REVEILLE Transcription Factors Contribute to the Nighttime Accumulation of Anthocyanins in ‘Red Zaosu’ (Pyrus Bretschneideri Rehd.) Pear Fruit Skin
Abstract
:1. Introduction
2. Results
2.1. The Anthocyanin Content Oscillated Diurnally and Mainly Increased over Night in ‘Red Zaosu’ Fruit Skin
2.2. The Expression Patterns of Candidate REVEILLE (RVE) TFs Correlated with the Nighttime Increase in the Anthocyanin Level in ‘Red Zaosu’ Fruit Skin
2.3. Overexpression of PbRVEs in ‘Zaosu’ Pear Fruit Promoted Anthocyanin Accumulation
2.4. PbRVE1a, 1b, 7 and 8 Promoted Anthocyanin Accumulation by Activating the Promoters of ALBGs in ‘Red Zaosu’ Pear Fruit
3. Discussion
3.1. PbRVE1a, 1b, 7 and 8 Expression Levels Correlated with Anthocyanin Accumulation during the Nighttime in ‘Red Zaosu’ Pear Fruit Skin
3.2. PbRVEs Promoted Anthocyanin Accumulation by Up-Regulating the Expression Levels of PbDFR and PbANS in Pear Fruit Skin
4. Materials and Methods
4.1. Plant Material and Treatments
4.2. Anthocyanin Content Measurements
4.3. Isolation of RVE Genes and Their Phylogenetic Analysis
4.4. RNA Isolation and an Expression Analysis Using Quantitative Real-Time PCR (qRT-PCR)
4.5. Transient Expression Assay in Pear Fruitlet Skins
4.6. Y1H Assay
4.7. Dual-Luciferase Assay
4.8. Statistical Analysis
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Abbreviations
ANS | Anthocyanidin synthase |
ALBGs | Anthocyanin late biosynthetic genes |
CDS | Coding DNA sequence |
DFR | Dihydroflavonol 4-reductase |
GUS | the β-glucuronidase gene |
HAS | Hours after sunrise of day 1 |
LUC | Firefly luciferase |
MCS | Multiple cloning sites |
MBW | MYB–bHLH–WDR |
OE | Overexpression |
qRT-PCR | Quantitative Real-Time PCR |
REN | Renilla luciferase |
RVE | REVEILLE |
SD | Selective synthetic dextrose medium |
SEs | the standard errors |
TF | Transcription factor |
UFGT | UDP-glucoside: flavonoid glucosyltransferase |
VIGS | Virus-induced gene silencing |
Y1H | Yeast one-hybrid assay |
References
- Shin, D.H.; Choi, M.; Kim, K.; Bang, G.; Cho, M.; Choi, S.B.; Choi, G.; Park, Y.I. HY5 regulates anthocyanin biosynthesis by inducing the transcriptional activation of the MYB75/PAP1 transcription factor in Arabidopsis. FEBS Lett. 2013, 587, 1543–1547. [Google Scholar] [CrossRef] [Green Version]
- Liu, Y.; Tikunov, Y.; Schouten, R.E.; Marcelis, L.F.M.; Visser, R.G.F.; Bovy, A. Anthocyanin Biosynthesis and Degradation Mechanisms in Solanaceous Vegetables: A Review. Front. Chem. 2018, 6, 52. [Google Scholar] [CrossRef]
- Sarma, A.D.; Sharma, R. Anthocyanin-DNA copigmentation complex: Mutual protection against oxidative damage. Phytochemistry 1999, 52, 1313–1318. [Google Scholar] [CrossRef]
- Tanaka, Y.; Sasaki, N.; Ohmiya, A. Biosynthesis of plant pigments: Anthocyanins, betalains and carotenoids. Plant J. 2008, 54, 733–749. [Google Scholar] [CrossRef]
- Shang, Y.; Venail, J.; Mackay, S.; Bailey, P.C.; Schwinn, K.E.; Jameson, P.E.; Martin, C.R.; Davies, K.M. The molecular basis for venation patterning of pigmentation and its effect on pollinator attraction in flowers of Antirrhinum. New Phytol. 2011, 189, 602–615. [Google Scholar] [CrossRef]
- Petroni, K.; Pilu, R.; Tonelli, C. Anthocyanins in corn: A wealth of genes for human health. Planta 2014, 240, 901–911. [Google Scholar] [CrossRef]
- Li, D.; Wang, P.; Luo, Y.; Zhao, M.; Chen, F. Health benefits of anthocyanins and molecular mechanisms: Update from recent decade. Crit. Rev. Food Sci. Nutr. 2015, 57, 1729–1741. [Google Scholar] [CrossRef]
