Abstract
Main conclusion
A novel Torenia phenotype having separate petals was obtained by the combination of NF-YA6-VP16 with a floral organ-specific promoter.
Abstract
Genetic engineering techniques helped in obtaining novel flower colors and shapes, in particular, by introducing functionally modified transcription factors (TFs) to ornamental flower species. Herein, we used functionally modified Arabidopsis TFs fused with the repression domain SRDX and the activation domain VP16 to screen for novel floral traits in Torenia fournieri Lind (torenia). We avoided undesired phenotypes unrelated to flowers by expressing these TFs through a floral organ-specific promoter belonging to the class-B genes, GLOBOSA (TfGLO). Fourteen constructs were produced to express functionally modified Arabidopsis TFs in which each of SRDX and VP16 was fused into 7 TFs that were used for the collective transformation of Torenia plants. Among the obtained transgenic plants, phenotypes with novel floral traits reflected in separate petals within normally gamopetalous flower lines. Sequencing analysis revealed that the transgenic plants contained nuclear factor-YA6 (NF-YA6) fused with the VP16. In the margin between the lips of the petals and tube in the TfGLOp:NF-YA6-VP16 plants, staminoid organs have been developed to separate petals. In the petals of the TfGLOp:NF-YA6-VP16 plants, the expression of a Torenia class C gene, PLENA (TfPLE), was found to be ectopically increased. Moreover, expression of TfPLE-VP16 under the control of the TfGLO promoter brought a similar staminoid phenotype observed in the TfGLOp:NF-YA6-VP16 plants. These results suggest that the introduction of the TfGLOp:NF-YA6-VP16 induced TfPLE expression, resulting in the formation of staminoid petals and separation of them.
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Abbreviations
- GLO:
-
GLOBOSA
- NF-Y:
-
Nuclear factor Y
- PLE:
-
PLENA
- SEM:
-
Scanning electron microscope
- TF:
-
Transcription factor
References
Aida R (2008) Torenia fournieri (torenia) as a model plant for transgenic studies. Plant Biotechnol 25:541–545. https://doi.org/10.5511/plantbiotechnology.25.541
An H, Roussot C, Suarez-Lopez P, Corbesier L, Vincent C, Pineiro M, Hepworth S, Mouradov A, Justin S, Turnbull C, Coupland G (2004) CONSTANS acts in the phloem to regulate a systemic signal that induces photoperiodic flowering of Arabidopsis. Development 131:3615–3626. https://doi.org/10.1242/dev.01231
Bartel DP (2009) MicroRNAs: target recognition and regulatory functions. Cell 136:215–233. https://doi.org/10.1016/j.cell.2009.01.002
Calvenzani V, Testoni B, Gusmaroli G, Lorenzo M, Gnesutta N, Petroni K, Mantovani R, Tonelli C (2012) Interactions and CCAAT-binding of Arabidopsis thaliana NF-Y subunits. PLoS ONE 7:e42902. https://doi.org/10.1371/journal.pone.0042902
Cartolano M, Castillo R, Efremova N, Kuckenberg M, Zethof J, Gerats T, Schwarz-Sommer Z, Vandenbussche M (2007) A conserved microRNA module exerts homeotic control over Petunia hybrida and Antirrhinum majus floral organ identity. Nat Genet 39:901–905. https://www.nature.com/articles/ng2056
Davies B, Motte P, Keck E, Saedler H, Sommer H, Schwarz-Sommer Z (1999) PLENA and FARINELLI: redundancy and regulatory interactions between two Antirrhinum MADS-box factors controlling flower development. EMBO J 15:4023–4034. https://doi.org/10.1093/emboj/18.14.4023
Fujiwara S, Sakamoto S, Kigoshi K, Suzuki K, Ohme-Takagi M (2014) VP16 fusion induces the multiple-knockout phenotype of redundant transcriptional repressors partly by Med25-independent mechanisms in Arabidopsis. FEBS Lett 588:3665–3672. https://doi.org/10.1016/j.febslet.2014.08.010
