Transcriptomic analysis of hub genes regulating albinism in light- and temperature-sensitive albino tea cultivars 'Zhonghuang 1' and 'Zhonghuang 2'.

Albino tea cultivars have high economic value because their young leaves contain enhanced free amino acids that improve the quality and properties of tea. Zhonghuang 1 (ZH1) and Zhonghuang 2 (ZH2) are two such cultivars widely planted in China; however, the environmental factors and molecular mechanisms regulating their yellow-leaf phenotype remain unclear. In this study, we demonstrated that both ZH1 and ZH2 are light- and temperature-sensitive. Under natural sunlight and low-temperature conditions, their young shoots were yellow with decreased chlorophyll and an abnormal chloroplast ultrastructure. Conversely, young shoots were green with increased chlorophyll and a normal chloroplast ultrastructure under shading and high-temperature conditions. RNA-seq analysis was performed for high light and low light conditions, and pairwise comparisons identified genes exhibiting different light responses between albino and green-leaf cultivars, including transcription factors, cytochrome P450 genes, and heat shock proteins. Weighted gene coexpression network analyses of RNA-seq data identified the modules related to chlorophyll differences between cultivars. Genes involved in chloroplast biogenesis and development, light signaling, and JA biosynthesis and signaling were typically downregulated in albino cultivars, accompanied by a decrease in JA-ILE content in ZH2 during the albino period. Furthermore, we identified the hub genes that may regulate the yellow-leaf phenotype of ZH1 and ZH2, including CsGDC1, CsALB4, CsGUN4, and a TPR gene (TEA010575.1), which were related to chloroplast biogenesis. This study provides new insights into the molecular mechanisms underlying leaf color formation in albino tea cultivars.

High-density genetic map construction and QTL mapping of a zigzag-shaped stem trait in tea plant (Camellia sinensis).

The highly unique zigzag-shaped stem phenotype in tea plants boasts significant ornamental value and is exceptionally rare. To investigate the genetic mechanism behind this trait, we developed BC1 artificial hybrid populations. Our genetic analysis revealed the zigzag-shaped trait as a qualitative trait. Utilizing whole-genome resequencing, we constructed a high-density genetic map from the BC1 population, incorporating 5,250 SNP markers across 15 linkage groups, covering 3,328.51 cM with an average marker interval distance of 0.68 cM. A quantitative trait locus (QTL) for the zigzag-shaped trait was identified on chromosome 4, within a 61.2 to 97.2 Mb range, accounting for a phenotypic variation explained (PVE) value of 13.62%. Within this QTL, six candidate genes were pinpointed. To better understand their roles, we analyzed gene expression in various tissues and individuals with erect and zigzag-shaped stems. The results implicated CsXTH (CSS0035625) and CsCIPK14 (CSS0044366) as potential key contributors to the zigzag-shaped stem formation. These discoveries lay a robust foundation for future functional genetic mapping and tea plant genetic enhancement.

Genome-wide association study, population structure, and genetic diversity of the tea plant in Guizhou Plateau.

BACKGROUND: Guizhou Plateau, as one of the original centers of tea plant, has a profound multi-ethnic cultural heritage and abundant tea germplasm resources. However, the impact of indigenous community factors on the genetic diversity, population structure and geographical distribution of tea plant is still unclear. RESULTS: Using the genotyping-by-sequencing (GBS) approach, we collected 415 tea plant accessions from the study sites, estimated genetic diversity, developed a core collection, and conducted a genome-wide association study (GWAS) based on 99,363 high-quality single-nucleotide polymorphisms (SNPs). A total of 415 tea accessions were clustered into six populations (GP01, GP02, GP03, GP04, GP05 and GP06), and the results showed that GP04 and GP05 had the highest and lowest genetic diversity (Pi = 0.214 and Pi = 0.145, respectively). Moreover, 136 tea accessions (33%) were selected to construct the core set that can represent the genetic diversity of the whole collection. By analyzing seven significant SNP markers associated with the traits such as the germination period of one bud and two leaves (OTL) and the germination period of one bud and three leaves (OtL), four candidate genes possibly related to OTL and OtL were identified. CONCLUSIONS: This study revealed the impact of indigenous communities on the population structure of 415 tea accessions, indicating the importance of cultural practices for protection and utilization of tea plant genetic resources. Four potential candidate genes associated with the OTL and OtL of tea plant were also identified, which will facilitate genetic research, germplasm conservation, and breeding.

