[1]

Chia TY, Gan CY, Murugaiyah V, Hashmi SF, Fatima T, et al. 2022. A narrative review on the phytochemistry, pharmacology and therapeutic potentials of Clinacanthus nutans (Burm. f. ) lindau leaves as an alternative source of future medicine. Molecules 27:139

doi: 10.3390/molecules27010139
[2]

Khoo LW, Mediani A, Zolkeflee NKZ, Leong SW, Ismail IS, et al. 2015. Phytochemical diversity of Clinacanthus nutans extracts and their bioactivity correlations elucidated by NMR based metabolomics. Phytochemistry Letters 14:123−33

doi: 10.1016/j.phytol.2015.09.015
[3]

Chiangchin S, Thongyim S, Pandith H, Kaewkod T, Tragoolpua Y, et al. 2023. Clinacanthus nutans genetic diversity and its association with anti-apoptotic, antioxidant, and anti-bacterial activities. Scientific Reports 13:19566

doi: 10.1038/s41598-023-46105-z
[4]

Fong SY, Piva T, Dekiwadia C, Urban S, Huynh T. 2016. Comparison of cytotoxicity between extracts of Clinacanthus nutans (Burm. f.) Lindau leaves from different locations and the induction of apoptosis by the crude methanol leaf extract in D24 human melanoma cells. BMC Complementary and Alternative Medicine 16:368

doi: 10.1186/s12906-016-1348-x
[5]

Bechtold U, Field B. 2018. Molecular mechanisms controlling plant growth during abiotic stress. Journal of Experimental Botany 69(11):2753−58

doi: 10.1093/jxb/ery157
[6]

Khan V, Jha A, Princi, Seth T, Iqbal N, et al. 2024. Exploring the role of jasmonic acid in boosting the production of secondary metabolites in medicinal plants: Pathway for future research. Industrial Crops and Products 220:119227

doi: 10.1016/j.indcrop.2024.119227
[7]

Zheng H, Fu X, Shao J, Tang Y, Yu M, et al. 2023. Transcriptional regulatory network of high-value active ingredients in medicinal plants. Trends in Plant Science 28:429−46

doi: 10.1016/j.tplants.2022.12.007
[8]

Atchley WR, Zhao J. 2007. Molecular architecture of the DNA-binding region and its relationship to classification of basic helix-loop-helix proteins. Molecular Biology and Evolution 24:192−202

doi: 10.1093/molbev/msl143
[9]

Ledent V, Vervoort M. 2001. The basic helix-loop-helix protein family: comparative genomics and phylogenetic analysis. Genome Research 11:754−70

doi: 10.1101/gr.177001
[10]

Sun X, Wang Y, Sui N. 2018. Transcriptional regulation of bHLH during plant response to stress. Biochemical and Biophysical Research Communications 503:397−401

doi: 10.1016/j.bbrc.2018.07.123
[11]

Klein ES, Simmons DM, Swanson LW, Rosenfeld MG. 1993. Tissue-specific RNA splicing generates an ankyrin-like domain that affects the dimerization and DNA-binding properties of a bHLH protein. Genes & Development 7:55−71

doi: 10.1101/gad.7.1.55
[12]

Wang K, Liu H, Mei Q, Yang J, Ma F, et al. 2023. Characteristics of bHLH transcription factors and their roles in the abiotic stress responses of horticultural crops. Scientia Horticulturae 310:111710

doi: 10.1016/j.scienta.2022.111710
[13]

Chinnusamy V, Ohta M, Kanrar S, Lee BH, Hong X, et al. 2003. ICE1: a regulator of cold-induced transcriptome and freezing tolerance in Arabidopsis. Genes & Development 17:1043−54

doi: 10.1101/gad.1077503
[14]

Wang F, Zhu H, Chen D, Li Z, Peng R, et al. 2016. A grape bHLH transcription factor gene, VvbHLH1, increases the accumulation of flavonoids and enhances salt and drought tolerance in transgenic Arabidopsis thaliana. Plant Cell, Tissue and Organ Culture (PCTOC) 125:387−98

doi: 10.1007/s11240-016-0953-1
[15]

