Abstract：The root of Sophora flavescens is rich in flavonoid secondary metabolites. Chalcone synthase (CHS) is the first rate-limiting enzyme in the flavonoid biosynthesis pathway, and its characteristics and activity directly affect the biosynthesis and accumulation of secondary metabolites. To analyze the phylogenetic relationship of CHS genes in S. flavescens and their expression profiles, and explore their roles in flavonoid biosynthesis, this study identified 21 CHS family members in S. flavescens using bioinformatics methods. Gene duplication contributed greatly to this family expansion, and the duplicate SfCHS genes underwent strong purifying selection. Then, these CHS members were subdivided into 3 groups (GroupⅠ,ⅡandⅢ) based on phylogenetic analysis. CHS family were not conserved during the evolution, but these members belonging to the same group were conserved in gene structure and motif distribution. The SfCHS04 promoter region contained the highest number of cis-elements for stress resistance. Moreover, based on the expression level of SfCHS genes among different tissues (root, stem, leaf, and flower), majority genes were specifically expressed in flowers. Based on the co-expression network analysis for transcriptome and metabolome data of the pod development, SfCHS04 and SfCHS20 were located in the center of the network, and the main regulatory transcription factors included members of ERF, MYB, WRKY, and NAC families, which played important roles in the development of S. flavescens pods and the accumulation of flavonoids. Finally, the quantitative of SfCHS genes using qPCR technology, which revealed that distinct SfCHS genes showed significant variations in expression levels during root development and among multiple tissues at different growth stages. In summary, the results not only clarify the phylogeny and expression profile of SfCHS genes, but also provide reference value for understanding the molecular mechanism of flavonoid biosynthesis and improving the medicinal value of S. flavescens through molecular breeding methods.
刘阿克, 卢俊洁, 刘林涛, 牛天增, 刘江天, 宋慧芳, 王茜. 苦参查尔酮合酶(CHS)基因家族的鉴定与表达谱分析[J]. 农业生物技术学报, 2025, 33(5): 1031-1043.
LIU A-Ke, LU Jun-Jie, LIU Lin-Tao, NIU Tian-Zeng, LIU Jiang-Tian, SONG Hui-Fang, WANG Xi. Identification and Expression Profile Analysis of Chalcone Synthase (CHS) Gene Family in Sophora flavescens. 农业生物技术学报, 2025, 33(5): 1031-1043.
[1] 梅志栋, 张贺, 刘晓妹, 等. 2015. 杧果查尔酮合成酶基因(CHS1)的克隆与表达分析[J]. 果树学报, 32(6): 1077-1084+1315.(Mei Z, Zhang H, Liu X, et al. 2015. Cloning and expression analysis of chalcone synthase gene [2] Abe I, Morita H.2010. Structure and function of the chalcone synthase superfamily of plant typeⅢpolyketide synthases[J]. Natural Product Reports, 27(6): 809-838. [3] Ahmad S, Jeridi M, Siddiqui S, et al.2023. Genome-wide identification, characterization, and expression analysis of the chalcone synthase gene family in Oryza sativa under abiotic stresses[J]. Plant Stress, 9: 100201. [4] Anguraj Vadivel A K, Krysiak K, Tian G, et al.2018. Genome-wide identification and localization of chalcone synthase family in soybean (Glycine max [L]Merr)[J]. BMC Plant Biology, 18(1): 325. [5] Austin M B, Noel J P.2003. The chalcone synthase superfamily of typeⅢpolyketide synthases[J]. Natural Product Reports, 20(1): 79-110. [6] Baozhu L, Ruonan F, Yanting F, et al.2022. The flavonoid biosynthesis regulator PFG3 confers drought stress tolerance in plants by promoting flavonoid accumulation[J]. Environmental and Experimental Botany, 196: 104792. [7] Baudry A, Caboche M, Lepiniec L.2006. TT8 controls its own expression in a feedback regulation involving TTG1 and homologous MYB and bHLH factors, allowing a strong and cell‐specific accumulation of flavonoids in Arabidopsis thaliana[J]. Plant Journal, 46(5): 768-779. [8] Baudry A, Heim MA, Dubreucq B, et al.2004. TT2, TT8, and TTG1 synergistically specify the expression of BANYULS and proanthocyanidin biosynthesis in Arabidopsis thaliana[J]. Plant Journal, 39(3): 366-380. [9] Chen S, Pan X, Li Y, et al.2016. Identification and characterization of chalcone synthase gene family members in Nicotiana tabacum[J]. Journal of Plant Growth Regulation, 36(2): 374-384. [10] Cheng W, Liu D, Guo M, et al.2022. Sophoraflavanone G suppresses the progression of triple‐negative breast cancer via the inactivation of EGFR-PI3K-AKT signaling[J]. Drug Development Research, 83(5): 1138-1151. [11] Daryanavard H, Postiglione A E, Mühlemann J K, et al.2023. Flavonols modulate plant development, signaling, and stress responses[J]. Current Opinion in Plant Biology, 72: 102350. [12] Deng X, Bashandy H, Ainasoja M, et al.2013. Functional diversification of duplicated chalcone synthase genes in anthocyanin biosynthesis of Gerbera hybrida[J]. New Phytologist, 