Depicting the genetic and metabolic panorama of chemical diversity in the tea plant.

As a frequently consumed beverage worldwide, tea is rich in naturally important bioactive metabolites. Combining genetic, metabolomic and biochemical methodologies, here, we present a comprehensive study to dissect the chemical diversity in tea plant. A total of 2837 metabolites were identified at high-resolution with 1098 of them being structurally annotated and 63 of them were structurally identified. Metabolite-based genome-wide association mapping identified 6199 and 7823 metabolic quantitative trait loci (mQTL) for 971 and 1254 compounds in young leaves (YL) and the third leaves (TL), respectively. The major mQTL (i.e., P < 1.05 × 10-5, and phenotypic variation explained (PVE) > 25%) were further interrogated. Through extensive annotation of the tea metabolome as well as network-based analysis, this study broadens the understanding of tea metabolism and lays a solid foundation for revealing the natural variations in the chemical composition of the tea plant. Interestingly, we found that galloylations, rather than hydroxylations or glycosylations, were the largest class of conversions within the tea metabolome. The prevalence of galloylations in tea is unusual, as hydroxylations and glycosylations are typically the most prominent conversions of plant specialized metabolism. The biosynthetic pathway of flavonoids, which are one of the most featured metabolites in tea plant, was further refined with the identified metabolites. And we demonstrated the further mining and interpretation of our GWAS results by verifying two identified mQTL (including functional candidate genes CsUGTa, CsUGTb, and CsCCoAOMT) and completing the flavonoid biosynthetic pathway of the tea plant.

Plant-derived monoterpene S-linalool and ß-ocimene generated by CsLIS and CsOCS-SCZ are key chemical cues for attracting parasitoid wasps for suppressing Ectropis obliqua infestation in Camellia sinensis L.

Insect-induced plant volatile organic compounds (VOCs) may function as either direct defence molecules to deter insects or indirect defence signals to attract the natural enemies of the invading insects. Tea (Camellia sinensis L.), an important leaf-based beverage crop, is mainly infested by Ectropis obliqua which causes the most serious damage. Here, we report a mechanistic investigation of tea plant-derived VOCs in an indirect defence mechanism against E. obliqua. Parasitoid wasp Parapanteles hyposidrae, a natural enemy of E. obliqua, showed strong electrophysiological response and selection behaviour towards S-linalool and ß-ocimene, two monoterpenes with elevated emission from E. obliqua-damaged tea plants. Larvae frass of E. obliqua, which also released S-linalool and ß-ocimene, was found to attract both mated female or male Pa. hyposidrae according to gas chromatography-electroantennogram detection and Y-tube olfactometer assays. In a field setting, both S-linalool and ß-ocimene were effective in recruiting both female and male Pa. hyposidrae wasps. To understand the molecular mechanism of monoterpenes-mediated indirect defence in tea plants, two novel monoterpene synthase genes, CsLIS and CsOCS-SCZ, involved in the biosynthesis of S-linalool or ß-ocimene, respectively, were identified and biochemically characterised. When the expression of these two genes in tea plants was inhibited by antisense oligodeoxynucleotide, both volatile emission and attraction of wasps were reduced. Furthermore, gene expression analysis suggested that the expression of CsLIS and CsOCS-SCZ is regulated by the jasmonic acid signalling pathway in the tea plant.