Ji Y, Xiao J, Shen Y, Ma D, Li Z, et al. 2014. Cloning and characterization of AabHLH1, a bHLH transcription factor that positively regulates artemisinin biosynthesis in Artemisia annuac. Plant & Cell Physiology 55:1592−604

doi: 10.1093/pcp/pcu090
[16]

Yuan M, Shu G, Zhou J, He P, Xiang L, et al. 2023. AabHLH113 integrates jasmonic acid and abscisic acid signaling to positively regulate artemisinin biosynthesis in Artemisia annua. New Phytologist 237:885−99

doi: 10.1111/nph.18567
[17]

Patra B, Pattanaik S, Schluttenhofer C, Yuan L. 2018. A network of jasmonate-responsive bHLH factors modulate monoterpenoid indole alkaloid biosynthesis in Catharanthus roseus. New Phytologist 217:1566−81

doi: 10.1111/nph.14910
[18]

Xu Y, Zhang H, Zhong Y, Jiang N, Zhong X, et al. 2022. Comparative genomics analysis of bHLH genes in cucurbits identifies a novel gene regulating cucurbitacin biosynthesis. Horticulture Research 9:uhac038

doi: 10.1093/hr/uhac038
[19]

Zhang X, Luo H, Xu Z, Zhu Y, Ji A, et al. 2015. Genome-wide characterisation and analysis of bHLH transcription factors related to tanshinone biosynthesis in Salvia miltiorrhiza. Scientific Reports 5:11244

doi: 10.1038/srep11244
[20]

Xu J, Xu H, Zhao H, Liu H, Xu L, et al. 2022. Genome-wide investigation of bHLH genes and expression analysis under salt and hormonal treatments in Andrographis paniculata. Industrial Crops and Products 183:114928

doi: 10.1016/j.indcrop.2022.114928
[21]

Qin Y, Li J, Chen J, Yao S, Li L, et al. 2024. Genome-wide characterization of the bHLH gene family in Gynostemma pentaphyllum reveals its potential role in the regulation of gypenoside biosynthesis. BMC Plant Biology 24:205

doi: 10.1186/s12870-024-04879-y
[22]

Wei K, Chen H. 2018. Comparative functional genomics analysis of bHLH gene family in rice, maize and wheat. BMC Plant Biology 18:309

doi: 10.1186/s12870-018-1529-5
[23]

Mistry J, Chuguransky S, Williams L, Qureshi M, Salazar GA, et al. 2021. Pfam: the protein families database in 2021. Nucleic Acids Research 49:D412−D419

doi: 10.1093/nar/gkaa913
[24]

Potter SC, Luciani A, Eddy SR, Park Y, Lopez R, et al. 2018. HMMER web server: 2018 update. Nucleic Acids Research 46:W200−W204

doi: 10.1093/nar/gky448
[25]

McGinnis S, Madden TL. 2004. BLAST: at the core of a powerful and diverse set of sequence analysis tools. Nucleic Acids Research 32:W20−W25

doi: 10.1093/nar/gkh435
[26]

Marchler-Bauer A, Derbyshire MK, Gonzales NR, Lu S, Chitsaz F, et al. 2015. CDD: NCBI's conserved domain database. Nucleic Acids Research 43:D222−D226

doi: 10.1093/nar/gku1221
[27]

Letunic I, Khedkar S, Bork P. 2021. SMART: recent updates, new developments and status in 2020. Nucleic Acids Research 49:D458−D460

doi: 10.1093/nar/gkaa937
[28]

Osorio D, Rondón-Villarreal P, Torres R. 2015. Peptides: a package for data mining of antimicrobial peptides. The R Journal 7(1):4−14

doi: 10.32614/rj-2015-001
[29]

Pires N, Dolan L. 2010. Origin and diversification of basic-helix-loop-helix proteins in plants. Molecular Biology and Evolution 27:862−74

doi: 10.1093/molbev/msp288
[30]

