The Carboxyl-Terminus of TRANSPARENT TESTA GLABRA1 Is Critical for Its Functions in Arabidopsis
Abstract
:1. Introduction
2. Results
2.1. Phenotypes in the ttg1δ15aa Mutant Are Identical to That in A TTG1 Loss-of-Function Mutant
2.2. A Single Nucleotide Substitution Led to the Production of A Truncated TTG1 Protein in the ttg1δ15aa Mutant
2.3. Expression of TTG1 under the Control of its Native Promoter Restored the Ttg1δ15aa Mutant Phenotypes
2.4. TTG1Δ15aa Is Unable to Interact with GL3 and with Regulated Downstream Target Genes
2.5. Deletion of the Last Three C-Terminalamino Acids Causes Loss-of-Function of TTG1
3. Discussion
4. Materials and Methods
4.1. Plant Materials and Growth Conditions
4.2. RNA Isolation and qRT-PCR
4.3. Constructs
4.4. Plant Transformation and Transgenic Plants Selection
4.5. DNA Isolation and PCR
4.6. Plasmid DNA Isolation, Protoplast Isolation and Transfection
4.7. Mucilage Production Assays
4.8. Anthocyanin Biosynthesis Assays
4.9. Trichome and Root Hair Formation Assays
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
- Walker, A.R.; Davison, P.A.; Bolognesiwinfield, A.C.; James, C.M.; Srinivasan, N.; Blundell, T.L.; Esch, J.J.; Marks, M.D.; Gray, J.C. The TRANSPARENT TESTA GLABRA1 locus, which regulates trichome differentiation and anthocyanin biosynthesis in Arabidopsis, encodes a WD40 repeat protein. Plant Cell 1999, 11, 1337–1349. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Tian, H.; Wang, S. TRANSPARENT TESTA GLABRA1, a key regulator in plants with multiple roles and multiple function mechanisms. Int. J. Mol. Sci. 2020, 21, 4881. [Google Scholar] [CrossRef]
- Koornneef, M. The complex syndrome of ttg mutants. Arabid Inf. Serv. 1981, 18, 45–51. [Google Scholar]
- Galway, M.E.; Masucci, J.D.; Lloyd, A.M.; Walbot, V.; Davis, R.W.; Schiefelbein, J.W. The TTG gene is required to specify epidermal cell fate and cell patterning in the Arabidopsis root. Dev. Biol. 1994, 166, 740–754. [Google Scholar] [CrossRef] [PubMed]
- Shirley, B.W.; Kubasek, W.L.; Storz, G.; Bruggemann, E.; Koornneef, M.; Ausubel, F.M.; Goodman, H.M. Analysis of Arabidopsis mutants deficient in flavonoid biosynthesis. Plant J. 1995, 8, 659–671. [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]
- Chen, M.; Zhang, B.; Li, C.; Kulaveerasingam, H.; Chew, F.T.; Yu, H. TRANSPARENT TESTA GLABRA1 regulates the accumulation of seed storage reserves in Arabidopsis. Plant Physiol. 2015, 169, 391–402. [Google Scholar] [CrossRef] [Green Version]
- Li, C.; Zhang, B.; Chen, B.; Ji, L.; Hao, Y. Site-specifific phosphorylation of TRANSPARENT TESTA GLABRA1 mediates carbon partitioning in Arabidopsis seeds. Nat. Commun. 2018, 9, 571. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Paffendorf, B.A.; Qassrawi, R.; Meys, A.M.; Trimborn, L.; Schrader, A. TRANSPARENT TESTA GLABRA 1 participates in flowering time regulation in Arabidopsis thaliana. Peer J. 2020, 8, e8303. [Google Scholar] [CrossRef] [Green Version]
- Yuan, F.; Leng, B.; Zhang, H.; Wang, X.; Han, G.; Wang, B. A WD40-repeat protein from the Recretohalophyte Limonium bicolor enhances trichome formation and salt tolerance in Arabidopsis. Front. Plant Sci. 2019, 10, 1456. [Google Scholar] [CrossRef]
- Kong, D.; Li, M.; Dong, Z.; Ji, H.; Li, X. Identification of TaWD40D, a wheat WD40 repeat-containing protein that is associated with plant tolerance to abiotic stresses. Plant Cell Rep. 2015, 34, 395–410. [Google Scholar] [CrossRef]
- Wang, Y.; Liu, R.; Chen, L.; Wang, Y.; Liang, Y.; Wu, X.; Li, B.; Wu, J.; Liang, Y.; Wang, X.; et al. Nicotiana tabacum TTG1 contributes to ParA1-induced signalling and cell death in leaf trichomes. J. Cell Sci. 2009, 122, 2673–2685. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Van Nocker, S.; Ludwig, P. The WD-repeat protein superfamily in Arabidopsis: Conservation and divergence in structure and function. BMC Genomics 2003, 4, 50. [Google Scholar] [CrossRef] [Green Version]