CIPK11 phosphorylates GSTU23 to promote cold tolerance in Camellia sinensis.

Cold stress negatively impacts the growth, development, and quality of Camellia sinensis (Cs, tea) plants. CBL-interacting protein kinases (CIPK) comprise a pivotal protein family involved in plant development and response to multiple environmental stimuli. However, their roles and regulatory mechanisms in tea plants (Camellia sinensis (L.) O. Kuntze) remain unknown. Here we show that CsCBL-interacting protein kinase 11 (CsCIPK11), whose transcript abundance was significantly induced at low temperatures, interacts and phosphorylates tau class glutathione S-transferase 23 (CsGSTU23). CsGSTU23 was also a cold-inducible gene and has significantly higher transcript abundance in cold-resistant accessions than in cold-susceptible accessions. CsCIPK11 phosphorylated CsGSTU23 at Ser37, enhancing its stability and enzymatic activity. Overexpression of CsCIPK11 in Arabidopsis thaliana resulted in enhanced cold tolerance under freezing conditions, while transient knockdown of CsCIPK11 expression in tea plants had the opposite effect, resulting in decreased cold tolerance and suppression of the C-repeat-binding transcription factor (CBF) transcriptional pathway under freezing stress. Furthermore, the transient overexpression of CsGSTU23 in tea plants increased cold tolerance. These findings demonstrate that CsCIPK11 plays a central role in the signaling pathway to cold signals and modulates antioxidant capacity by phosphorylating CsGSTU23, leading to improved cold tolerance in tea plants.

Biocontrol potential of plant growth-promoting rhizobacteria against plant disease and insect pest.

Biological control is a promising approach to enhance pathogen and pest control to ensure high productivity in cash crop production. Therefore, PGPR biofertilizers are very suitable for application in the cultivation of tea plants (Camellia sinensis) and tobacco, but it is rarely reported so far. In this study, production of a consortium of three strains of PGPR were applied to tobacco and tea plants. The results demonstrated that plants treated with PGPR exhibited enhanced resistance against the bacterial pathogen Pseudomonas syringae (PstDC3000). The significant effect in improving the plant's ability to resist pathogen invasion was verified through measurements of oxygen activity, bacterial colony counts, and expression levels of resistance-related genes (NPR1, PR1, JAZ1, POD etc.). Moreover, the application of PGPR in the tea plantation showed significantly reduced population occurrences of tea green leafhoppers (Empoasca onukii Matsuda), tea thrips (Thysanoptera:Thripidae), Aleurocanthus spiniferus (Quaintanca) and alleviated anthracnose disease in tea seedlings. Therefore, PGPR biofertilizers may serve as a viable biological control method to improve tobacco and tea plant yield and quality. Our findings revealed part of the mechanism by which PGPR helped improve plant biostresses resistance, enabling better application in agricultural production.

Recent Progress Regarding Jasmonates in Tea Plants: Biosynthesis, Signaling, and Function in Stress Responses.

Tea plants have to adapt to frequently challenging environments due to their sessile lifestyle and perennial evergreen nature. Jasmonates regulate not only tea plants' responses to biotic stresses, including herbivore attack and pathogen infection, but also tolerance to abiotic stresses, such as extreme weather conditions and osmotic stress. In this review, we summarize recent progress about jasmonaic acid (JA) biosynthesis and signaling pathways, as well as the underlying mechanisms mediated by jasmontes in tea plants in responses to biotic stresses and abiotic stresses. This review provides a reference for future research on the JA signaling pathway in terms of its regulation against various stresses of tea plants. Due to the lack of a genetic transformation system, the JA pathway of tea plants is still in the preliminary stages. It is necessary to perform further efforts to identify new components involved in the JA regulatory pathway through the combination of genetic and biochemical methods.

Changes in the Antioxidant Potential of Camellia sinensis Cultures under the Influence of Phenolic Precursors.

