Genes & Genomics

, Volume 41, Issue 11, pp 1341–1355 | Cite as

Genome-wide characterization of the NUCLEAR FACTOR-Y (NF-Y) family in Citrus grandis identified CgNF-YB9 involved in the fructose and glucose accumulation

  • Yiting Mai
  • Lanya Shui
  • Kaisen Huo
  • Jun NiuEmail author
Research Article



Nuclear factor Y (NF-Y) is increasingly known to be involved in many aspects of plant growth and development. To date, the systematic characterization of NF-Y family has never been reported in Citrus grandis.


Genome-wide characterization of C. grandis NF-Y (CgNF-Y) family and analysis of their role in sucrose metabolism.


NF-Y conserved models were employed to identify CgNF-Y genes from genomic data. Phylogenetic tree was generated by the neighbor-joining method using program MEGA 7.0. Based on our previous transcriptomic data, the transcription levels were calculated by RSEM software and were clustered by ShortTime-series Expression Miner. The plant expression vector of CgNF-YB9 was constructed using In-Fusion Cloning and transferred into tobacco by leaf disc transformation method. Soluble sugars and gene expressions were analysis by HPLC and qRT-PCR, respectively.


A total of 24 CgNF-Y genes (6 CgNF-YAs, 13 CgNF-YBs and 5 CgNF-YCs) were identified with conserved domains. Phylogenetic analysis of the NF-Y proteins indicated that NF-YA, NF-YB and NF-YC could be categorized into four, five and three clades, respectively. Expression profiling analysis reflected spatio-temporally distinct expression patterns for CgNF-Y genes. Importantly, we observed a positive correlation between the expression level of CgNF-YB9 and the content of soluble sugar. Moreover, CgNF-YB9-corelated genes were enriched in carbohydrate metabolism. In CgNF-YB9 overexpression lines, sucrose content showed a decrease, whereas glucose and fructose contents displayed an increase. As expected, the transcription levels of sucrose-phosphate synthase and vacuolar invertase in transgenic Line 3 were observed with significantly down- and up-regulated, respectively.


The structure, phylogenetic relationship and expression pattern of 24 CgNF-Y genes were identified, and CgNF-YB9 was involved in sucrose metabolism.


Citrus grandis NF-Y family CgNF-YB9 Sucrose metabolism 



Days after flowering


Fragment per kilobase of exon model per million mapped reads


Gene Ontology


High-performance liquid chromatography


Histone-fold motif






Kyoto Encyclopedia of Genes and Genomes


Multiple Em for Motif Elicitation


Nuclear factor Y


Pummelo juice sacs


Sucorse-phosphate phosphatase


Sucrose-phosphate synthase


Sucrose synthase


Vacuolar invertase


Wild tobacco



This research was supported by Natural Science Foundation of Hainan Province (317045) and Hainan university research funded projects (KYQD(ZR)1701).

Author contributions

JN conceived and designed the experiments. YM and LS wrote the manuscript and performed the analysis. KH conducted the transgenosis and detection.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

13258_2019_862_MOESM1_ESM.docx (21 kb)
Supplementary material 1 (DOCX 20 kb) Supplementary Table S1 The primers for qRT-PCR analysis
13258_2019_862_MOESM2_ESM.xls (418 kb)
Supplementary material 2 (XLS 418 kb) Supplementary Table S2 The CgNF-YB9-corelated genes