Edgar RC. 2004. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Research 32:1792−97

doi: 10.1093/nar/gkh340
[31]

Minh BQ, Schmidt HA, Chernomor O, Schrempf D, Woodhams MD, et al. 2020. IQ-TREE 2: new models and efficient methods for phylogenetic inference in the genomic era. Molecular Biology and Evolution 37:1530−34

doi: 10.1093/molbev/msaa015
[32]

Subramanian B, Gao S, Lercher MJ, Hu S, Chen WH. 2019. Evolview v3: a webserver for visualization, annotation, and management of phylogenetic trees. Nucleic Acids Research 47:W270−W275

doi: 10.1093/nar/gkz357
[33]

Bailey TL, Johnson J, Grant CE, Noble WS. 2015. The MEME suite. Nucleic Acids Research 43:W39−W49

doi: 10.1093/nar/gkv416
[34]

Shen W, Sipos B, Zhao L. 2024. SeqKit2: a Swiss army knife for sequence and alignment processing. iMeta 3:e191

doi: 10.1002/imt2.191
[35]

Lescot M, Déhais P, Thijs G, Marchal K, Moreau Y, et al. 2002. PlantCARE, a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences. Nucleic Acids Research 30:325−27

doi: 10.1093/nar/30.1.325
[36]

Horton NJ, Kleinman K. 2015. Using R and RStudio for data management, statistical analysis, and graphics. 2nd Edition. New York: Chapman & Hall/CRC Press. doi: 10.1201/b18151

[37]

Wang Y, Tang H, DeBarry JD, Tan X, Li J, et al. 2012. MCScanX: a toolkit for detection and evolutionary analysis of gene synteny and collinearity. Nucleic Acids Research 40:e49

doi: 10.1093/nar/gkr1293
[38]

Chen C, Wu Y, Li J, Wang X, Zeng Z, et al. 2023. TBtools-II: a 'one for all, all for one' bioinformatics platform for biological big-data mining. Molecular Plant 16:1733−42

doi: 10.1016/j.molp.2023.09.010
[39]

Zhang Z, Li J, Zhao XQ, Wang J, Wong GK, et al. 2006. KaKs_Calculator: calculating Ka and Ks through model selection and model averaging. Genomics, Proteomics & Bioinformatics 4:259−63

doi: 10.1016/S1672-0229(07)60007-2
[40]

Szklarczyk D, Franceschini A, Wyder S, Forslund K, Heller D, et al. 2015. STRING v10: protein–protein interaction networks, integrated over the tree of life. Nucleic Acids Research 43:D447−D452

doi: 10.1093/nar/gku1003
[41]

Berman HM, Westbrook J, Feng Z, Gilliland G, Bhat TN, et al. 2000. The Protein Data Bank. Nucleic Acids Research 28:235−42

doi: 10.1093/nar/28.1.235
[42]

Høie MH, Kiehl EN, Petersen B, Nielsen M, Winther O, et al. 2022. NetSurfP-3.0: accurate and fast prediction of protein structural features by protein language models and deep learning. Nucleic Acids Research 50:W510−W515

doi: 10.1093/nar/gkac439
[43]

Waterhouse A, Bertoni M, Bienert S, Studer G, Tauriello G, et al. 2018. SWISS-MODEL: homology modelling of protein structures and complexes. Nucleic Acids Research 46:W296−W303

doi: 10.1093/nar/gky427
[44]

DeLano WL. 2002. PyMOL: an open-source molecular graphics tool. CCP4 Newsletter on Protein Crystallography 40. pp. 82−92 https://legacy.ccp4.ac.uk/newsletters/newsletter40/11_pymol.pdf

[45]

Chen S, Zhou Y, Chen Y, Gu J. 2018. fastp: an ultra-fast all-in-one FASTQ preprocessor. Bioinformatics 34:i884−i890

doi: 10.1093/bioinformatics/bty560
[46]

Kim D, Paggi JM, Park C, Bennett C, Salzberg SL. 2019. Graph-based genome alignment and genotyping with HISAT2 and HISAT-genotype. Nature Biotechnology 37:907−15

doi: 10.1038/s41587-019-0201-4
[47]

