GmNF-YC4 delays soybean flowering and maturation by directly repressing GmFT2a and GmFT5a expression

Abstract

Flowering time and growth period are key agronomic traits which directly affect soybean (Glycine max (L.) Merr.) adaptation to diverse latitudes and farming systems. The FLOWERING LOCUS T (FT) homologs GmFT2a and GmFT5a integrate multiple flowering regulation pathways and significantly advance flowering and maturity in soybean. Pinpointing the genes responsible for regulating GmFT2a and GmFT5a will improve our understanding of the molecular mechanisms governing growth period in soybean. In this study, we identified the Nuclear Factor Y-C (NFY-C) protein GmNF-YC4 as a novel flowering suppressor in soybean under long-day (LD) conditions. GmNF-YC4 delays flowering and maturation by directly repressing the expression of GmFT2a and GmFT5a. In addition, we found that a strong selective sweep event occurred in the chromosomal region harboring the GmNF-YC4 gene during soybean domestication. The GmNF-YC4^Hap3 allele was mainly found in wild soybean (Glycine soja Siebold & Zucc.) and has been eliminated from G. max landraces and improved cultivars, which predominantly contain the GmNF-YC4^Hap1 allele. Furthermore, the Gmnf-yc4 mutants displayed notably accelerated flowering and maturation under LD conditions. These alleles may prove to be valuable genetic resources for enhancing soybean adaptability to higher latitudes.

Keywords

CRISPR/Cas9 / early flowering and maturity / GmNFYC4 / regional adaptability / soybean

References

[1] Abe, M.,Kobayashi, Y.,Yamamoto, S.,Daimon, Y.,Yamaguchi, A.,Ikeda, Y.,Ichinoki, H.,Notaguchi, M.,Goto, K., and Araki, T. (2005). FD, a bZIP protein mediating signals from the floral pathway integrator FT at the shoot apex. Science 309:1052–1056.

[2] Bäurle, I., and Dean, C. (2006). The timing of developmental transitions in plants. Cell 125:655–664.

[3] Cai, Y.,Chen, L.,Liu, X.,Guo, C.,Sun, S.,Wu, C.,Jiang, B.,Han, T., and Hou, W. (2018). CRISPR/Cas9-mediated targeted mutagenesis of GmFT2a delays flowering time in soya bean. Plant Biotechnol. J. 16:176–185.

[4] Cai, Y.,Liu, Y.,Fan, Y.,Li, X.,Yang, M.,Xu, D.,Wang, H.,Deng, X.W., and Li, J. (2023). MYB112 connects light and circadian clock signals to promote hypocotyl elongation in Arabidopsis. Plant Cell 35:3485–3503.

[5] Cai, Y.,Wang, L.,Chen, L.,Wu, T.,Liu, L.,Sun, S.,Wu, C.,Yao, W.,Jiang, B.,Yuan, S., et al. (2020). Mutagenesis of GmFT2a and GmFT5a mediated by CRISPR/Cas9 contributes for expanding the regional adaptability of soybean. Plant Biotechnol. J. 18:298–309.

[6] Chen, L.,Cai, Y.,Liu, X.,Yao, W.,Guo, C.,Sun, S.,Wu, C.,Jiang, B.,Han, T., and Hou, W. (2018). Improvement of soybean Agrobacterium-mediated transformation efficiency by adding glutamine and asparagine into the culture media. Int. J. Mol. Sci. 19:3039.

[7] Chen, L.,Nan, H.,Kong, L.,Yue, L.,Yang, H.,Zhao, Q.,Fang, C.,Li, H.,Cheng, Q.,Lu, S., et al. (2020). Soybean AP1 homologs control flowering time and plant height. J. Integr. Plant Biol. 62:1868–1879.

[8] Chi, Y.,Huang, F.,Liu, H.,Yang, S., and Yu, D. (2011). An APETALA1-like gene of soybean regulates flowering time and specifies floral organs. J. Plant Physiol. 168:2251–2259.

[9] Doebley, J.F.,Gaut, B.S., and Smith, B.D. (2006). The molecular genetics of crop domestication. Cell 127:1309–1321.

[10] Dong, L.,Cheng, Q.,Fang, C.,Kong, L.,Yang, H.,Hou, Z.,Li, Y.,Nan, H.,Zhang, Y.,Chen, Q., et al. (2022). Parallel selection of distinct T of 5 alleles drove the adaptation of cultivated and wild soybean to high latitudes. Mol. Plant 15:308–321.