The viability, productivity and survival of higher plants under the adverse factors influence are largely determined by the functional activity of the antioxidant system. The aim of our work was to investigate changes in formation of high-molecular (superoxide dismutase and peroxidase) and low-molecular (phenolics, including flavanols and proanthocyanidins) antioxidants in callus culture of Camellia sinensis under influence of phenolic precursors (L-phenylalanine-3 mM, trans-cinnamic acid-1 mM, naringenin-0.5 mM). According to the data obtained, the effect of precursors on tea callus cultures did not lead to significant increasing of superoxide dismutase and peroxidase activity in most cases. However, it led to the increased accumulation of the total phenolics content, as well as flavanols and proanthocyanidins contents. For C. sinensis callus cultures, the most promising regulator of phenolic compounds was L-phenylalanine, in the presence of which its content increased almost twice. Thus, the exogenous effect of various precursors is possible to use for the targeted regulation of certain phenolics classes accumulation in plant cells.

Genome-wide identification of the PYL gene family of tea plants (Camellia sinensis) revealed its expression profiles under different stress and tissues.

BACKGROUND: PYL (Pyrabactin resistance 1-like) protein is a receptor of abscisic acid (ABA), which plays an important role in ABA signaling and influences plant growth and development and stress response. However, studies on PYL gene family in tea plants have not been reported. RESULTS: In this study, we identified 20 PYL genes from the reference genome of tea plant ('Shuchazao'). Phylogeny analysis indicated that PYLs from tea and other plant species were clustered into seven groups. The promoter region of PYL genes contains a large number of cis-elements related to hormones and stresses. A large number of PYL genes responding to stress were found by analyzing the expression levels of abiotic stress and biotic stress transcriptome data. For example, CSS0047272.1 were up-regulated by drought stress, and CSS0027597.1 could respond to both anthracnose disease and geometrid feeding treatments. In addition, 10 PYL genes related to growth and development were verified by RT-qPCR and their tissue expression characteristics were revealed. CONCLUSIONS: Our results provided a comprehensive characteristic of the PYL gene family in tea plants and provided an important clue for further exploring its functions in the growth and development, and resistance to stress of tea plants.

Physiological and transcriptomic analyses reveal tea plant (Camellia sinensis L.) adapts to extreme freezing stress during winter by regulating cell wall structure.

Tea plants grown in high-latitude areas are often damaged by extreme freezing temperatures in winter, leading to huge economic losses. Here, the physiological and gene expression characteristics of two tea cultivars (Xinyang No. 10 (XY10), a freezing-tolerant cultivar and Fudingdabaicha (FDDB), a freezing-sensitive cultivar) during overwintering in northern China were studied to better understand the regulation mechanisms of tea plants in response to natural freezing stress. Samples were collected at a chill (D1), freezing (D2) and recovery (D3) temperature in winter. TEM analysis of integrated leaf ultrastructure at D2 revealed lower malondialdehyde and relative electrical conductivity in XY10 than in FDDB, with serious cell structure damage in the latter, indicating XY10 was more resistant to freezing stress. Differential gene expression analysis among the different samples over winter time highlighted the following gene functions in cell wall metabolism (CesAs, COBLs, XTHs, PGs, PMEs), transcription factors (ERF1B and MYC2), and signal transduction (CDPKs and CMLs). The expression pattern of cellulose and pectin-related genes suggested higher accumulation of cellulosic and pectic materials in the cell wall of XY10, agreeing with the results of cell wall and its components. These results indicated that under the regulation of cell wall genes, the freezing-resistant tea cultivar can better maintain a well-knit cell wall structure with sufficient substances to survive natural freezing damage. This study demonstrated the crucial role of cell wall in tea plant resistance to natural freezing stress and provided important candidate genes for breeding of freezing-resistant tea cultivars.

Comprehensive analysis of the laccase gene family in tea plant highlights its roles in development and stress responses.

