lyase (PAL) activity was enhanced and inhibited at 15/10 and 35/30 °C, respectively. Cinnamate-4-hydroxylase (C4H) activity was relatively stable under the three temperature conditions, and the p-coumarate CoA ligase (4CL) activity showed different trends under the three temperature conditions. The concentrations of flavonoid constituents (quercetin, kaempferol, and isorhamnetin) were decreased and increased under the 35/30 and 15/10 °C conditions, respectively. Low temperature promoted soluble sugar accumulation, while temperature had a limited impact on the accumulation of soluble protein. The pattern of change in the total flavonoid concentration was not always in agreement with PAL activity due to its complex pathway. Indoleacetic acid (IAA) and gibberellin (GA) changes shared similar patterns and had limited effects on flavonoid accumulation, while abscisic acid (ABA) acted as a promotor of flavonoid accumulation under high-temperature conditions. The total flavonoids achieved the highest content under the 15/10 °C treatment on the 40th day. Therefore, the lower temperature (15/10 °C) is more favorable for flavonoid accumulation and will provide a theoretical basis for further study.

Transcriptional profiling of long noncoding RNAs associated with leaf-color mutation in Ginkgo biloba L.

BACKGROUND: Long noncoding RNAs (lncRNAs) play an important role in diverse biological processes and have been widely studied in recent years. However, the roles of lncRNAs in leaf pigment formation in ginkgo (Ginkgo biloba L.) remain poorly understood. RESULTS: In this study, lncRNA libraries for mutant yellow-leaf and normal green-leaf ginkgo trees were constructed via high-throughput sequencing. A total of 2044 lncRNAs were obtained with an average length of 702 nt and typically harbored 2 exons. We identified 238 differentially expressed lncRNAs (DELs), 32 DELs and 49 differentially expressed mRNAs (DEGs) that constituted coexpression networks. We also found that 48 cis-acting DELs regulated 72 target genes, and 31 trans-acting DELs regulated 31 different target genes, which provides a new perspective for the regulation of the leaf-color mutation. Due to the crucial regulatory roles of lncRNAs in a wide range of biological processes, we conducted in-depth studies on the DELs and their targets and found that the chloroplast thylakoid membrane subcategory and the photosynthesis pathways (ko00195) were most enriched, suggesting their potential roles in leaf coloration mechanisms. In addition, our correlation analysis indicates that eight DELs and 68 transcription factors (TFs) might be involved in interaction networks. CONCLUSIONS: This study has enriched the knowledge concerning lncRNAs and provides new insights into the function of lncRNAs in leaf-color mutations, which will benefit future selective breeding of ginkgo.