Gnesutta N, Kumimoto RW, Swain S, Chiara M, Siriwardana C, Horner DS, Holt BF, Mantovani R (2017) CONSTANS imparts DNA sequence specificity to the histone fold NF-YB/NF-YC dimer. Plant Cell 29:1516–1532. https://doi.org/10.1105/tpc.16.00864
Gnesutta N, Mantovani R, Fornara F (2018) Plant flowering: imposing DNA specificity on histone-fold subunits. Trends Plant Sci 23:293–301. https://doi.org/10.1016/j.tplants.2017.12.005
Gómez-Mena C, de Folter S, Costa MMR, Angenent GC, Sablowski R (2005) Transcriptional program controlled by the floral homeotic gene AGAMOUS during early organogenesis. Development 132:429–438. https://doi.org/10.1242/dev.01600
Gusmaroli G, Tonellia C, Mantovani R (2002) Regulation of the CCAAT-Binding NF-Y subunits in Arabidopsis thaliana. Gene 283:41–48. https://doi.org/10.1016/S0378-1119(01)00323-7
Hanano S, Goto K (2011) Arabidopsis TERMINAL FLOWER1 is involved in the regulation of flowering time and inflorescence development through transcriptional repression. Plant Cell 23:3172–3184. https://doi.org/10.1105/tpc.111.088641
Hiratsu K, Matsui K, Koyama T, Ohme-Takagi M (2003) Dominant repression of target genes by chimeric repressors that include the EAR motif, a repression domain, in Arabidopsis. Plant J 34:733–739. https://doi.org/10.1046/j.1365-313X.2003.01759.x
Hong RL, Hamaguchi L, Busch MA, Weigel D (2003) Regulatory elements of the floral homeotic gene AGAMOUS identified by phylogenetic footprinting and shadowing. Plant Cell 15:1296–1309. https://doi.org/10.1105/tpc.009548
Huijser P, Klein J, Lönnig WE, Meijer H, Saedler H, Sommer H (1992) Bracteomania, an inflorescence anomaly, is caused by the loss of function of the MADS-box gene squamosa in Antirrhinum majus. EMBO J 11:1239–1249. https://doi.org/10.1002/j.1460-2075.1992.tb05168.x
Jones-Rhoades MW, Bartel DP, Bartel B (2006) MicroRNAs and their regulatory roles in plants. Annu Rev Plant Biol 57:19–53. https://doi.org/10.1146/annurev.arplant.57.032905.105218
Kasajima I, Ohtsubo N, Sasaki K (2017) Combination of Cyclamen persicum Mill. floral gene promoters and chimeric repressors for the modification of ornamental traits in Torenia fournieri Lind. Hortic Res 4:17008. https://www.nature.com/articles/hortres20178
Koo SC, Bracko O, Park MS, Schwab R, Chun HJ, Park KM, Seo JS, Grbic V, Balasubramanian S, Schmid M, Godard F, Dae-Jin Y, Lee SY, Cho MJ, Weigel D, Kim MC (2010) Control of lateral organ development and flowering time by the Arabidopsis thaliana MADS-box Gene AGAMOUS-LIKE6. Plant J 62:807–816. https://doi.org/10.1111/j.1365-313X.2010.04192.x
Kosugi S, Ohashi Y, Nakajima K, Arai Y (1990) An improved assay for ß-glucuronidase in transformed cells: methanol almost completely suppresses a putative endogenous ß-glucuronidase activity. Plant Sci 70:133–140. https://doi.org/10.1016/0168-9452(90)90042-M
Koyama T, Mitsuda N, Seki M, Shinozaki K, Ohme-Takagi M (2010) TCP transcription factors regulate the activities of ASYMMETRIC LEAVES1 and miR164, as well as the auxin response, during differentiation of leaves in Arabidopsis. Plant Cell 22:3574–3588. https://doi.org/10.1105/tpc.110.075598
Laloum T, De Mita S, Gamas P, Baudin M, Niebel A (2013) CCAAT-box binding transcription factors in plants: Y so many? Trends Plant Sci 18:157–166. https://doi.org/10.1016/j.tplants.2012.07.004
Li XY, Hooft van Huijsduijnen R, Mantovani R, Benoist C, Mathis D (1992) Intron-exon organization of the NF-Y genes. Tissue-specific splicing modifies an activation domain. J Biol Chem 267:8984–8990. https://doi.org/10.1016/S0021-9258(19)50377-5