BACKGROUND: Laccase (LAC) is the pivotal enzyme responsible for the polymerization of monolignols and stress responses in plants. However, the roles of LAC genes in plant development and tolerance to diverse stresses are still largely unknown, especially in tea plant (Camellia sinensis), one of the most economically important crops worldwide. RESULTS: In total, 51 CsLAC genes were identified, they were unevenly distributed on different chromosomes and classified into six groups based on phylogenetic analysis. The CsLAC gene family had diverse intron-exon patterns and a highly conserved motif distribution. Cis-acting elements in the promoter demonstrated that promoter regions of CsLACs encode various elements associated with light, phytohormones, development and stresses. Collinearity analysis identified some orthologous gene pairs in C. sinensis and many paralogous gene pairs among C. sinensis, Arabidopsis and Populus. Tissue-specific expression profiles revealed that the majority of CsLACs had high expression in roots and stems and some members had specific expression patterns in other tissues, and the expression patterns of six genes by qRT‒PCR were highly consistent with the transcriptome data. Most CsLACs showed significant variation in their expression level under abiotic (cold and drought) and biotic (insect and fungus) stresses via transcriptome data. Among them, CsLAC3 was localized in the plasma membrane and its expression level increased significantly at 13 d under gray blight treatment. We found that 12 CsLACs were predicted to be targets of cs-miR397a, and most CsLACs showed opposite expression patterns compared to cs-miR397a under gray blight infection. Additionally, 18 highly polymorphic SSR markers were developed, these markers can be widely used for diverse genetic studies of tea plants. CONCLUSIONS: This study provides a comprehensive understanding of the classification, evolution, structure, tissue-specific profiles, and (a)biotic stress responses of CsLAC genes. It also provides valuable genetic resources for functional characterization towards enhancing tea plant tolerance to multiple (a)biotic stresses.

Population structure analysis to explore genetic diversity and geographical distribution characteristics of wild tea plant in Guizhou Plateau.

BACKGROUND: Tea, the second largest consumer beverage in the world after water, is widely cultivated in tropical and subtropical areas. However, the effect of environmental factors on the distribution of wild tea plants is unclear. RESULTS: A total of 159 wild tea plants were collected from different altitudes and geological types of the Guizhou Plateau. Using the genotyping-by-sequencing method, 98,241 high-quality single nucleotide polymorphisms were identified. Genetic diversity, population structure analysis, principal component analysis, phylogenetic analysis, and linkage disequilibrium were performed. The genetic diversity of the wild tea plant population from the Silicate Rock Classes of Camellia gymnogyna was higher than that from the Carbonate Rock Classes of Camellia tachangensis. In addition, the genetic diversity of wild tea plants from the second altitude gradient was significantly higher than that of wild tea plants from the third and first altitude gradients. Two inferred pure groups (GP01 and GP02) and one inferred admixture group (GP03) were identified by population structure analysis and were verified by principal component and phylogenetic analyses. The highest differentiation coefficients were determined for GP01 vs. GP02, while the lowest differentiation coefficients were determined for GP01 vs. GP03. CONCLUSIONS: This study revealed the genetic diversity and geographical distribution characteristics of wild tea plants in the Guizhou Plateau. There are significant differences in genetic diversity and evolutionary direction between Camellia tachangensis with Carbonate Rock Classes at the first altitude gradient and Camellia gymnogyna with Silicate Rock Classes at the third altitude gradient. Geological environment, soil mineral element content, soil pH, and altitude markedly contributed to the genetic differentiation between Camellia tachangensis and Camellia gymnogyna.

Comprehensive analysis and expression profiles of the AP2/ERF gene family during spring bud break in tea plant (Camellia sinensis).