Liao Y, Smyth GK, Shi W. 2013. The Subread aligner: fast, accurate and scalable read mapping by seed-and-vote. Nucleic Acids Research 41:e108

doi: 10.1093/nar/gkt214
[48]

An C, Lu L, Yao Y, Liu R, Cheng Y, et al. 2025. Selection and validation of reference genes in clinacanthus nutans under abiotic stresses, MeJA treatment, and in different tissues. International Journal of Molecular Sciences 26:2483

doi: 10.3390/ijms26062483
[49]

De Masi F, Grove CA, Vedenko A, Alibés A, Gisselbrecht SS, et al. 2011. Using a structural and logics systems approach to infer bHLH-DNA binding specificity determinants. Nucleic Acids Research 39:4553−63

doi: 10.1093/nar/gkr070
[50]

Kato M, Hata N, Banerjee N, Futcher B, Zhang MQ. 2004. Identifying combinatorial regulation of transcription factors and binding motifs. Genome Biology 5:R56

doi: 10.1186/gb-2004-5-8-r56
[51]

Xu G, Guo C, Shan H, Kong H. 2012. Divergence of duplicate genes in exon-intron structure. Proceedings of the National Academy of Sciences of the United States of America 109:1187−92

doi: 10.1073/pnas.1109047109
[52]

Zhang W, Zhang J, Fan Y, Dong J, Gao P, et al. 2024. RNA sequencing analysis reveals PgbHLH28 as the key regulator in response to methyl jasmonate-induced saponin accumulation in Platycodon grandiflorus. Horticulture Research 11:uhae058

doi: 10.1093/hr/uhae058
[53]

Goossens J, Mertens J, Goossens A. 2017. Role and functioning of bHLH transcription factors in jasmonate signalling. Journal of Experimental Botany 68:1333−47

doi: 10.1093/jxb/erw440
[54]

Zhou Z, Wu M, Sun B, Li J, Li J, et al. 2024. Identification of transcription factor genes responsive to MeJA and characterization of a LaMYC2 transcription factor positively regulates lycorine biosynthesis in Lycoris aurea. Journal of Plant Physiology 296:154218

doi: 10.1016/j.jplph.2024.154218
[55]

Upadhyay R, Saini R, Shukla PK, Tiwari KN. 2025. Role of secondary metabolites in plant defense mechanisms: a molecular and biotechnological insights. Phytochemistry Reviews 24:953−83

doi: 10.1007/s11101-024-09976-2
[56]

Chang A, Lin L, Kangzhuo Y, Shengzhen C, Bingrui W, et al. 2023. 药用植物鳄嘴花(忧遁草)的研究概况及质量标志物预测分析 [Comprehensive review on research of Ezuihua (Clinacanthus nutans) and predictive analysis on quality marker (Q-marker)]. 中华中医药学刊 [Chinese Archives of Traditional Chinese Medicine] 41:25−35

doi: 10.13193/j.issn.1673-7717.2023.12.005
[57]

Lin CM, Chen HH, Lung CW, Chen HJ. 2023. Antiviral and immunomodulatory activities of Clinacanthus nutans (Burm. f.) Lindau. International Journal of Molecular Sciences 24:10789

doi: 10.3390/ijms241310789
[58]

Lei P, Jiang Y, Zhao Y, Jiang M, Ji X, et al. 2024. Functions of basic helix-loop-helix (bHLH) proteins in the regulation of plant responses to cold, drought, salt, and iron deficiency: a comprehensive review. Journal of Agricultural and Food Chemistry 72:10692−709

doi: 10.1021/acs.jafc.3c09665
[59]

Gao F, Dubos C. 2024. The arabidopsis bHLH transcription factor family. Trends in Plant Science 29:668−80

doi: 10.1016/j.tplants.2023.11.022
[60]