Mantovani R (1999) The molecular biology of the CCAAT-binding factor NF-Y. Gene 239:15–27. https://doi.org/10.1016/S0378-1119(99)00368-6
Marzougui S, Sugimoto K, Yamanouchi U, Shimono M, Hoshino T, Hori K, Kobayashi M, Ishiyama K, Yano M (2012) Mapping and characterization of seed dormancy QTLs using chromosome segment substitution lines in rice. Theor Appl Genet 124:893–902. https://link.springer.com/article/https://doi.org/10.1007/s00122-011-1753-y
Mitsuda N, Umemura Y, Ikeda M, Shikata M, Koyama T, Matsui K, Narumi T, Aida R, Sasaki K, Hiyama T et al (2008) FioreDB: a database of phenotypic information induced by the chimeric repressor silencing technology (CRES-T) in Arabidopsis and floricultural plants. Plant Biotechnol 25:37–44. https://doi.org/10.5511/plantbiotechnology.25.37
Mitsuda N, Takiguchi Y, Shikata M, Sage-Ono K, Ono M, Sasaki K, Yamaguchi H, Narumi T, Tanaka Y, Sugiyama M et al (2011b) The new FioreDB database provides comprehensive information on plant transcription factors and phenotypes induced by CRES-T in ornamental and model plants. Plant Biotechnol 28:123–130. https://doi.org/10.5511/plantbiotechnology.11.0106a
Mitsuda N, Matsui K, Ikeda M, Nakata M, Oshima Y, Nagatoshi Y, Ohme-Takagi M (2011a) CRES-T, an effective gene silencing system utilizing chimeric repressors. Methods Mol Biol 754:87–105. https://link.springer.com/protocol/https://doi.org/10.1007/978-1-61779-154-3_5
Mu J, Tan H, Hong S, Liang Y, 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 6:188–201. https://doi.org/10.1093/mp/sss061
Myers ZA, Holt BF III (2018) NUCLEAR FACTOR-Y: still complex after all these years? Curr Opin Plant Biol 45:96–102. https://doi.org/10.1016/j.pbi.2018.05.015
Nardini M, Gnesutta N, Donati G, Gatta R, Forni C, Fossati A, Vonrhein C, Moras C, Romier C, Bolognesi M, Mantovani R (2013) Sequence-specific transcription factor NF-Y displays histone-like DNA binding and H2B-like ubiquitination. Cell 152:132–143. https://doi.org/10.1016/j.cell.2012.11.047
Niki T, Hirai M, Niki T, Kanno A, Nishijima T (2012) Role of floral homeotic genes in the morphology of forchlorfenuron-induced paracorollas in Torenia fournieri Lind. J Jpn Soc Hort Sci 81:204–212. https://doi.org/10.2503/jjshs1.81.204
Nishijima T, Shima K (2006) Change in flower morphology of Torenia fournieri Lind. induced by forchlorfenuron application. Sci Hortic 109:254–261. https://doi.org/10.1016/j.scienta.2006.05.005
Notaguchi M, Abe M, Kimura T, Daimon Y, Kobayashi T, Yamaguchi A, Tomita Y, Dohi K, Mori M, Araki T (2008) Long-distance, graft-transmissible action of Arabidopsis FLOWERING LOCUS T protein to promote flowering. Plant Cell Physiol 49:1645–1658. https://doi.org/10.1093/pcp/pcn154
Ó’Maoiléidigh DS, Graciet E, Wellmer F (2014) Gene networks controlling Arabidopsis thaliana flower development. New Phytol 201:16–30. https://doi.org/10.1111/nph.12444
Petroni K, Kumimoto RW, Gnesutta N, Calvenzani V, Fornari M, Tonelli C, Holt BF III, Mantovani R (2012) The promiscuous life of plant NUCLEAR FACTOR Y transcription factors. Plant Cell 24:4777–4792. https://doi.org/10.1105/tpc.112.105734
Riechmann JL, Heard J, Martin G., Reuber L, Jiang C, Keddie J, Adam L, Pineda O, Ratcliffe OJ, Samaha RR, Creelman R, Pilgrim M, Broun P, Zhang JZ, Ghandehari D, Sherman BK, Yu G (2000) Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes. Science 290:2105–2110. https://science.sciencemag.org/content/290/5499/2105.abstract
Samad AFA, Sajad M, Nazaruddin N, Fauzi IA, Murad AMA, Zainal Z, Ismail I (2017) MicroRNA and transcription factor: key players in plant regulatory network. Front Plant Sci 8:565. https://doi.org/10.3389/fpls.2017.00565
