The MabZIP5-MaMYB69 module cooperates with MaERF55 to modulate banana fruit ripening via cell wall degradation

Abstract

The MYB protein family comprises numerous transcription factors with important functions in various biological processes in plants; however, their role in modulating banana fruit ripening has been rarely investigated. In this study, we identified an R2R3-type MYB gene, MaMYB69, which promotes fruit ripening by directly modulating cell wall degradation. The expression of MaMYB69, which encodes a nucleus-localized transcription activator, was induced by ethylene and upregulated during banana ripening. MaMYB69 activated the expression of MaPE, MaPL1, MaGAL, and MaPG3 by directly targeting their promoters. MaMYB69 overexpression in tomato and banana accelerated fruit ripening and stimulated cell wall-modifying gene expression. Moreover, MaMYB69 interacted with MaERF55 and strengthened its transactivation activity through downstream target genes. MabZIP5, an aroma biosynthesis regulator, acted directly upstream of MaMYB69, enhancing its transcription. MaMYB69 also formed a homodimer with itself and activated its expression. Collectively, our results show that MaMYB69 is pivotal in controlling banana ripening and softening and improve our understanding of the regulatory cascades involved in fruit ripening. MaMYB69 may serve as a potential target for improving fruit quality and extending the shelf life.

Acknowledgements

This work received financial support from the Natural Science Foundation of China (32172544, 32261160375), the special fund for scientific innovation strategy-construction of high level Academy of Agriculture Science (R2023PY-JG003), the earmarked fund for CARS (CARS-31-01), Guangdong Special Support Program (NYLJ2024010), and IAEA CRP D23033.

Authors contributions

Experimental work and data analysis were performed by F.L. and X.S. Data analysis was conducted by J.C., O.S., To.D., T.D., Y.L., H.G., and F.B. The manuscript was prepared by F.L. and X.S. Results interpretation involved W.H., Q.Y., Ch.L., C.H., C.L., and Y.L. Experiments design, coordination, and manuscript revision were handled by G.Y. and F.B.

Data availability

The article incorporates all experimental data generated in this study.

Conflicts of interest statement

No competing interests are declared by the authors.

Supplementary material

Supplementary material is available at Horticulture Research online.

References

[1] Brumos J. Gene regulation in climacteric fruit ripening. Curr Opin Plant Biol. 2021; 63:102042

[2] Li XM, Wang XM, Zhang Y. et al. Regulation of fleshy fruit ripen-ing: from transcription factors to epigenetic modifications. Hortic Res. 2022;9:uhac013

[3] Peng ZZ, Liu GS, Li HL. et al. Molecular and genetic events determining the softening of fleshy fruits: a comprehensive review. Int J Mol Sci. 2022; 23:12482

[4] Shi YN, Li BJ, Su GQ. et al. Transcriptional regulation of fleshy fruit texture. J Integr Plant Biol. 2022; 64:1649-72

[5] Maduwanthi SDT, Marapana RAUJ. Induced ripening agents and their effect on fruit quality of banana. Int J Food Sci. 2019; 2019:1-8

[6] Zhai ZF, Xiao YQ, Wang YY. et al. Abscisic acid-responsive transcription factors PavDof2/6/15 mediate fruit softening in sweet cherry. Plant Physiol. 2022; 190:2501-18

[7] Xiao L, Li TT, Jiang GX. et al. Cell wall proteome analysis of banana fruit softening using iTRAQ technology. J Proteome. 2019; 209:103506

[8] Wang DD, Samsulrizal NH, Yan C. et al. Characterization of CRISPR mutants targeting genes modulating pectin degrada-tion in ripening tomato. Plant Physiol. 2019; 179:544-57

[9] Zhang WW, Zhao SQ, Gu S. et al. FvWRKY48 binds to the pec-tate lyase FvPLA promoter to control fruit softening in Fragaria vesca. Plant Physiol. 2022; 189:1037-49