- Wang, L.S.; Stoner, G.D. Anthocyanins and their role in cancer prevention. Cancer Lett. 2008, 269, 281–290. [Google Scholar] [CrossRef] [Green Version]
- De Pascual-Teresa, S. Molecular mechanisms involved in the cardiovascular and neuroprotective effects of anthocyanins. Arch. Biochem. Biophys. 2014, 559, 68–74. [Google Scholar] [CrossRef] [Green Version]
- Sakuta, M. Diversity in plant red pigments: Anthocyanins and betacyanins. Plant Biotechnol. Rep. 2013, 8, 37–48. [Google Scholar] [CrossRef]
- Xu, W.; Dubos, C.; Lepiniec, L. Transcriptional control of flavonoid biosynthesis by MYB-bHLH-WDR complexes. Trends Plant Sci. 2015, 20, 176–185. [Google Scholar] [CrossRef] [PubMed]
- Lin-Wang, K.; Micheletti, D.; Palmer, J.; Volz, R.; Lozano, L.; Espley, R.; Hellens, R.P.; Chagne, D.; Rowan, D.D.; Troggio, M.; et al. High temperature reduces apple fruit colour via modulation of the anthocyanin regulatory complex. Plant Cell Environ. 2011, 34, 1176–1190. [Google Scholar] [CrossRef] [PubMed]
- Jaakola, L. New insights into the regulation of anthocyanin biosynthesis in fruits. Trends Plant Sci. 2013, 18, 477–483. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Xu, W.; Grain, D.; Bobet, S.; Le Gourrierec, J.; Thevenin, J.; Kelemen, Z.; Lepiniec, L.; Dubos, C. Complexity and robustness of the flavonoid transcriptional regulatory network revealed by comprehensive analyses of MYB-bHLH-WDR complexes and their targets in Arabidopsis seed. New Phytol. 2014, 202, 132–144. [Google Scholar] [CrossRef]
- Katia, P.; Chiara, T. Recent advances on the regulation of anthocyanin synthesis in reproductive organs. Plant Sci. 2011, 181, 219–229. [Google Scholar]
- Gonzalez, A.; Zhao, M.; Leavitt, J.M.; Lloyd, A.M. Regulation of the anthocyanin biosynthetic pathway by the TTG1/bHLH/Myb transcriptional complex in Arabidopsis seedlings. Plant J. 2008, 53, 814–827. [Google Scholar] [CrossRef]
- Maier, A.; Schrader, A.; Kokkelink, L.; Falke, C.; Welter, B.; Iniesto, E.; Rubio, V.; Uhrig, J.F.; Hulskamp, M.; Hoecker, U. Light and the E3 ubiquitin ligase COP1/SPA control the protein stability of the MYB transcription factors PAP1 and PAP2 involved in anthocyanin accumulation in Arabidopsis. Plant J. 2013, 74, 638–651. [Google Scholar] [CrossRef]
- Xie, X.B.; Li, S.; Zhang, R.F.; Zhao, J.; Chen, Y.C.; Zhao, Q.; Yao, Y.X.; You, C.X.; Zhang, X.S.; Hao, Y.J. The bHLH transcription factor MdbHLH3 promotes anthocyanin accumulation and fruit colouration in response to low temperature in apples. Plant Cell Environ. 2012, 35, 1884–1897. [Google Scholar] [CrossRef]
- Xie, Y.; Chen, P.; Yan, Y.; Bao, C.; Li, X.; Wang, L.; Shen, X.; Li, H.; Liu, X.; Niu, C.; et al. An atypical R2R3 MYB transcription factor increases cold hardiness by CBF-dependent and CBF-independent pathways in apple. New Phytol. 2018, 218, 201–218. [Google Scholar] [CrossRef]
- Zhai, R.; Wang, Z.; Zhang, S.; Meng, G.; Song, L.; Wang, Z.; Li, P.; Ma, F.; Xu, L. Two MYB transcription factors regulate flavonoid biosynthesis in pear fruit (Pyrus bretschneideri Rehd.). J. Exp. Bot. 2016, 67, 1275–1284. [Google Scholar] [CrossRef] [Green Version]
- Feng, S.; Wang, Y.; Yang, S.; Xu, Y.; Chen, X. Anthocyanin biosynthesis in pears is regulated by a R2R3-MYB transcription factor PyMYB10. Planta 2010, 232, 245–255. [Google Scholar] [CrossRef] [PubMed]
