Duplicated chalcone synthase(CHS)genes modulate flavonoid production in tea plants in response to light stress

In tea plants,the abundant flavonoid compounds are responsible for the health benefits for the human body and define the astringent flavor profile.While the downstream mechanisms of flavonoid biosynthesis have been extensively studied,the role of chalcone synthase(CHS)in this secondary metabolic process in tea plants remains less clear.In this study,we compared the evolutionary profile of the flavonoid metabolism pathway and discovered that gene duplication of CHS occurred in tea plants.We identified three CsCHS genes,along with a CsCHS-like gene,as potential candidates for further functional investigation.Unlike the CsCHS-like gene,the CsCHS genes effectively restored flavonoid production in Arabidopsis chs-mutants.Additionally,CsCHS transgenic tobacco plants exhibited higher flavonoid compound accumulation compared to their wild-type counterparts.Most notably,our examination of promoter and gene expression levels for the selected CHS genes revealed distinct responses to UV-B stress in tea plants.Our findings suggest that environmental factors such as UV-B exposure could have been the key drivers behind the gene duplication events in CHS.

Camellia sinensis CsMYB4a participates in regulation of stamen growth by interaction with auxin signaling transduction repressor CsAUX/IAA4

Subgroup 4(Sg4)members of the R2R3-MYB are generally known as negative regulators of the phenylpropanoid pathway in plants.Our previous research showed that a R2R3-MYB Sg4 member from Camellia sinensis(CsMYB4a)inhibits expression of some genes in the phenylpropanoid pathway,but its physiological function in the tea plant remained unknown.Here,CsMYB4a was found to be highly expressed in anther and filaments,and participated in regulating filament growth.Transcriptome analysis and exogenous auxin treatment showed that the target of CsMYB4a might be the auxin signal pathway.Auxin/indole-3-acetic acid 4(AUX/IAA4),a repressor in auxin signal transduction,was detected from a yeast two-hybrid screen using CsMYB4a as bait.Gene silencing assays showed that both CsIAA4 and CsMYB4a regulate filament growth.Tobacco plants overexpressing CsIAA4 were insensitive to exogenous a-NAA,consistent with overexpression of CsMYB4a.Protein-protein interaction experiments revealed that CsMYB4a interacts with N-terminal of CsIAA4 to prevent CsIAA4 degradation.Knock out of the endogenous NtIAA4 gene,a CsIAA4 homolog,in tobacco alleviated filament growth inhibition and a-NAA insensitivity in plants overexpressing CsMYB4a.All results strongly suggest that CsMYB4a works synergistically with CsIAA4 and participates in regulation of the auxin pathway in stamen.

Al-induced CsUGT84J2 enhances flavonol and auxin accumulation to promote root growth in tea plants

Although Al is not necessary or even toxic to most plants,it is beneficial for the growth of tea plants.However,the mechanism through which Al promotes root growth in tea plants remains unclear.In the present study,we found that flavonol glycoside levels in tea roots increased following Al treatment,and the Al-induced UDP glycosyltransferase CsUGT84J2 was involved in this mechanism.Enzyme activity assays revealed that rCsUGT84J2 exhibited catalytic activity on multiple types of substrates,including phenolic acids,flavonols,and auxins in vitro.Furthermore,metabolic analysis with UPLC-QqQ-MS/MS revealed significantly increased flavonol and auxin glycoside accumulation in CsUGT84J2-overexpressing Arabidopsis thaliana.In addition,the expression of genes involved in the flavonol pathway as well as in the auxin metabolism,transport,and signaling pathways was remarkably enhanced.Additionally,lateral root growth and exogenous Al stress tolerance were significantly improved in transgenic A.thaliana.Moreover,gene expression and metabolic accumulation related to phenolic acids,flavonols,and auxin were upregulated in CsUGT84J2-overexpressing tea plants but downregulated in CsUGT84J2-silenced tea plants.In conclusion,Al treatment induced CsUGT84J2 expression,mediated flavonol and auxin glycosylation,and regulated endogenous auxin homeostasis in tea roots,thereby promoting the growth of tea plants.Our findings lay the foundation for studying the precise mechanisms through which Al promotes the growth of tea plants.