BACKGROUND: AP2/ERF transcription factors (AP2/ERFs) are important regulators of plant physiological and biochemical metabolism. Evidence suggests that AP2/ERFs may be involved in the regulation of bud break in woody perennials. Green tea is economically vital in China, and its production value is significantly affected by the time of spring bud break of tea plant. However, the relationship between AP2/ERFs in tea plant and spring bud break remains largely unknown. RESULTS: A total of 178 AP2/ERF genes (CsAP2/ERFs) were identified in the genome of tea plant. Based on the phylogenetic analysis, these genes could be classified into five subfamilies. The analysis of gene duplication events demonstrated that whole genome duplication (WGD) or segmental duplication was the primary way of CsAP2/ERFs amplification. According to the result of the Ka/Ks value calculation, purification selection dominated the evolution of CsAP2/ERFs. Furthermore, gene composition and structure analyses of CsAP2/ERFs indicated that different subfamilies contained a variety of gene structures and conserved motifs, potentially resulting in functional differences among five subfamilies. The promoters of CsAP2/ERFs also contained various signal-sensing elements, such as abscisic acid responsive elements, light responsive elements and low temperature responsive elements. The evidence presented here offers a theoretical foundation for the diverse functions of CsAP2/ERFs. Additionally, the expressions of CsAP2/ERFs during spring bud break of tea plant were analyzed by RNA-seq and grouped into clusters A-F according to their expression patterns. The gene expression changes in clusters A and B were more synchronized with the spring bud break of tea plant. Moreover, several potential correlation genes, such as D-type cyclin genes, were screened out through weighted correlation network analysis (WGCNA). Temperature and light treatment experiments individually identified nine candidate CsAP2/ERFs that may be related to the spring bud break of tea plant. CONCLUSIONS: This study provides new evidence for role of the CsAP2/ERFs in the spring bud break of tea plant, establishes a theoretical foundation for analyzing the molecular mechanism of the spring bud break of tea plant, and contributes to the improvement of tea cultivars.

Transcriptomic and metabolomic analyses reveals keys genes and metabolic pathways in tea (Camellia sinensis) against six-spotted spider mite (Eotetranychus Sexmaculatus).

BACKGROUND: Six-spotted spider mite (Eotetranychus sexmaculatus) is one of the most damaging pests of tea (Camellia sinensis). E. sexmaculatus causes great economic loss and affects tea quality adversely. In response to pests, such as spider mites, tea plants have evolved resistance mechanisms, such as expression of defense-related genes and defense-related metabolites. RESULTS: To evaluate the biochemical and molecular mechanisms of resistance in C. sinensis against spider mites, "Tianfu-5" (resistant to E. sexmaculatus) and "Fuding Dabai" (susceptible to E. sexmaculatus) were inoculated with spider mites. Transcriptomics and metabolomics based on RNA-Seq and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) technology were used to analyze changes in gene expression and metabolite content, respectively. RNA-Seq data analysis revealed that 246 to 3,986 differentially expressed genes (DEGs) were identified in multiple compared groups, and these DEGs were significantly enriched in various pathways, such as phenylpropanoid and flavonoid biosynthesis, plant-pathogen interactions, MAPK signaling, and plant hormone signaling. Additionally, the metabolome data detected 2,220 metabolites, with 194 to 260 differentially abundant metabolites (DAMs) identified in multiple compared groups, including phenylalanine, lignin, salicylic acid, and jasmonic acid. The combined analysis of RNA-Seq and metabolomic data indicated that phenylpropanoid and flavonoid biosynthesis, MAPK signaling, and Ca2+-mediated PR-1 signaling pathways may contribute to spider mite resistance. CONCLUSIONS: Our findings provide insights for identifying insect-induced genes and metabolites and form a basis for studies on mechanisms of host defense against spider mites in C. sinensis. The candidate genes and metabolites identified will be a valuable resource for tea breeding in response to biotic stress.

Genome-wide association study of leaf-related traits in tea plant in Guizhou based on genotyping-by-sequencing.