- Wu, J.; Wang, Z.; Shi, Z.; Zhang, S.; Ming, R.; Zhu, S.; Khan, M.A.; Tao, S.; Korban, S.S.; Wang, H.; et al. The genome of the pear (Pyrus bretschneideri Rehd.). Genome Res. 2013, 23, 396–408. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Harmer, S.L. The circadian system in higher plants. Annu. Rev. Plant Biol. 2009, 60, 357–377. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Deikman, J.; Hammer, P.E. Induction of Anthocyanin Accumulation by Cytokinins in Arabidopsis thaliana. Plant Physiol. 1995, 108, 47–57. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Dubos, C.; Le Gourrierec, J.; Baudry, A.; Huep, G.; Lanet, E.; Debeaujon, I.; Routaboul, J.M.; Alboresi, A.; Weisshaar, B.; Lepiniec, L. MYBL2 is a new regulator of flavonoid biosynthesis in Arabidopsis thaliana. Plant J. 2008, 55, 940–953. [Google Scholar] [CrossRef]
- Nguyen, N.H.; Jeong, C.Y.; Kang, G.H.; Yoo, S.D.; Hong, S.W.; Lee, H. MYBD employed by HY5 increases anthocyanin accumulation via repression of MYBL2 in Arabidopsis. Plant J. 2015, 84, 1192–1205. [Google Scholar] [CrossRef] [Green Version]
- Perez-Garcia, P.; Ma, Y.; Yanovsky, M.J.; Mas, P. Time-dependent sequestration of RVE8 by LNK proteins shapes the diurnal oscillation of anthocyanin biosynthesis. Proc. Natl. Acad. Sci. USA 2015, 112, 5249–5253. [Google Scholar] [CrossRef] [Green Version]
- Espley, R.V.; Hellens, R.P.; Putterill, J.; Stevenson, D.E.; Kutty-Amma, S.; Allan, A.C. Red colouration in apple fruit is due to the activity of the MYB transcription factor, MdMYB10. Plant J. 2007, 49, 414–427. [Google Scholar] [CrossRef] [Green Version]
- Li, K.T.; Zhang, J.; Kang, Y.H.; Chen, M.C.; Song, T.T.; Geng, H.; Tian, J.; Yao, Y.C. McMYB10 Modulates the Expression of a Ubiquitin Ligase, McCOP1 During Leaf Coloration in Crabapple. Front. Plant Sci. 2018, 9, 704. [Google Scholar] [CrossRef] [Green Version]
- Bai, S.; Tao, R.; Tang, Y.; Yin, L.; Ma, Y.; Ni, J.; Yan, X.; Yang, Q.; Wu, Z.; Zeng, Y.; et al. BBX16, a B-box protein, positively regulates light-induced anthocyanin accumulation by activating MYB10 in red pear. Plant Biotechnol. J. 2019, 17, 1985–1997. [Google Scholar] [CrossRef] [Green Version]
- Yao, G.; Ming, M.; Allan, A.C.; Gu, C.; Li, L.; Wu, X.; Wang, R.; Chang, Y.; Qi, K.; Zhang, S.; et al. Map-based cloning of the pear gene MYB114 identifies an interaction with other transcription factors to coordinately regulate fruit anthocyanin biosynthesis. Plant J. 2017, 92, 437–451. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Nguyen, N.H.; Lee, H. MYB-related transcription factors function as regulators of the circadian clock and anthocyanin biosynthesis in Arabidopsis. Plant Signal. Behav. 2016, 11, e1139278. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Xing, H.; Wang, P.; Cui, X.; Zhang, C.; Wang, L.; Liu, X.; Yuan, L.; Li, Y.; Xie, Q.; Xu, X. LNK1 and LNK2 recruitment to the evening element require morning expressed circadian related MYB-like transcription factors. Plant Signal. Behav. 2015, 10, e1010888. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Wang, Z.-Y.; Kenigsbuch, D.; Sun, L.; Harel, E.; Ong, M.S.; Tobin, E.M. A Myb-related transcription factor is involved in the phytochrome regulation of an Arabidopsis Lhcb gene. Plant Cell 1997, 9, 491–507. [Google Scholar]