- Ouyang, Y.; Huang, X.; Lu, Z.; Yao, J. Genomic survey, expression profile and co-expression network analysis of OsWD40 family in rice. BMC Genomics 2012, 13, 100. [Google Scholar] [CrossRef] [Green Version]
- Neer, E.J.; Schmidt, C.J.; Nambudripad, R.; Smith, T.F. The ancient regulatory protein family of WD-repeat proteins. Nature 1994, 371, 297–300. [Google Scholar] [CrossRef]
- Chen, S.; Wang, S. GLABRA2, a common regulator for epidermal cell fate determination and anthocyanin biosynthesis in Arabidopsis. Int. J. Mol. Sci. 2019, 20, 4997. [Google Scholar] [CrossRef] [Green Version]
- Schiefelbein, J. Cell-fate specification in the epidermis: A common patterning mechanism in the root and shoot. Curr. Opin. Plant Biol. 2003, 6, 74–78. [Google Scholar] [CrossRef]
- Lin, Q.; Aoyama, T. Pathways for epidermal cell differentiation via the homeobox gene GLABRA2: Update on the roles of the classic regulator. J. Integr. Plant Biol. 2012, 54, 729–737. [Google Scholar]
- Wang, S.; Chen, J.G. Regulation of cell fate determination by single-repeat R3 MYB transcription factors in Arabidopsis. Front. Plant Sci. 2014, 5, 133. [Google Scholar] [CrossRef] [Green Version]
- Golz, J.F.; Allen, P.J.; Li, S.F.; Parish, R.W.; Jayawardana, N.; Bacic, A.; Doblin, M.S. Layers of regulation—Insights into the role of transcription factors controlling mucilage production in the Arabidopsis thaliana seed coat. Plant Sci. 2018, 272, 179–192. [Google Scholar] [CrossRef]
- Pesch, M.; Hulskamp, M. Creating a two-dimensional pattern de novo during Arabidopsis trichome and root hair initiation. Curr. Opin. Genet. Dev. 2004, 14, 422–427. [Google Scholar] [CrossRef] [PubMed]
- Schellmann, S.; Schnittger, A.; Kirik, V.; Wada, T.; Okada, K.; Beermann, A.; Thumfahrt, J.; Jürgens, G.; Hulskamp, M. TRIPTYCHON and CAPRICE mediate lateral inhibition during trichome and root hair patterning in Arabidopsis. EMBO J. 2002, 21, 5036–5046. [Google Scholar] [CrossRef] [Green Version]
- Masucci, J.D.; Rerie, W.G.; Foreman, D.R.; Zhang, M.; Galway, M.E.; Marks, M.D.; Schiefelbein, J. The homeobox gene GLABRA2 is required for position-dependent cell differentiation in the root epidermis of Arabidopsis thaliana. Development 1996, 122, 1253–1260. [Google Scholar] [CrossRef]
- Schiefelbein, J.; Huang, L.; Zheng, X. Regulation of epidermal cell fate in Arabidopsis roots: The importance of multiple feedback loops. Front. Plant Sci. 2014, 5, 47. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Baudry, A.; Heim, M.A.; Debreucq, B.; Caboche, M.; Weisshaar, B.; Lepiniec, L. TT2, TT8, and TTG1 synergistically specify the expression of BANYULS and proanthocyanidin biosynthesis in Arabidopsis thaliana. Plant J. 2004, 39, 366–380. [Google Scholar] [CrossRef]
- Zhang, F.; Gonzalez, A.; Zhao, M.; Payne, C.T.; Lloyd, A. A network of redundant bHLH proteins functions in all TTG1-dependent pathways of Arabidopsis. Development 2003, 130, 4859–4869. [Google Scholar] [CrossRef] [Green Version]
- Nesi, N.; Debeaujon, I.; Jond, C.; Pelletier, G.; Caboche, M.; Lepiniec, L. The TT8 gene encodes a basic helix-loop-helix domain protein required for expression of DFR and BAN genes in Arabidopsis siliques. Plant Cell 2000, 12, 1863–1878. [Google Scholar] [CrossRef] [Green Version]
- 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]
- Petroni, K.; Tonelli, C. Recent advances on the regulation of anthocyanin synthesis in reproductive Organs. Plant Sci. 2011, 181, 219–229. [Google Scholar] [CrossRef]
- Xu, W.; Grain, D.; Bobet, S.; Le Gourrierec, J.; Thévenin, 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]