Functional diversity of subgroup 5 R2R3-MYBs promoting proanthocyanidin biosynthesis and their key residues and motifs in tea plant

The tea plant(Camellia sinensis)is rich in polyphenolic compounds.Particularly,flavan-3-ols and proanthocyanidins(PAs)are essential for the flavor and disease-resistance property of tea leaves.The fifth subgroup of R2R3-MYB transcription factors comprises the primary activators of PA biosynthesis.This study showed that subgroup 5 R2R3-MYBs in tea plants contained at least nine genes belonging to the TT2,MYB5,and MYBPA types.Tannin-rich plants showed an expansion in the number of subgroup 5 R2R3-MYB genes compared with other dicotyledonous and monocot plants.The MYBPA-type genes of tea plant were slightly expanded.qRT–PCR analysis and GUS staining analysis of promoter activity under a series of treatments revealed the differential responses of CsMYB5s to biotic and abiotic stresses.In particular,CsMYB5a,CsMYB5b,and CsMYB5e responded to high-intensity light,high temperature,MeJA,and mechanical wounding,whereas CsMYB5f and CsMYB5g were only induced by wounding.Three genetic transformation systems(C.sinensis,Nicotiana tabacum,and Arabidopsis thaliana)were used to verify the biological function of CsMYB5s.The results show that CsMYB5a,CsMYB5b,and CsMYB5e could promote the gene expression of CsLAR and CsANR.However,CsMYB5f and CsMYB5g could only upregulate the gene expression of CsLAR but not CsANR.A series of site-directed mutation and domain-swapping experiments were used to verify functional domains and key amino acids of CsMYB5s responsible for the regulation of PA biosynthesis.This study aimed to provide insight into the induced expression and functional diversity model of PA biosynthesis regulation in tea plants.

Roles of YABBY tanscription factors in the regulation of leaf development and abiotic stress responses in Camellia sinensis

The formation of leaf polarity directly determines the development of leaf spreading and type,and affects the physiological functions of photosynthesis,transpiration and stress resistance of plants.Many famous Chinese teas are highly dependent on the morphological characteristics and size of the tea leaves.To date,there have been few reports on the establishment of tea leaf polarity and leaf type development.The plant-specific transcription factor YABBY family gene has been reported to be involved in the formation of leaf polarity and growth.Herein we describe the isolation and functional characterization of transcription factor YABBY genes of the tea plant(Camellia sinensis).Six YABBY transcription factors were isolated from tea plants and classified into five clusters in the protein phylogenetic tree.Among them,CsFILa and CsFILb were highly expressed in buds and mature petals,and their expression decreased with the stage of leaf development.The Arabidopsis transgenic lines of CsFILa and CsFILb showed abaxial curled and long-narrow leaves.The above results indicated that CsFILa and CsFILb regulated the leaf growth of tea plants.The growth and expansion rate of leaf on the picked shoots decreased significantly after being plucked from tea plants,especially those suffering from drought stress.In this process,CsFILa expression levels decreased 60%−80%compared to the control.These data indicated YABBY genes play important roles in the leaf development of Camellia sinensis.

Removal of the C4-domain preserves the drought tolerance enhanced by CsMYB4a and eliminates the negative impact of this transcription factor on plant growth

The MYB4 transcription factor family regulates plant traits.However,their overexpression often results in undesirable side effects like growth reduction.We have reported a green tea(Camellia sinensis)MYB4 transcription factor(CsMYB4)that represses the phenylpropanoid and shikimate pathways and stunts plant growth and development.In the current study,we observed that in CsMYB4a transgenic tobacco(Nicotiana tabacum)plants,primary metabolism was altered,including sugar and amino acid metabolism,which demonstrated a pleiotropic regulation by CsMYB4a.The CsMYB4a transgenic tobacco plants had improved drought tolerance,which correlated to alterations in carbohydrate metabolism and an increase in proline content,as revealed by metabolic profiling and transcriptomic analysis.To mitigate the undesirable repressive side effects on plant traits,including dwarfism,shrunken leaves,and shorter roots of CsMYB4a transgenic plants,we deleted the C4 domain of CsMYB4a to obtain a CsMYB4a-DC4 variant and then overexpressed it in transgenic plants(CsMYB4a-DC4).These CsMYB4a-DC4 plants displayed a normal growth and had improved drought tolerance.Metabolite analysis demonstrated that the contents of carbohydrates and proline were increased in these transgenic plants.Our findings suggest that an approriate modification of TFs can generate novel crop traits,thus providing potential agricultural benefits and expanding its application to various crops.

Publisher Correction:Removal of the C4-domain preserves the drought tolerance enhanced by CsMYB4a and eliminates the negative impact of this transcription factor on plant growth

Publisher Correction:aBIOTECH https://doi.org/10.1007/s42994-024-00149-5 The original article has been updated to correct two errors introduced during production:In this article Xiu Li the same as Mingzhuo Li and Guoliang Ma should have been denoted as equally contributing authors.