  1. Albani D, Robert LS (1995) Cloning and characterization of a Brassica napus gene encoding a homologue of the B subunit of a heteromeric CCAAT-binding factor. Gene 167:209–213PubMedGoogle Scholar
  2. Bailey TL, Johnson J, Grant CE, Noble WS (2015) The MEME suite. Nucleic Acids Res 43:W39PubMedPubMedCentralGoogle Scholar
  3. Baudin M, Laloum T, Lepage A, RãPodas C, Ariel F, Frances L, Crespi M, Gamas P, Blanco FA, Zanetti ME (2015) A phylogenetically conserved group of nuclear factor-Y transcription factors interact to control nodulation in legumes. Plant Physiol 169:2761–2773PubMedPubMedCentralGoogle Scholar
  4. Bieniawska Z, Barratt D, Garlick A, Thole V, Nj Martin C, Zrenner R, Smith A (2010) Analysis of the sucrose synthase gene family in Arabidopsis. Plant J 49:810–828Google Scholar
  5. Boivin EB, Lepage É, Matton DP, Crescenzo GD, Jolicoeur M (2011) Transient expression of antibodies in suspension plant cell suspension cultures is enhanced when co-transformed with the tomato bushy stunt virus p19 viral suppressor of gene silencing. Biotechnol Progr 26:1534–1543Google Scholar
  6. Cao S, Kumimoto RW, Siriwardana CL, Risinger JR (2011) Identification and characterization of NF-Y transcription factor families in the monocot model plant Brachypodium Distachyon. PLoS One 6:e21805PubMedPubMedCentralGoogle Scholar
  7. Ceribelli M, Dolfini D, Merico D, Gatta R, Viganã AM, Pavesi G, Mantovani R (2008) The histone-like NF-Y is a bifunctional transcription factor. Mol Cell Biol 28:2047–2058PubMedPubMedCentralGoogle Scholar
  8. Chourey PS, Taliercio EW, Carlson SJ, Ruan YL (1998) Genetic evidence that the two isozymes of sucrose synthase present in developing maize endosperm are critical, one for cell wall integrity and the other for starch biosynthesis. Mol Gen Genet 259:88–96PubMedGoogle Scholar
  9. Deng S, Mai Y, Niu J (2019) Fruit characteristics, soluble sugar compositions and transcriptome analysis during the development of Citrus maxima “seedless”, and identification of SUS and INV genes involved in sucrose degradation. Gene 689:131–140PubMedGoogle Scholar
  10. Dieter H, Ulrich K, Bernhard G (2012) Homologous NF-YC2 subunit from Arabidopsis and tobacco is activated by photooxidative stress and induces flowering. Int J Mol Sci 13:3458–3477Google Scholar
  11. Dolfini D, Gatta R, Mantovani R (2012) NF-Y and the transcriptional activation of CCAAT promoters. Crit Rev Biochem Mol Biol 47:29–49PubMedGoogle Scholar
  12. Ernst J, Bar-Joseph Z (2006) STEM: a tool for the analysis of short time series gene expression data. BMC Bioinform 7:191Google Scholar
  13. Fallahi H, Scofield GN, Badger MR, Chow WS, Furbank RT, Ruan Y-L (2008) Localization of sucrose synthase in developing seed and siliques of Arabidopsis thaliana reveals diverse roles for SUS during development. J Exp Bot 59:3283–3295PubMedPubMedCentralGoogle Scholar
  14. Feng ZJ, He GH, Zheng WJ, Lu PP, Chen M, Gong YM, Ma YZ, Xu ZS (2015) Foxtail millet NF-Y families: genome-wide survey and evolution analyses identified two functional genes important in abiotic stresses. Front Plant Sci 6:1142PubMedPubMedCentralGoogle Scholar
  15. Fornari M, Calvenzani V, Masiero S, Tonelli C, Petroni K (2013) The Arabidopsis NF-YA3 and NF-YA8 genes are functionally redundant and are required in early embryogenesis. PLoS One 8:e82043PubMedPubMedCentralGoogle Scholar
  16. Gusmaroli G, Tonelli C, Mantovani R (2002) Regulation of novel members of the Arabidopsis thaliana CCAAT-binding nuclear factor Y subunits. Gene 283:41–48PubMedGoogle Scholar
  17. Helin T, Xiaohui Y, Fengxia Z, Xiu Z, Cunmin Q, Jinye M, Fuyou F, Jiana L, Rongzhan G, Hongsheng Z (2011) Enhanced seed oil production in canola by conditional expression of Brassica napus LEAFY COTYLEDON1 and LEC1-LIKE in developing seeds. Plant Physiol 156:1577–1588Google Scholar
  18. Junker A, Mönke G, Rutten T, Keilwagen J, Seifert M, Thi TM, Renou JP, Balzergue S, Viehöver P, Hähnel U (2012) Elongation-related functions of LEAFY COTYLEDON1 during the development of Arabidopsis thaliana. Plant J 71:427–442PubMedGoogle Scholar