[11] Dong, L.,Li, S.,Wang, L.,Su, T.,Zhang, C.,Bi, Y.,Lai, Y.,Kong, L.,Wang, F.,Pei, X., et al. (2023). The genetic basis of high-latitude adaptation in wild soybean. Curr. Biol. 33:252–262.e254.

[12] Hackenberg, D.,Keetman, U., and Grimm, B. (2012). Homologous NF-YC2 subunit from Arabidopsis and tobacco is activated by photooxidative stress and induces flowering. Int. J. Mol. Sci. 13:3458–3477.

[13] Hellens, R.P.,Allan, A.C.,Friel, E.N.,Bolitho, K.,Grafton, K.,Templeton, M.D.,Karunairetnam, S.,Gleave, A.P., and Laing, W.A. (2005). Transient expression vectors for functional genomics, quantification of promoter activity and RNA silencing in plants. Plant Methods 1:13.

[14] Hwang, Y.-H.,Kim, S.-K.,Lee, K.C.,Chung, Y.S.,Lee, J.H., and Kim, J.-K. (2016). Functional conservation of rice OsNF-YB/YC and Arabidopsis AtNF-YB/YC proteins in the regulation of flowering time. Plant Cell Rep. 35:857–865.

[15] Jia, Z.,Jiang, B.,Gao, X.,Yue, Y.,Fei, Z.,Sun, H.,Wu, C.,Sun, S.,Hou, W., and Han, T. (2015). GmFULa, a FRUITFULL homolog, functions in the flowering and maturation of soybean. Plant Cell Rep. 34:121–132.

[16] Kim, M.Y.,Van, K.,Kang, Y.J.,Kim, K.H., and Lee, S.H. (2012). Tracing soybean domestication history: From nucleotide to genome. Breed. Sci. 61:445–452.

[17] Kong, F.,Liu, B.,Xia, Z.,Sato, S.,Kim, B.M.,Watanabe, S.,Yamada, T.,Tabata, S.,Kanazawa, A.,Harada, K., et al. (2010). Two coordinately regulated homologs of FLOWERING LOCUS T are involved in the control of photoperiodic flowering in soybean. Plant Physiol. 154:1220–1231.

[18] Kou, K.,Yang, H.,Li, H.,Fang, C.,Chen, L.,Yue, L.,Nan, H.,Kong, L.,Li, X.,Wang, F., et al. (2022). A functionally divergent SOC1 homolog improves soybean yield and latitudinal adaptation. Curr. Biol. 32:1728–1742.

[19] Kumimoto, R.W.,Zhang, Y.,Siefers, N., and Holt, 3rd, B.F. (2010). NF-YC3, NF-YC4 and NF-YC9 are required for CONSTANS-mediated, photoperiod-dependent flowering in Arabidopsis thaliana. Plant J. 63:379–391.

[20] Lei, Y.,Lu, L.,Liu, H.Y.,Li, S.,Xing, F., and Chen, L.L. (2014). CRISPR-P: A web tool for synthetic single-guide RNA design of CRISPR-system in plants. Mol. Plant 7:1494–1496.

[21] Li, C.,Distelfeld, A.,Comis, A., and Dubcovsky, J. (2011). Wheat flowering repressor VRN2 and promoter CO₂ compete for interactions with NUCLEAR FACTOR-Y complexes. Plant J. 67:763–773.

[22] Li, H.,Du, H.,Huang, Z.,He, M.,Kong, L.,Fang, C.,Chen, L.,Yang, H.,Zhang, Y.,Liu, B., et al. (2023). The AP2/ERF transcription factor TOE4b regulates photoperiodic flowering and grain yield per plant in soybean. Plant Biotechnol. J. 21:1682–1694.

[23] Li, X.,Fang, C.,Yang, Y.,Lv, T.,Su, T.,Chen, L.,Nan, H.,Li, S.,Zhao, X.,Lu, S., et al. (2021). Overcoming the genetic compensation response of soybean florigens to improve adaptation and yield at low latitudes. Curr. Biol. 31:3755–3767.