BACKGROUND: Studying the genetic characteristics of tea plant (Camellia spp.) leaf traits is essential for improving yield and quality through breeding and selection. Guizhou Plateau, an important part of the original center of tea plants, has rich genetic resources. However, few studies have explored the associations between tea plant leaf traits and single nucleotide polymorphism (SNP) markers in Guizhou. RESULTS: In this study, we used the genotyping-by-sequencing (GBS) method to identify 100,829 SNP markers from 338 accessions of tea germplasm in Guizhou Plateau, a region with rich genetic resources. We assessed population structure based on high-quality SNPs, constructed phylogenetic relationships, and performed genome-wide association studies (GWASs). Four inferred pure groups (G-I, G-II, G-III, and G-IV) and one inferred admixture group (G-V), were identified by a population structure analysis, and verified by principal component analyses and phylogenetic analyses. Through GWAS, we identified six candidate genes associated with four leaf traits, including mature leaf size, texture, color and shape. Specifically, two candidate genes, located on chromosomes 1 and 9, were significantly associated with mature leaf size, while two genes, located on chromosomes 8 and 11, were significantly associated with mature leaf texture. Additionally, two candidate genes, located on chromosomes 1 and 2 were identified as being associated with mature leaf color and mature leaf shape, respectively. We verified the expression level of two candidate genes was verified using reverse transcription quantitative polymerase chain reaction (RT-qPCR) and designed a derived cleaved amplified polymorphism (dCAPS) marker that co-segregated with mature leaf size, which could be used for marker-assisted selection (MAS) breeding in Camellia sinensis. CONCLUSIONS: In the present study, by using GWAS approaches with the 338 tea accessions population in Guizhou, we revealed a list of SNPs markers and candidate genes that were significantly associated with four leaf traits. This work provides theoretical and practical basis for the genetic breeding of related traits in tea plant leaves.

Regulation of biosynthesis of the main flavor-contributing metabolites in tea plant (Camellia sinensis): A review.

In the process of adapting to the environment, tea plants (Camellia sinensis) endow tea with unique flavor and health functions, which should be attributed to secondary metabolites, including catechins, L-theanine, caffeine and terpene volatiles. Since the content of these flavor-contributing metabolites are mainly determined by the growth of tea plant, it is very important to understand their alteration and regulation mechanisms. In the present work, we first summarize the distribution, change characteristics of the main flavor-contributing metabolites in different cultivars, organs and under environmental stresses of tea plant. Subsequently, we discuss the regulating mechanisms involved in the biosynthesis of these metabolites based on the existing evidence. Finally, we propose the remarks and perspectives on the future study relating flavor-contributing metabolites. This review would contribute to the acceleration of research on the characteristic secondary metabolites and the breeding programs in tea plants.

Multi-Omics Analysis Reveals Mechanisms of Strong Phosphorus Adaptation in Tea Plant Roots.

Low phosphorus (P) is a major limiting factor for plant growth in acid soils, which are preferred by tea plants. This study aims to investigate the unique mechanisms of tea plant roots adaptation to low-P conditions. Tea plant roots were harvested for multi-omics analysis after being treated with 0 µmol·L-1 P (0P) and 250 µmol·L-1 P (250P) for 30 days. Under 250P conditions, root elongation was significantly inhibited, and the density of lateral roots was dramatically increased. This suggests that 250P may inhibit the elongation of tea plant roots. Moreover, the P concentration in roots was about 4.58 times higher than that under 0P, indicating that 250P may cause P toxicity in tea plant roots. Contrary to common plants, the expression of CsPT1/2 in tea plant roots was significantly increased by four times at 250P, which indicated that tea plant roots suffering from P toxicity might be due to the excessive expression of phosphate uptake-responsible genes under 250P conditions. Additionally, 94.80% of P-containing metabolites accumulated due to 250P stimulation, most of which were energy-associated metabolites, including lipids, nucleotides, and sugars. Especially the ratio of AMP/ATP and the expression of energy sensor CsSnRKs were inhibited by P application. Therefore, under 250P conditions, P over-accumulation due to the excessive expression of CsPT1/2 may inhibit energy metabolism and thus the growth of tea plant roots.