Sasaki K (2018) Utilization of transcription factors for controlling floral morphogenesis in horticultural plants. Breeding Sci 68:88–98. https://doi.org/10.1270/jsbbs.17114
Sasaki K, Ohtsubo N (2020) Production of multi-petaled Torenia fournieri flowers by functional disruption of two class C MADS-box genes. Planta 251:101. https://doi.org/10.1007/s00425-020-03393-3
Sasaki K, Yamaguchi H, Kasajima I, Narumi T, Ohtsubo N (2016) Generation of novel floral traits using a combination of floral organ-specific promoters and a chimeric repressor in Torenia fournieri Lind. Plant Cell Physiol 57:1319–1331. https://doi.org/10.1093/pcp/pcw081
Scott MP (2000) Development: the natural history of genes. Cell 100:27–40. https://doi.org/10.1016/S0092-8674(00)81681-5
Shikata M, Narumi T, Yamaguchi H, Sasaki K, Aida R, Oshima Y, Takiguchi Y, Ohme-Takagi M, Mitsuda N, Ohtsubo N (2011) Efficient production of novel floral traits in torenia by collective transformation with chimeric repressors of Arabidopsis transcription factors. Plant Biotechnol 28:189–199. https://doi.org/10.5511/plantbiotechnology.10.1216a
Siefers N, Dang KK, Kumimoto RW, Bynum WET, Tayrose G, Holt BF III (2009) Tissue-specific expression patterns of Arabidopsis NF-Y transcription factors suggest potential for extensive combinatorial complexity. Plant Physiol 149:625–641. https://doi.org/10.1104/pp.108.130591
Siriwardana CL, Gnesutta N, Kumimoto RW, Jones DS, Myers ZA, Mantovani R, Holt BF III (2016) NUCLEAR FACTOR Y, subunit A (NF-YA) proteins positively regulate flowering and act through FLOWERING LOCUS T. PLoS Genet 12:e1006496. https://doi.org/10.1371/journal.pgen.1006496
Soltis DE, Ma H, Frohlich MW, Soltis PS, Albert VA, Oppenheimer DG, Altman NS, dePamphilis C, Leebens-Mack J (2007) The floral genome: an evolutionary history of gene duplication and shifting patterns of gene expression. Trends Plant Sci 12:358–367. https://doi.org/10.1016/j.tplants.2007.06.012
Tasaki K, Nakatsuka A, Cheon KS, Kobayashi N (2014) Expression of MADS-box genes in narrow-petaled cultivars of Rhododendron macrosepalum Maxim. J Jpn Soc Hort Sci 83:52–58. https://doi.org/10.2503/jjshs1.CH-030
Theißen G, Melzer R, Rümpler F (2016) MADS-domain transcription factors and the floral quartet model of flower development: linking plant development and evolution. Development 143:3259–3271. https://dev.biologists.org/content/143/18/3259.long
Triezenberg SJ, Kingsbury RC, McKnight SL (1988) Functional dissection of VP16, the trans-activator of herpes simplex virus immediate early gene expression. Genes Dev 2:718–729. http://genesdev.cshlp.org/content/2/6/718.short
Yamaguchi H, Sasaki K, Shikata M, Aida R, Ohtsubo N (2011) Trehalose drastically extends the in vitro vegetative culture period and facilitates maintenance of Torenia fournieri plants. Plant Biotechnol 28:263–266. https://doi.org/10.5511/plantbiotechnology.11.0124c
Zhao H, Wu D, Kong F, Lin K, Zhang H, Li G (2017) The Arabidopsis thaliana Nuclear Factor Y transcription factors. Front Plant Sci 7:2045. https://doi.org/10.3389/fpls.2016.02045
Acknowledgements
We thank Ms. Miyuki Tsuruoka, Ms. Yuko Namekawa, and Ms. Ayumi Takimoto for maintaining the transgenic plants, and Dr. Mitsuko Kishi-Kaboshi for helpful advices.
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Sekiguchi, N., Sasaki, K., Oshima, Y. et al. Ectopic expression of AtNF-YA6-VP16 in petals results in a novel petal phenotype in Torenia fournieri. Planta 255, 105 (2022). https://doi.org/10.1007/s00425-022-03876-5
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DOI: https://doi.org/10.1007/s00425-022-03876-5