  19. Klann EM, Chetelat RT, Bennett AB (1993) Expression of acid invertase gene controls sugar composition in tomato (Lycopersicon) fruit. Plant Physiol 103:863PubMedPubMedCentralGoogle Scholar
  20. Komatsu A, Moriguchi T, Koyama K, Omura M, Akihama T (2002) Analysis of sucrose synthase genes in citrus suggests different roles and phylogenetic relationships. J Exp Bot 53:61–71PubMedGoogle Scholar
  21. Kumimoto RW, Adam L, Hymus GJ, Repetti PP, Reuber TL, Marion CM, Hempel FD, Ratcliffe OJ (2008) The nuclear factor Y subunits NF-YB2 and NF-YB3 play additive roles in the promotion of flowering by inductive long-day photoperiods in Arabidopsis. Planta 228:709–723PubMedGoogle Scholar
  22. Kusnetsov V, Landsberger M, Meurer J, Oelmüller R, Kusnetsov V, Landsberger M, Meurer J, Oelmüller R (1999) The assembly of the CAAT-box binding complex at a photosynthesis gene promoter is regulated by light, cytokinin, and the stage of the plastids. J Biol Chem 274:36009–36014PubMedGoogle Scholar
  23. Lee H, Fischer RL, Goldberg RB, Harada JJ (2003) Arabidopsis LEAFY COTYLEDON1 represents a functionally specialized subunit of the CCAAT binding transcription factor. Proc Natl Acad Sci USA 100:2152–2156PubMedGoogle Scholar
  24. Leloir LF, Cardini CE (1955) The biosynthesis of sucrose phosphate. J Biol Chem 214:157PubMedGoogle Scholar
  25. Li YJ, Fang Y, Fu YR, Huang JG, Wu CA, Zheng CC (2013) NFYA1 is involved in regulation of postgermination growth arrest under salt stress in Arabidopsis. PLoS One 8:e61289PubMedPubMedCentralGoogle Scholar
  26. Li S, Li K, Ju Z, Cao D, Fu D, Zhu H, Zhu B, Luo Y (2016) Genome-wide analysis of tomato NF-Y factors and their role in fruit ripening. BMC Genom 17:36Google Scholar
  27. Lotan T, Ohto M, Yee KM, West MA, Lo R, Kwong RW, Yamagishi K, Fischer RL, Goldberg RB, Harada JJ (1998) Arabidopsis LEAFY COTYLEDON1 is sufficient to induce embryo development in vegetative cells. Cell 93:1195–1205PubMedGoogle Scholar
  28. Luger K, Mäder AW, Richmond RK, Sargent DF, Richmond TJ (1997) Crystal structure of the nucleosome core particle at 2.8 Å resolution. Nature 389:251–260PubMedGoogle Scholar
  29. Maity SN, Crombrugghe BD (1998) Role of the CCAAT-binding protein CBF/NF-Y in transcription. Trends Biochem Sci 23:174–178PubMedGoogle Scholar
  30. Mantovani R (1999) The molecular biology of the CCAAT-binding factor NF-Y. Gene 239:15–27PubMedGoogle Scholar
  31. Mu J, Tan H, Hong S, Yan L, Zuo J (2013) Arabidopsis transcription factor genes NF-YA1, 5, 6, and 9 play redundant roles in male gametogenesis, embryogenesis, and seed development. Mol Plant 6:188–201PubMedGoogle Scholar
  32. Nelson DE, Repetti PP, Adams TR, Creelman RA, Wu J, Warner DC, Anstrom DC, Bensen RJ, Castiglioni PP, Donnarummo MG (2007) Plant nuclear factor Y (NF-Y) B subunits confer drought tolerance and lead to improved corn yields on water-limited acres. Proc Natl Acad Sci USA 104:16450–16455PubMedGoogle Scholar
  33. Oldfield AJ, Yang P, Conway AE, Cinghu S, Freudenberg JM, Yellaboina S, Jothi R (2014) Histone-fold domain protein NF-Y promotes chromatin accessibility for cell type-specific master transcription factors. Mol Cell 55:708–722PubMedPubMedCentralGoogle Scholar
  34. Quach TN, Nguyen HT, Valliyodan B, Joshi T, Xu D, Nguyen HT (2015) Genome-wide expression analysis of soybean NF-Y genes reveals potential function in development and drought response. Mol Genet Genom 290:1095–1115Google Scholar
  35. Riechmann JL, Heard J, Martin G, Reuber L, Jiang C, Keddie J, Adam L, Pineda O, Ratcliffe OJ, Samaha RR (2000) Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes. Science 290:2105–2110PubMedGoogle Scholar
  36. Rípodas C, Castaingts M, Clúa J, Blanco F, Zanetti ME (2015) Annotation, phylogeny and expression analysis of the nuclear factor Y gene families in common bean (Phaseolus vulgaris). Front Plant Sci 5:761PubMedPubMedCentralGoogle Scholar
  37. Ruan YL (2014) Sucrose metabolism: gateway to diverse carbon use and sugar signaling. Annu Rev Plant Biol 65:33–67PubMedGoogle Scholar
  38. Sato H, Mizoi J, Tanaka H, Maruyama K, Qin F, Osakabe Y, Morimoto K, Ohori T, Kusakabe K, Nagata M (2014) Arabidopsis DPB3-1, a DREB2A interactor, specifically enhances heat stress-induced gene expression by forming a heat stress-specific transcriptional complex with NF-Y subunits. Plant Cell 26:4954–4973PubMedPubMedCentralGoogle Scholar