201(4): 1469-1483. [13] Durbin M L, Mccaig B, Clegg M T2000. Molecular evolution of the chalcone synthase multigene family in the morning glory genome[J]. Plant Molecular Biology, 42(1): 79-92. [14] El-Gebali S, Mistry J, Bateman A, et al.2019. The Pfam protein families database in 2019[J]. Nucleic Acids Research, 47(D1): D427-d432. [15] Gonzalez A, Zhao M, Leavitt J M, et al.2007. Regulation of the anthocyanin biosynthetic pathway by the TTG1/bHLH/Myb transcriptional complex in Arabidopsis seedlings[J]. The Plant Journal, 53(5): 814-827. [16] Hu T, Gao Z Q, Hou J M, et al.2020. Identification of biosynthetic pathways involved in flavonoid production in licorice by RNA-seq based transcriptome analysis[J]. Plant Growth Regulation, 92(1): 15-28. [17] Huang X C, Tang H, Wei X, et al.2024. The gradual establishment of complex coumarin biosynthetic pathway in Apiaceae[J]. Nature Communications, 15(1): 6864. [18] Langfelder P, Horvath S.2008. WGCNA: An R package for weighted correlation network analysis[J]. BMC Bioinformatics, 9: 559. [19] Lescot M.2002. PlantCARE, a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences[J]. Nucleic Acids Research, 30(1): 325-327. [20] Lin Z, Qu X.2022. Emerging diversity in polyketide synthase[J]. Tetrahedron Letters, 110: 154183. [21] Liu S, Meng Z, Zhang H, et al.2022. Identification and characterization of thirteen gene families involved in flavonoid biosynthesis in Ginkgo biloba[J]. Industrial Crops and Products, 188: 115576. [22] Noel J P, Ferrer J L, Jez J M, et al.1999. Structure of chalcone synthase and the molecular basis of plant polyketide biosynthesis[J]. Nature Structural Biology, 6(8): 775-784. [23] Otasek D, Morris J H, Bouças J, et al.2019. Cytoscape Automation: Empowering workflow-based network analysis[J]. Genome Biology, 20(1): 185. [24] Pandith S A, Ramazan S, Khan M I, et al.2019. Chalcone synthases (CHSs): The symbolic type Ⅲ polyketide synthases[J]. Planta, 251(1): 15. [25] Prakash A, Jeffryes M, Bateman A, et al.2017. The HMMER web server for protein sequence similarity search[J]. Current Protocols in Bioinformatics, 60(1): 3.15.11-13.15.23. [26] Qiao X, Li Q, Yin H, et al.2019. Gene duplication and evolution in recurring polyploidization-diploidization cycles in plants[J]. Genome Biology, 20(1): 38. [27] Qu Z, Wang W, Adelson D L.2023. Chromosomal level genome assembly of medicinal plant Sophora flavescens[J]. Scientific Data, 10(1): 572. [28] Schröder J.2000. Evolution of Metabolic Pathways[M]. Elsevier, Kidlington, Britain. pp. 55-89. [29] Shen B.2003. Polyketide biosynthesis beyond the typeⅠ,Ⅱand Ⅲ polyketide synthase paradigms[J]. Current Opinion in Chemical Biology, 7(2): 285-295. [30] Shen N, Wang T, Gan Q, et al.2022. Plant flavonoids: Classification, distribution, biosynthesis, and antioxidant activity[J]. Food Chemistry, 383: 132531. [31] Sun P, Zhao W, Wang Q, et al.2022. Chemical diversity, biological activities and traditional uses of and important Chinese herb Sophora[J]. Phytomedicine, 100: 154054. [32] Wang G, Yang B, Wu J, et al.2018a. Identification of genes involved in flavonoid biosynthesis of chinese narcissus (Narcissus tazetta L. var. chinensis)[J]. Plant Molecular Biology Reporter, 36(5-6): 812-821. [33] Wang Z, Yu Q, Shen W, et al.2018b. Functional study of CHS gene family members in citrus revealed a novel CHS gene affecting the production of flavonoids[J]. BMC Plant Biology, 18(1): 189. [34] Yang T, Zhang T, Li Y, et al.2023. Genome-wide identification and expression analysis of the chalcone synthase (CHS) gene family in Dendrobium catenatum[J]. Agronomy, 13(6): 1488. [35] Yang Z, Nielsen R.2000. Estimating synonymous and nonsynonymous nubstitution rates under realistic evolutionary models[J]. Molecular Biology and Evolution, 17(1): 32-43. [36] Yin X, Yang D, Liu Y, et al.2023. Sophora moorcroftiana genome analysis suggests association between sucrose metabolism and drought adaptation[J]. Plant Physiology, 191(2): 844-848. [37] Zhang F S, Wang Q Y, Pu Y J, et al.2017. Identification of genes involved in flavonoid biosynthesis in Sophora japonica through transcriptome sequencing[J]. Chemistry & Biodiversity, 14(12): e1700369. [38] Zhang Z.2022. KaKs_Calculator 3.0: Calculating selective pressure on coding and non-coding sequences[J]. Genomics, Proteomics and Bioinformatics, 20(3): 536-540. [39] Zhong Y, Xie Y, Zhang D, et al.2024. Integrated metabolomic and transcriptomic analysis of metabolic diversity and biosynthesis of glucosinolates and flavonoids in various cultivars of radish microgreens[J]. Food Bioscience, 59: 104055. [40] Zhu L, Ding Y, Wang S, et al.2022. Genome-wide identification, characterization, and expression analysis of CHS gene family members in Chrysanthemum nankingense[J]. Genes, 13(11): 2145.