Wu Y, Wu S, Wang X, Mao T, Bao M, et al. 2022. Genome-wide identification and characterization of the bHLH gene family in an ornamental woody plant Prunus mume. Horticultural Plant Journal 8:531−44

doi: 10.1016/j.hpj.2022.01.004
[61]

Liang J, Fang Y, An C, Yao Y, Wang X, et al. 2023. Genome-wide identification and expression analysis of the bHLH gene family in passion fruit (Passiflora edulis) and its response to abiotic stress. International Journal of Biological Macromolecules 225:389−403

doi: 10.1016/j.ijbiomac.2022.11.076
[62]

Flagel LE, Wendel JF. 2009. Gene duplication and evolutionary novelty in plants. New Phytologist 183:557−64

doi: 10.1111/j.1469-8137.2009.02923.x
[63]

Jain M, Tyagi AK, Khurana JP. 2008. Genome-wide identification, classification, evolutionary expansion and expression analyses of homeobox genes in rice. The FEBS Journal 275:2845−61

doi: 10.1111/j.1742-4658.2008.06424.x
[64]

Liu H, Lyu HM, Zhu K, Van de Peer Y, Max Cheng ZM. 2021. The emergence and evolution of intron-poor and intronless genes in intron-rich plant gene families. The Plant Journal 105:1072−82

doi: 10.1111/tpj.15088
[65]

Wu Y, Wen J, Xia Y, Zhang L, Du H. 2022. Evolution and functional diversification of R2R3-MYB transcription factors in plants. Horticulture Research 9:uhac058

doi: 10.1093/hr/uhac058
[66]

Ye D, Liu J, Tian X, Wen X, Zhang Y, et al. 2024. Genome-wide identification of bHLH gene family and screening of candidate gene in response to salt stress in kiwifruit. Environmental and Experimental Botany 222:105774

doi: 10.1016/j.envexpbot.2024.105774
[67]

He J, Yao L, Pecoraro L, Liu C, Wang J, et al. 2023. Cold stress regulates accumulation of flavonoids and terpenoids in plants by phytohormone, transcription process, functional enzyme, and epigenetics. Critical Reviews in Biotechnology 43:680−97

doi: 10.1080/07388551.2022.2053056
[68]

Jiang M, Niu Y, Wen G, Zhao C, Gao Q, et al. 2024. Evolutionary studies of the basic helix–loop–helix (bHLH) IVc gene family in plants and the role of AtILR3 in Arabidopsis response to ABA stress. Physiologia Plantarum 176:e14128

doi: 10.1111/ppl.14128
[69]

Liao HS, Yang CC, Hsieh MH. 2022. Nitrogen deficiency-and sucrose-induced anthocyanin biosynthesis is modulated by HISTONE DEACETYLASE15 in Arabidopsis. Journal of Experimental Botany 73:3726−42

doi: 10.1093/jxb/erac067
[70]

Le Hir R, Castelain M, Chakraborti D, Moritz T, Dinant S, et al. 2017. AtbHLH68 transcription factor contributes to the regulation of ABA homeostasis and drought stress tolerance in Arabidopsis thaliana. Physiologia Plantarum 160:312−27

doi: 10.1111/ppl.12549
[71]

Kim D, Jeon SJ, Yanders S, Park S, Kim HS, et al. 2022. MYB3 plays an important role in lignin and anthocyanin biosynthesis under salt stress condition in Arabidopsis. Plant Cell Reports 41:1549−60

doi: 10.1007/s00299-022-02878-7
[72]

He Z, Wang Z, Nie X, Qu M, Zhao H, et al. 2022. UNFERTILIZED EMBRYO SAC 12 phosphorylation plays a crucial role in conferring salt tolerance. Plant Physiology 188:1385−401

doi: 10.1093/plphys/kiab549
[73]

Chen HC, Hsieh-Feng V, Liao PC, Cheng WH, Liu LY, et al. 2017. The function of OsbHLH068 is partially redundant with its homolog, AtbHLH112, in the regulation of the salt stress response but has opposite functions to control flowering in Arabidopsis. Plant Molecular Biology 94:531−48

doi: 10.1007/s11103-017-0624-6