  39. Sawamura M, Kuriyama T (1988) Quantitative determination of volatile constituents in the pummelo (Citrus grandis Osbeck forma Tosa-buntan). J Agric Food Chem 36:385–392Google Scholar
  40. Shi H, Ye T, Zhong B, Liu X, Jin R, Chan Z (2014) AtHAP5A modulates freezing stress resistance in Arabidopsis through binding to CCAAT motif of AtXTH21. New Phytol 9:554–567Google Scholar
  41. Siefers N, Dang KK, Kumimoto RW, Bynum WE, Iv Tayrose G, Holt BF (2009) Tissue-specific expression patterns of arabidopsis NF-Y transcription factors suggest potential for extensive combinatorial complexity. Plant Physiol 149:625–641PubMedPubMedCentralGoogle Scholar
  42. Sls P, Cps M, Sousa AO, Camillo LR, Araújo CP, Alcantara GM, Camargo DS, Cidade LC, de Almeida AF, Mgc C (2018) Genome-wide characterization and expression analysis of citrus NUCLEAR FACTOR-Y (NF-Y) transcription factors identified a novel NF-YA gene involved in drought-stress response and tolerance. PLoS One 13:6Google Scholar
  43. Stephenson TJ, Mcintyre CL, Collet C, Xue GP (2007) Genome-wide identification and expression analysis of the NF-Y family of transcription factors in Triticum aestivum. Plant Mol Biol 65:77–92PubMedGoogle Scholar
  44. Sturm A (1999) Invertases. Primary structures, functions, and roles in plant development and sucrose partitioning. Plant Physiol 121:1–8PubMedPubMedCentralGoogle Scholar
  45. Wang X, Xu Y, Zhang S, Li C, Huang Y, Cheng J, Wu G, Tian S, Chen C, Liu Y (2017) Genomic analyses of primitive, wild and cultivated citrus provide insights into asexual reproduction. Nat Genet 49:765–772PubMedGoogle Scholar
  46. Warpeha KM, Snehali U, Jennifer Y, Julia A, Hawkins SI, Lapik YR, Mary Beth A, Kaufman LS (2007) The GCR47, GPA1, PRN1, NF-Y signal chain mediates both blue light and abscisic acid responses in Arabidopsis. Plant Physiol 143:1590–1600PubMedPubMedCentralGoogle Scholar
  47. Waterhouse AM, Procter JB, Martin DM, Clamp M, Barton GJ (2009) Jalview Version 2—a multiple sequence alignment editor and analysis workbench. Bioinformatics 25:1189–1191PubMedPubMedCentralGoogle Scholar
  48. West M, Yee KM, Danao J, Zimmerman JL, Fischer RL, Goldberg RB, Harada JJ (1994) LEAFY COTYLEDON1 is an essential regulator of late embryogenesis and cotyledon identity in Arabidopsis. Plant Cell 6:1731–1745PubMedPubMedCentralGoogle Scholar
  49. Xing Y, Zhang S, Olesen JT, Rich A, Guarente L (1994) Subunit interaction in the CCAAT-binding heteromeric complex is mediated by a very short α-helix in HAP2. Proc Natl Acad Sci USA 91:3009–3013PubMedGoogle Scholar
  50. Xu Q, Chen LL, Ruan X, Chen D, Zhu A, Chen C, Bertrand D, Jiao WB, Hao BH, Lyon MP (2013) The draft genome of sweet orange (Citrus sinensis). Nat Genet 45:U59–U92Google Scholar
  51. Yan WH, Wang P, Chen HX, Zhou HJ, Li QP, Wang CR, Ding ZH, Zhang YS, Yu SB, Xing YZ (2011) A major QTL, Ghd8, plays pleiotropic roles in regulating grain productivity, plant height and heading date in rice. Mol Plant 4:319–330Google Scholar
  52. Yang W, Lu Z, Xiong Y, Yao J (2017) Genome-wide identification and co-expression network analysis of the Os NF-Y gene family in rice. Crop J 5:21–31Google Scholar
  53. Zanetti ME, Rípodas C, Niebel A (2017) Plant NF-Y transcription factors: key players in plant-microbe interactions, root development and adaptation to stress ☆. BBA Gene Regul Mech 1860:645Google Scholar
  54. Zemzoumi K, Frontini M, Bellorini M, Mantovani R (1999) NF-Y histone fold alpha1 helices help impart CCAAT specificity. J Mol Biol 286:327–337PubMedGoogle Scholar
  55. Zhang JJ, Xue HW (2013) OsLEC1/OsHAP3E participates in the determination of meristem identity in both vegetative and reproductive developments of rice. J Integr Plant Biol 55:232–249PubMedGoogle Scholar

Copyright information

© The Genetics Society of Korea 2019

Authors and Affiliations

  1. 1.Key Laboratory of Tropical Biological Resources of Ministry of Education, School of Life and PharmaceuticalHainan UniversityHaikouChina

Personalised recommendations