Aluminum Supplementation Mediates the Changes in Tea Plant Growth and Metabolism in Response to Calcium Stress.

Tea plants are more sensitive to variations in calcium concentration compared to other plants, whereas a moderate aluminum concentration facilitates the growth and development of tea plants. Aluminum and calcium show a competitive interaction with respect to the uptake of elements, consequently exerting physiological effects on plants. To further explore these interactions, in this study, we used the solution culture method to treat tea plants with two calcium concentrations (0.8 mM and 5.6 mM) and three aluminum concentrations (0 mM, 0.4 mM, and 1 mM). We then determined the influence of the combined treatments on root growth and quality compound accumulation in the tissues by a combination of phenotype, gene expression, and metabolite analyses. Moderate aluminum supplementation (0.4 mM) alleviated the inhibition of root growth caused by high calcium stress. High calcium stress significantly inhibited the accumulation of most amino acids (e.g., Glutamic acid, Citulline, and Arginine) and organic acids (e.g., a-ketoglutaric acid) in the roots, stems, and leaves, whereas aluminum deficiency significantly increased most amino acids in the roots and leaves (except Serine, Alanine, and Phenylalanine in the roots and Ser in the leaves), with a more than two-fold increase in Arg and Lysine. High calcium stress also induced the accumulation of secondary metabolites such as epigallocatechin gallate and procyanidin in the roots, whereas aluminum supplementation significantly reduced the contents of flavonol glycosides such as quercetin, rutin, myricitrin, and kaempferitrin, as well as caffeine, regardless of calcium concentration. Aluminum supplementation reversed some of the changes in the contents of leaf metabolites induced by calcium stress (e.g., 4-dihydroquercetin, apigenin C-pentoside, phenethylamine, and caffeine). Overall, calcium stress caused severe growth inhibition and metabolic disorders in tea plants, which could be reversed by aluminum supplementation, particularly in maintaining the root tips and the accumulation of secondary metabolites. These results provide a theoretical basis for improving calcium-aluminum nutrient management to promote tea plant growth and quality.

CsCIPK11-Regulated Metalloprotease CsFtsH5 Mediates the Cold Response of Tea Plants.

Photosystem II repair in chloroplasts is a critical process involved in maintaining a plant's photosynthetic activity under cold stress. FtsH (filamentation temperature-sensitive H) is an essential metalloprotease that is required for chloroplast photosystem II repair. However, the role of FtsH in tea plants and its regulatory mechanism under cold stress remains elusive. In this study, we cloned a FtsH homolog gene in tea plants, named CsFtsH5, and found that CsFtsH5 was located in the chloroplast and cytomembrane. RT-qPCR showed that the expression of CsFtsH5 was increased with leaf maturity and was significantly induced by light and cold stress. Transient knockdown CsFtsH5 expression in tea leaves using antisense oligonucleotides resulted in hypersensitivity to cold stress, along with higher relative electrolyte leakage and lower Fv/Fm values. To investigate the molecular mechanism underlying CsFtsH5 involvement in the cold stress, we focused on the calcineurin B-like-interacting protein kinase 11 (CsCIPK11), which had a tissue expression pattern similar to that of CsFtsH5 and was also upregulated by light and cold stress. Yeast two-hybrid and dual luciferase (Luc) complementation assays revealed that CsFtsH5 interacted with CsCIPK11. Furthermore, the Dual-Luc assay showed that CsCIPK11-CsFtsH5 interaction might enhance CsFtsH5 stability. Altogether, our study demonstrates that CsFtsH5 is associated with CsCIPK11 and plays a positive role in maintaining the photosynthetic activity of tea plants in response to low temperatures.