[10] Nie HM, Shi Y, Geng XQ. et al. CRISRP/Cas9-mediated targeted mutagenesis of tomato polygalacturonase gene (SlPG) delays fruit softening. Front Plant Sci. 2022; 13:729128

[11] Yang L, Huang W, Xiong FJ. et al. Silencing of SlPL, which encodes a pectate lyase in tomato, confers enhanced fruit firmness, prolonged shelf-life and reduced susceptibility to grey mould. Plant Biotechnol J. 2017; 15: 1544-55

[12] Jiménez NP, Bjornson M, Famula RA. et al. Loss-of-function mutations in the fruit softening gene POLYGALACTURONASE1 doubled fruit firmness in strawberry. Hortic Res. 2025;12:uhae315

[13] Paniagua C, Blanco-Portales R, Barceló-Muñoz M. et al. Anti-sense down-regulation of the strawberry β-galactosidase gene FaβGal4 increases cell wall galactose levels and reduces fruit softening. J Exp Bot. 2016; 67:619-31

[14] Ortega Salazar I, Crum D, Sbodio AO. et al. Double CRISPR knockout of pectin degrading enzymes improves tomato shelf-life while ensuring fruit quality. Plants People Planet. 2024; 6:330-40

[15] Shi YN, Vrebalov J, Zheng H. et al. A tomato LATERAL ORGAN BOUNDARIES transcription factor, SlLOB1, predominantly reg-ulates cell wall and softening components of ripening. Proc Natl Acad Sci USA. 2021; 118:e2102486118

[16] Gao Y, Zhang YP, Fan ZQ. et al. Mutagenesis of SlNAC4 by CRISPR/Cas9 alters gene expression and softening of ripening tomato fruit. Vegetable Res. 2021; 1:1-12

[17] Wang MM, Wu Y, Zhan W. et al. The apple transcription fac-tor MdZF-HD11 regulates fruit softening by promoting Mdβ-GAL18 expression. J Exp Bot. 2024; 75:819-36

[18] Fu CC, Han C, Wei YX. et al. Two NAC transcription factors regulated fruit softening through activating xyloglucan endo-transglucosylase/hydrolase genes during kiwifruit ripening. Int J Biol Macromol. 2024; 263:130678

[19] Wang J, Lu XM, Kuang MM. et al. Transcriptome analysis iden-tified a transcriptional repressor, BES1-INTERACTING MYC-LIKE1 regulating rapid postharvest fruit softening of Actinidia arguta. Postharvest Biol Technol. 2024; 217:113107

[20] Liu F, Dou TX, Hu CH. et al. WRKY transcription factor MaWRKY49 positively regulates pectate lyase genes during fruit ripening of Musa acuminata. Plant Physiol Biochem. 2023; 194:643-50

[21] Wu CJ, Cai DL, Li J. et al. Banana MabHLH28 positively reg-ulates the expression of softening-related genes to medi-ate fruit ripening independently or via cooperating with MaWRKY49/111. Hortic Res. 2024;11:uhae053

[22] Chen TH, Wei W, Shan W. et al. MaHDA6-MaNAC154 mod-ule regulates the transcription of cell wall modification genes during banana fruit ripening. Postharvest Biol Technol. 2023; 198:112237

[23] Wu XY, Xia M, Su P. et al. MYB transcription factors in plants: a comprehensive review of their discovery, structure, classifica-tion, functional diversity and regulatory mechanism. Int J Biol Macromol. 2024; 282:136652

[24] Zhang YS, Xu Y, Huang DM. et al. Research progress on the MYB transcription factors in tropical fruit. Trop Plants. 2022; 1:1-15

[25] Cao HH, Chen J, Yue M. et al. Tomato transcriptional repressor MYB70 directly regulates ethylene-dependent fruit ripening. Plant J. 2020; 104:1568-81

[26] Jian W, Cao HH, Yuan S. et al. SlMYB75, an MYB-type tran-scription factor, promotes anthocyanin accumulation and enhances volatile aroma production in tomato fruits. Hortic Res. 2019; 6:22