[24] Li, X.,Zhou, H.,Cheng, L.,Ma, N.,Cui, B.,Wang, W.,Zhong, Y., and Liao, H. (2022). Shoot-to-root translocated GmNN1/FT2a triggers nodulation and regulates soybean nitrogen nutrition. PLoS Biol. 20: e3001739.

[25] Lin, X.,Liu, B.,Weller, J.L.,Abe, J., and Kong, F. (2021). Molecular mechanisms for the photoperiodic regulation of flowering in soybean. J. Integr. Plant Biol. 63:981–994.

[26] Liu, H.,Guo, Y.,Wang, H.,Yang, W.,Yang, J.,Zhang, J.,Liu, D.,El-Kassaby, Y.A., and Li, W. (2022). Involvement of PtCOL5-PtNF-YC4 in reproductive cone development and gibberellin signaling in Chinese pine. Plant Sci. 323:111383.

[27] Liu, Y.,Du, H.,Li, P.,Shen, Y.,Peng, H.,Liu, S.,Zhou, G.A.,Zhang, H.,Liu, Z.,Shi, M., et al. (2020). Pan-genome of wild and cultivated soybeans. Cell 182:162–176.e113.

[28] Liu, Y.,Zhang, Y.,Liu, X.,Shen, Y.,Tian, D.,Yang, X.,Liu, S.,Ni, L.,Zhang, Z.,Song, S., et al. (2023). SoyOmics: A deeply integrated database on soybean multi-omics. Mol. Plant 16:794–797.

[29] Lu, L.,Wei, W.,Li, Q.T.,Bian, X.H.,Lu, X.,Hu, Y.,Cheng, T.,Wang, Z.Y.,Jin, M.,Tao, J.J., et al. (2021b). A transcriptional regulatory module controls lipid accumulation in soybean. New Phytol. 231:661–678.

[30] Lu, L.,Wei, W.,Tao, J.J.,Lu, X.,Bian, X.H.,Hu, Y.,Cheng, T.,Yin, C.C.,Zhang, W.K.,Chen, S.Y., et al. (2021a). Nuclear factor Y subunit GmNFYA competes with GmHDA13 for interaction with GmFVE to positively regulate salt tolerance in soybean. Plant Biotechnol. J. 19:2362–2379.

[31] Lu, S.,Dong, L.,Fang, C.,Liu, S.,Kong, L.,Cheng, Q.,Chen, L.,Su, T.,Nan, H.,Zhang, D., et al. (2020). Stepwise selection on homeologous PRR genes controlling flowering and maturity during soybean domestication. Nat. Genet. 52:428–436.

[32] Luo, X.,Gao, Z.,Wang, Y.,Chen, Z.,Zhang, W.,Huang, J.,Yu, H., and He, Y. (2018). The NUCLEAR FACTOR-CONSTANS complex antagonizes Polycomb repression to de-repress FLOWERING LOCUS T expression in response to inductive long days in Arabidopsis. Plant J. 95:17–29.

[33] Lyu, X.,Li, Y.H.,Li, Y.,Li, D.,Han, C.,Hong, H.,Tian, Y.,Han, L.,Liu, B., and Qiu, L.J. (2023). The domestication-associated L1 gene encodes a eucomic acid synthase pleiotropically modulating pod pigmentation and shattering in soybean. Mol. Plant 16:1178–1191.

[34] Mallano, A.I.,Li, W.,Tabys, D.,Chao, C.,Yang, Y.,Anwar, S.,Almas, H.I.,Nisa, Z.U., and Li, Y. (2021). The soybean GmNFY-B1 transcription factor positively regulates flowering in transgenic Arabidopsis. Mol. Biol. Rep. 48:1589–1599.

[35] Nan, H.,Cao, D.,Zhang, D.,Li, Y.,Lu, S.,Tang, L.,Yuan, X.,Liu, B., and Kong, F. (2014). GmFT2a and GmFT5a redundantly and differentially regulate flowering through interaction with and upregulation of the bZIP transcription factor GmFDL19 in soybean. PLoS One 9: e97669.

[36] Ni, Z.,Hu, Z.,Jiang, Q., and Zhang, H. (2013). GmNFYA3, a target gene of miR169, is a positive regulator of plant tolerance to drought stress. Plant Mol. Biol. 82:113–129.

[37] O’Conner, S.,Zheng, W.,Qi, M.,Kandel, Y.,Fuller, R.,Whitham, S.A., and Li, L. (2021). GmNF-YC4-2 increases protein, exhibits broad disease resistance and expedites maturity in soybean. Int. J. Mol. Sci. 22:3586.