- Deng, Y.; Lu, S. Biosynthesis and regulation of phenylpropanoids in plants. Crit. Rev. Plant Sci. 2017, 36, 1–34. [Google Scholar] [CrossRef]
- Francoz, E.; Ranocha, P.; Burlat, V.; Dunand, C. Arabidopsis seed mucilage secretory cells: Regulation and dynamics. Trends Plant Sci. 2015, 20, 515–524. [Google Scholar] [CrossRef] [PubMed]
- Voiniciuc, C.; Yang, B.; Schmidt, M.H.; Günl, M.; Usadel, B. Starting to gel: How Arabidopsis seed coat epidermal cells produce specialized secondary cell walls. Int. J. Mol. Sci. 2015, 16, 3452–3473. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Lloyd, A.; Brockman, A.; Aguirre, L.; Campbell, A.; Bean, A.; Cantero, A.; Gonzalez, A. Advances in the MYB-bHLH-WD repeat (MBW) pigment regulatory model: Addition of a WRKY factor and co-option of an anthocyanin MYB for betalain regulation. Plant Cell Physiol. 2017, 58, 1431–1441. [Google Scholar] [CrossRef] [Green Version]
- Johnson, C.S.; Kolevski, B.; Smyth, D.R. TRANSPARENT TESTA GLABRA2, a trichome and seed coat development gene of Arabidopsis, encodes a WRKY transcription factor. Plant Cell. 2002, 14, 1359–1375. [Google Scholar] [CrossRef] [Green Version]
- Western, T.L.; Young, D.S.; Dean, G.H.; Tan, W.L.; Samuels, A.L.; Haughn, G.W. MUCILAGE-MODIFIED4 encodes a putative pectin biosynthetic enzyme developmentally regulated by APETALA2, TRANSPARENT TESTA GLABRA1, and GLABRA2 in the Arabidopsis seed coat. Plant Physiol. 2004, 134, 296–306. [Google Scholar] [CrossRef] [Green Version]
- Wang, S.; Hubbard, L.; Chang, Y.; Guo, J.; Schiefelbein, J.; Chen, J.G. Comprehensive analysis of single-repeat R3 MYB proteins in epidermal cell patterning and their transcriptional regulation in Arabidopsis. BMC Plant Biol. 2008, 8, 81. [Google Scholar] [CrossRef] [Green Version]
- Miller, J.C.; Chezem, W.R.; Clay, N.K. Ternary WD40 repeat-containing protein complexes: Evolution, composition and roles in plant immunity. Front. Plant Sci. 2015, 6, 1108. [Google Scholar] [CrossRef] [Green Version]
- Payne, C.T.; Zhang, F.; Lloyd, A.M. GL3 encodes a bHLH protein that regulates trichome development in Arabidopsis through interaction with GL1 and TTG1. Genetics 2000, 156, 1349–1362. [Google Scholar] [CrossRef]
- Wang, S.; Chen, J.G. Arabidopsis transient expression analysis reveals that activation of GLABRA2 may require concurrent binding of GLABRA1 and GLABRA3 to the promoter of GLABRA2. Plant Cell Physiol. 2008, 49, 1792–1804. [Google Scholar] [CrossRef] [Green Version]
- Zimmermann, I.M.; Heim, M.A.; Weisshaar, B.; Uhrig, J.F. Comprehensive identification of Arabidopsis thaliana MYB transcription factors interacting with R/B-like BHLH proteins. Plant J. Cell Mol. Biol. 2004, 40, 22–34. [Google Scholar] [CrossRef]
- Long, Y.; Schiefelbein, J. Novel TTG1 mutants modify root-hair pattern formation in Arabidopsis. Front. Plant Sci. 2020, 11, 383. [Google Scholar] [CrossRef]
- Wang, X.; Wang, X.; Hu, Q.; Dai, X.; Tian, H.; Zheng, K.; Wang, X.; Mao, T.; Chen, J.G.; Wang, S. Characterization of an activation-tagged mutant uncovers a role of GLABRA2 in anthocyanin biosynthesis in Arabidopsis. Plant J. 2015, 83, 300–311. [Google Scholar] [CrossRef]
- Larkin, J.C.; Oppenheimer, D.G.; Lloyd, A.M.; Paparozzi, E.T.; Marks, M.D. Roles of the GLABROUS1 and TRANSPARENT TESTA GLABRA genes in Arabidopsis trichome development. Plant Cell 1994, 6, 1065–1076. [Google Scholar] [CrossRef]
- Larkin, J.C.; Walker, J.D.; Bolognesi-Winfifield, A.C.; Gray, J.C.; Walker, A.R. Allele-specifific interactions between ttg and gl1 during trichome development in Arabidopsis thaliana. Genetics 1999, 151, 1591–1604. [Google Scholar] [CrossRef]