[27] Jin WM, Wang H, Li MF. et al. The R2R3 MYB transcription factor PavMYB10.1 involves in anthocyanin biosynthesis and determines fruit skin colour in sweet cherry (Prunus avium L.). Plant Biotechnol J. 2016; 14:2120-33

[28] Fan ZQ, Ba LJ, Shan W. et al. A banana R2R3-MYB transcription factor MaMYB3 is involved in fruit ripening through modulation of starch degradation by repressing starch degradation-related genes and MabHLH6. Plant J. 2018; 96: 1191-205

[29] Wei W, Yang YY, Lakshmanan P. et al. Proteasomal degradation of MaMYB60 mediated by the E3 ligase MaBAH1 causes high temperature-induced repression of chlorophyll catabolism and green ripening in banana. Plant Cell. 2023; 35:1408-28

[30] Cai JF, Mo XL, Wen CJ. et al. FvMYB79 positively regulates strawberry fruit softening via transcriptional activation of FvPME38. Int J Mol Sci. 2021; 23:101

[31] Yang L, He JL, Qin SJ. et al. MYB transcription factor MdMYB44 positively regulates fruit crispness by directly activating the expression of pectin methylesterase MdMPE3 in apple. Plant Physiol Biochem. 2025; 224:109936

[32] Yang YY, Shan W, Yang TW. et al. MaMYB4 is a negative reg-ulator and a substrate of RING-type E3 ligases MaBRG2/3 in controlling banana fruit ripening. Plant J. 2022; 110: 1651-69

[33] Suriyamoorthy P, Madhuri A, Tangirala S. et al. Comprehensive review on banana fruit allergy: pathogenesis, diagnosis, management, and potential modification of allergens through food processing. Plant Food Hum Nutr. 2022; 77:159-71

[34] Al-Dairi M, Pathare PB, Al-Yahyai R. et al. Postharvest qual-ity, technologies, and strategies to reduce losses along the supply chain of banana: a review. Trends Food Sci Technol. 2023; 134:177-91

[35] Liu F, Sun XL, He J. et al. Membrane-associated NAC transcrip-tion factor MaNAC169 is a positive regulator during banana fruit ripening. Postharvest Biol Technol. 2025; 223:113451

[36] Li B, Fan RY, Yang QS. et al. Genome-wide identification and characterization of the NAC transcription factor family in Musa acuminata and expression analysis during fruit ripening. Int J Mol Sci. 2020; 21:634

[37] Wang Z, Yao XM, Jia CH. et al. Genome-wide characterization and analysis of R2R3-MYB genes related to fruit ripening and stress response in banana (Musa acuminata L. AAA Group, cv ’Cavendish’). Plants. 2022; 12:152

[38] Kelemen Z, Sebastian A, Xu W. et al. Analysis of the DNA-binding activities of the Arabidopsis R2R3-MYB transcription factor family by one-hybrid experiments in yeast. PLoS One. 2015; 10:e0141044

[39] Chang Y, Chen G, Yang G. et al. The PcERF5 promotes antho-cyanin biosynthesis in red-fleshed pear (Pyrus communis) through both activating and interacting with PcMYB transcrip-tion factors. J Integr Agric. 2023; 22:2687-704

[40] Guo YF, Zhang YL, Shan W. et al. Identification of two tran-scriptional activators MabZIP4/5 in controlling aroma biosyn-thetic genes during banana ripening. J Agric Food Chem. 2018; 66:6142-50

[41] Yang YL, Xu Y, Feng BZ. et al. Regulatory networks of bZIPs in drought, salt and cold stress response and signaling. Plant Sci. 2025; 352:112399

[42] Moya-León MA, Mattus-Araya E, Herrera R. Molecular events occurring during softening of strawberry fruit. Front Plant Sci. 2019; 10:615

[43] Asif MH, Lakhwani D, Pathak S. et al. Transcriptome analy-sis of ripe and unripe fruit tissue of banana identifies major metabolic networks involved in fruit ripening process. BMC Plant Biol. 2014; 14:316