[38] Olsen, K.M., and Wendel, J.F. (2013). A bountiful harvest: Genomic insights into crop domestication phenotypes. Annu. Rev. Plant Biol. 64:47–70.

[39] Qi, M.,Zheng, W.,Zhao, X.,Hohenstein, J.D.,Kandel, Y.,O’Conner, S.,Wang, Y.,Du, C.,Nettleton, D.,MacIntosh, G.C., et al. (2019). QQS orphan gene and its interactor NF-YC4 reduce susceptibility to pathogens and pests. Plant Biotechnol. J. 17:252–263.

[40] Quach, T.N.,Nguyen, H.T.,Valliyodan, B.,Joshi, T.,Xu, D., and Nguyen, H.T. (2015). Genome-wide expression analysis of soybean NF-Y genes reveals potential function in development and drought response. Mol. Genet. Genomics 290:1095–1115.

[41] Wang, Y.,Xu, C.,Sun, J.,Dong, L.,Li, M.,Liu, Y.,Wang, J.,Zhang, X.,Li, D.,Sun, J., et al. (2021). GmRAV confers ecological adaptation through photoperiod control of flowering time and maturity in soybean. Plant Physiol. 187:361–377.

[42] Wei, Q.,Ma, C.,Xu, Y.,Wang, T.,Chen, Y.,Lü J.,Zhang, L.,Jiang, C.-Z.,Hong, B., and Gao, J. (2017). Control of chrysanthemum flowering through integration with an aging pathway. Nat. Commun. 8:829.

[43] Wigge, P.A.,Kim, M.C.,Jaeger, K.E.,Busch, W.,Schmid, M.,Lohmann, J.U., and Weigel, D. (2005). Integration of spatial and temporal information during floral induction in Arabidopsis. Science 309:1056–1059.

[44] Wu, F.,Sedivy, E.J.,Price, W.B.,Haider, W., and Hanzawa, Y. (2017). Evolutionary trajectories of duplicated FT homologues and their roles in soybean domestication. Plant J. 90:941–953.

[45] Xu, H.,Li, Y.,Zhang, K.,Li, M.,Fu, S.,Tian, Y.,Qin, T.,Li, X.,Zhong, Y., and Liao, H. (2021). miR169c-NFYA-C-ENOD40 modulates nitrogen inhibitory effects in soybean nodulation. New Phytol. 229:3377–3392.

[46] Yang, S.,Miao, L.,He, J.,Zhang, K.,Li, Y., and Gai, J. (2019). Dynamic transcriptome changes related to oil accumulation in developing soybean seeds. Int. J. Mol. Sci. 20:2202.

[47] Yu, T.F.,Liu, Y.,Fu, J.D.,Ma, J.,Fang, Z.W.,Chen, J.,Zheng, L.,Lu, Z.W.,Zhou, Y.B.,Chen, M., et al. (2021). The NF-Y-PYR module integrates the abscisic acid signal pathway to regulate plant stress tolerance. Plant Biotechnol. J. 19:2589–2605.

[48] Yue, L.,Li, X.,Fang, C.,Chen, L.,Yang, H.,Yang, J.,Chen, Z.,Nan, H.,Chen, L.,Zhang, Y., et al. (2021). FT5a interferes with the Dt1-AP1 feedback loop to control flowering time and shoot determinacy in soybean. J. Integr. Plant Biol. 63:1004–1020.

[49] Zeng, X.,Liu, H.,Du, H.,Wang, S.,Yang, W.,Chi, Y.,Wang, J.,Huang, F., and Yu, D. (2018). Soybean MADS-box gene GmAGL1 promotes flowering via the photoperiod pathway. BMC Genomics 19:51.

[50] Zhuang, Q.,Xue, Y.,Yao, Z.,Zhu, S.,Liang, C.,Liao, H., and Tian, J. (2021). Phosphate starvation responsive GmSPX5 mediates nodule growth through interaction with GmNF-YC4 in soybean (Glycine max). Plant J. 108:1422–1438.

RIGHTS & PERMISSIONS

2024 The Authors. Journal of Integrative Plant Biology published by John Wiley & Sons Australia, Ltd on behalf of Institute of Botany, Chinese Academy of Sciences.