- Andronis, C.; Barak, S.; Knowles, S.M.; Sugano, S.; Tobin, E.M. The clock protein CCA1 and the bZIP transcription factor HY5 physically interact to regulate gene expression in Arabidopsis. Mol. Plant 2008, 1, 58–67. [Google Scholar] [CrossRef] [Green Version]
- Zhai, R.; Liu, X.T.; Feng, W.T.; Chen, S.S.; Xu, L.F.; Wang, Z.G.; Zhang, J.L.; Li, P.M.; Ma, F.W. Different biosynthesis patterns among flavonoid 3-glycosides with distinct effects on accumulation of other flavonoid metabolites in pears (Pyrus bretschneideri Rehd.). PLoS ONE. 2014, 9, e91945. [Google Scholar] [CrossRef]
- Wolfe, K.L.; Liu, R.H. Apple peels as a value-added food ingredient. J. Agric. Food Chem. 2003, 51, 1676–1683. [Google Scholar] [CrossRef]
- Giusti, M.M.; Wrolstad, R.E. Characterization and Measurement of Anthocyanins by UV-Visible Spectroscopy. Curr. Protoc. Food Anal. Chem. 2001, F1–F2. [Google Scholar] [CrossRef]
- Wang, Z.; Du, H.; Zhai, R.; Song, L.; Ma, F.; Xu, L. Transcriptome Analysis Reveals Candidate Genes Related to Color Fading of ‘Red Bartlett’ (Pyrus communis L.). Front. Plant Sci. 2017, 8, 455. [Google Scholar] [CrossRef] [Green Version]
- Hellens, R.P.; Allan, A.C.; Friel, E.N.; Bolitho, K.; Grafton, K.; Templeton, M.D.; Karunairetnam, S.; Gleave, A.P.; Laing, W.A. Transient expression vectors for functional genomics, quantification of promoter activity and RNA silencing in plants. Plant Methods 2005, 1, 13. [Google Scholar] [CrossRef] [Green Version]
- Spolaore, S.; Trainotti, L.; Casadoro, G. A simple protocol for transient gene expression in ripe fleshy fruit mediated by Agrobacterium. J. Exp. Bot. 2001, 52, 845–850. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Fillatti, J.J.; Kiser, J.; Rose, R.; Comai, L. Efficient transfer of a glyphosate tolerance gene into tomato using a binary Agrobacterium tumefaciens vector. Biotechnology 1987, 5, 726–730. [Google Scholar] [CrossRef]
Correlation Coefficients (Pearson’s) | ||||||||
---|---|---|---|---|---|---|---|---|
Genes | PbMYB9 | PbMYB10 | PbMYB10b | PbbHLH33a | PbRVE1a | PbRVE1b | PbRVE7 | PbRVE8 |
PbANS | 0.441 ** | 0.22 | −0.01 | 0.07 | 0.750 ** | 0.775 ** | 0.728 ** | 0.757 ** |
PbDFR | 0.416 ** | 0.17 | −0.02 | 0.08 | 0.734 ** | 0.782 ** | 0.746 ** | 0.777 ** |
PbUFGT | 0.355 * | 0.335 * | 0.19 | 0.06 | 0.779 ** | 0.859 ** | 0.818 ** | 0.855 ** |
© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
Share and Cite
Li, X.; Wu, T.; Liu, H.; Zhai, R.; Wen, Y.; Shi, Q.; Yang, C.; Wang, Z.; Ma, F.; Xu, L. REVEILLE Transcription Factors Contribute to the Nighttime Accumulation of Anthocyanins in ‘Red Zaosu’ (Pyrus Bretschneideri Rehd.) Pear Fruit Skin. Int. J. Mol. Sci. 2020, 21, 1634. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21051634
Li X, Wu T, Liu H, Zhai R, Wen Y, Shi Q, Yang C, Wang Z, Ma F, Xu L. REVEILLE Transcription Factors Contribute to the Nighttime Accumulation of Anthocyanins in ‘Red Zaosu’ (Pyrus Bretschneideri Rehd.) Pear Fruit Skin. International Journal of Molecular Sciences. 2020; 21(5):1634. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21051634
Chicago/Turabian StyleLi, Xieyu, Ting Wu, Hanting Liu, Rui Zhai, Yao Wen, Qianrong Shi, Chengquan Yang, Zhigang Wang, Fengwang Ma, and Lingfei Xu. 2020. "REVEILLE Transcription Factors Contribute to the Nighttime Accumulation of Anthocyanins in ‘Red Zaosu’ (Pyrus Bretschneideri Rehd.) Pear Fruit Skin" International Journal of Molecular Sciences 21, no. 5: 1634. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21051634