- Yoshida, Y.; Sano, R.; Wada, T.; Takabayashi, J.; Okada, K. Jasmonic acid control of GLABRA3 links inducible defense and trichome patterning in Arabidopsis. Development 2009, 136, 1039–1048. [Google Scholar] [CrossRef] [Green Version]
- Bharti, A.K.; Khurana, J.P. Molecular characterization of transparent testa (tt) mutants of Arabidopsis thaliana(ecotype Estland) impaired in flavonoid biosynthetic pathway. Plant Sci. 2003, 165, 1321–1332. [Google Scholar] [CrossRef]
- Tian, H.; Wang, X.; Guo, H.; Cheng, Y.; Hou, C.; Chen, J.G.; Wang, S. NTL8 regulates trichome formation in Arabidopsis by directly activating R3 MYB genes TRY and TCL1. Plant Physiol. 2017, 174, 2363–2375. [Google Scholar] [CrossRef] [Green Version]
- Zheng, K.; Tian, H.; Hu, Q.; Guo, H.; Yang, L.; Cai, L.; Wang, X.; Liu, B.; Wang, S. Ectopic expression of R3 MYB transcription factor gene OsTCL1 in Arabidopsis, but not rice, affects trichome and root hair formation. Sci Rep. 2016, 6, 19254. [Google Scholar] [CrossRef] [PubMed]
- Wang, S.; Tiwari, S.B.; Hagen, G.; Guilfoyle, T.J. Auxin response factor7 restores the expression of auxin-responsive genes in mutant Arabidopsis leaf mesophyll protoplasts. Plant Cell 2005, 17, 1979–1993. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Tiwari, S.B.; Wang, X.; Hagen, G.; Guilfoyle, T.J. AUX/IAA proteins are active repressors, and their stability and activity are modulated by auxin. Plant Cel l 2001, 13, 2809–2822. [Google Scholar] [CrossRef] [Green Version]
- Gao, X.; Chen, J.; Dai, X.; Zhang, D.; Zhao, Y. An effective strategy for reliably isolating heritable and Cas9-free Arabidopsis mutants generated by CRISPR/Cas9-mediated genome editing. Plant Physiol. 2016, 171, 1794–1800. [Google Scholar] [CrossRef] [Green Version]
- Clough, S.J.; Bent, A.F. Floral dip: A simplifified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J. 1998, 16, 735–743. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Chen, S.; Zhang, N.; Zhang, Q.; Zhou, G.; Tian, H.; Hussain, S.; Ahmed, S.; Wang, T.; Wang, S. Genome editing to integrate seed size and abiotic stress tolerance traits in Arabidopsis reveals a role for DPA4 and SOD7 in the regulation of inflorescence architecture. Int. J. Mol. Sci. 2019, 20, 2695. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Wang, S.; Barron, C.; Schiefelbein, J.; Chen, J.G. Distinct relationships between GLABRA2 and single-repeat R3 MYB transcription factors in the regulation of trichome and root hair patterning in Arabidopsis. New Phytol. 2010, 185, 387–400. [Google Scholar] [CrossRef] [PubMed] [Green Version]
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Wang, Y.; Tian, H.; Wang, W.; Wang, X.; Zheng, K.; Hussain, S.; Lin, R.; Wang, T.; Wang, S. The Carboxyl-Terminus of TRANSPARENT TESTA GLABRA1 Is Critical for Its Functions in Arabidopsis. Int. J. Mol. Sci. 2021, 22, 10039. https://doi.org/10.3390/ijms221810039
Wang Y, Tian H, Wang W, Wang X, Zheng K, Hussain S, Lin R, Wang T, Wang S. The Carboxyl-Terminus of TRANSPARENT TESTA GLABRA1 Is Critical for Its Functions in Arabidopsis. International Journal of Molecular Sciences. 2021; 22(18):10039. https://doi.org/10.3390/ijms221810039
Chicago/Turabian StyleWang, Yating, Hainan Tian, Wei Wang, Xutong Wang, Kaijie Zheng, Saddam Hussain, Rao Lin, Tianya Wang, and Shucai Wang. 2021. "The Carboxyl-Terminus of TRANSPARENT TESTA GLABRA1 Is Critical for Its Functions in Arabidopsis" International Journal of Molecular Sciences 22, no. 18: 10039. https://doi.org/10.3390/ijms221810039
APA StyleWang, Y., Tian, H., Wang, W., Wang, X., Zheng, K., Hussain, S., Lin, R., Wang, T., & Wang, S. (2021). The Carboxyl-Terminus of TRANSPARENT TESTA GLABRA1 Is Critical for Its Functions in Arabidopsis. International Journal of Molecular Sciences, 22(18), 10039. https://doi.org/10.3390/ijms221810039