[44] Kuang JF, Wu CJ, Guo YF. et al. Deciphering transcriptional regulators of banana fruit ripening by regulatory network analysis. Plant Biotechnol J. 2021; 19:477-89

[45] Fu CC, Chen HJ, Gao HY. et al. Two papaya MYB proteins function in fruit ripening by regulating some genes involved in cell-wall degradation and carotenoid biosynthesis. J Sci Food Agric. 2020; 100:4442-8

[46] Jiang GX, Zhang DD, Li ZW. et al. Alternative splicing of MaMYB16L regulates starch degradation in banana fruit dur-ing ripening. J Integr Plant Biol. 2021; 63:1341-52

[47] Jeong CY, Kim JH, Lee WJ. et al. AtMyb56 regulates antho-cyanin levels via the modulation of AtGPT2 expression in response to sucrose in Arabidopsis. Mol Cells. 2018; 41: 351-61

[48] Xue YS, Shan YF, Yao JL. et al. The transcription factor PbrMYB24 regulates lignin and cellulose biosynthesis in stone cells of pear fruits. Plant Physiol. 2023; 192:1997-2014

[49] Xu Q, Yin XR, Zeng JK. et al. Activator- and repressor-type MYB transcription factors are involved in chilling injury induced flesh lignification in loquat via their interactions with the phenylpropanoid pathway. J Exp Bot. 2014; 65: 4349-59

[50] Wu CJ, Shan W, Liu XC. et al. Phosphorylation of transcription factor bZIP21 by MAP kinase MPK6-3 enhances banana fruit ripening. Plant Physiol. 2022; 188:1665-85

[51] Wu CJ, Su XG, Shan W. et al. Banana MaWRKY49 and MaWRKY111 cooperate with MabZIP21 to activate the tran-scription of MaACS1 and MaACO1 during fruit ripening. Postharvest Biol Technol. 2022; 194:112087

[52] Rajput R, Naik J, Stracke R. et al. Interplay between R2R3 MYB-type activators and repressors regulates proanthocyani-din biosynthesis in banana (Musa acuminata). New Phytol. 2022; 236:1108-27

[53] Zeng JK, Li X, Xu Q. et al. EjAP2-1, an AP2/ERF gene, is a novel regulator of fruit lignification induced by chilling injury, via interaction with EjMYB transcription factors. Plant Biotechnol J. 2015; 13:1325-34

[54] Zhang YY, Yin XR, Xiao YW. et al. An ETHYLENE RESPONSE FACTOR-MYB transcription complex regulates furaneol biosynthesis by activating QUINONE OXIDOREDUCTASE expression in strawberry. Plant Physiol. 2018; 178:189-201

[55] Jiang SH, Sun QG, Zhang TL. et al. MdMYB114 regulates antho-cyanin biosynthesis and functions downstream of MdbZIP4-like in apple fruit. J Plant Physiol. 2021; 257:153353

[56] Wang YY, Zhang XD, Zhao YR. et al. Transcription factor PyHY5 binds to the promoters of PyWD40 and PyMYB10 and regulates its expression in red pear ’Yunhongli No. 1’. Plant Physiol Biochem. 2020; 154:665-74

[57] Wu CJ, Shan W, Liang SM. et al. MaMPK2 enhances MabZIP93-mediated transcriptional activation of cell wall modify-ing genes during banana fruit ripening. Plant Mol Biol. 2019; 101:113-27

[58] Liu F, Sun XL, Sheng O. et al. Genome-wide analysis of the tri-helix gene family reveals that MaGT21 modulates fruit ripen-ing by regulating the expression of MaACO1 in Musa acumi-nata. Plant Physiol Biochem. 2024; 216:109089

[59] Fillatti JJ, Kiser J, Rose R. et al. Efficient transfer of a glyphosate tolerance gene into tomato using a binary Agrobacterium tumefaciens vector. Nat Biotechnol. 1987; 5:726-30