Wheat wounding-responsive HD-Zip IV transcription factor GL7 is predominantly expressed in grain and activates genes encoding defensins
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Several classes of transcription factors are involved in the activation of defensins. A new type of the transcription factor responsible for the regulation of wheat grain specific defensins was characterised in this work.
HD-Zip class IV transcription factors constitute a family of multidomain proteins. A full-length cDNA of HD-Zip IV, designated TaGL7 was isolated from the developing grain of bread wheat, using a specific DNA sequence as bait in the Y1H screen. 3D models of TaGL7 HD complexed with DNA cis-elements rationalised differences that underlined accommodations of binding and non-binding DNA, while the START-like domain model predicted binding of lipidic molecules inside a concave hydrophobic cavity. The 3′-untranslated region of TaGL7 was used as a probe to isolate the genomic clone of TdGL7 from a BAC library prepared from durum wheat. The spatial and temporal activity of the TdGL7 promoter was tested in transgenic wheat, barley and rice. TdGL7 was expressed mostly in ovary at fertilisation and its promoter was active in a liquid endosperm during cellularisation and later in the endosperm transfer cells, aleurone, and starchy endosperm. The pattern of TdGL7 expression resembled that of genes that encode grain-specific lipid transfer proteins, particularly defensins. In addition, GL7 expression was upregulated by mechanical wounding, similarly to defensin genes. Co-bombardment of cultured wheat cells with TdGL7 driven by constitutive promoter and seven grain or root specific defensin promoters fused to GUS gene, revealed activation of four promoters. The data confirmed the previously proposed role of HD-Zip IV transcription factors in the regulation of genes that encode lipid transfer proteins involved in lipid transport and defence. The TdGL7 promoter could be used to engineer cereal grains with enhanced resistance to insects and fungal infections.
KeywordsBarley Biotechnology Molecular model Rice Structural bioinformatics Wheat Wounding Yeast-1-hybrid
Basic leucine zipper
Days after pollination
Ethylene-responsive element binding factor
GLABRA2-like clone 7
Heat-shock transcription factor
Protein data bank
Standard errors of mean
Steroidogenic acute regulatory protein-related lipid-transfer
3′ Untranslated region
We acknowledge the contributions of Ainur Ismagul and Serik Eliby in plant transformation. We thank Margaret Pallotta for technical assistance with BAC library screening and promoter cloning. Ursula Langridge, Lorraine Carruthers and Alex Kovalchuk are thanked for their assistance with growing of plants in the glasshouse. This work was supported by the Australian Research Council (Grant No. LP120100201 to M.H. and S.L.), the Grains Research and Development Corporation and the Government of South Australia.
Conceived, designed experiments and analysed data: NK and SL. Cloning and Y1H: NB and WW. Transient expression assays: NR and OE. Q-PCR experiments: NS. Plant transformation and analysis: RS, AATJ, NK. 3D molecular modelling and bioinformatics: MH. Discussed the data: SL, NK, PL and MH. Writing of the manuscript: SL. Contributed to writing: MH.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
- Dezar CA, Giacomelli JI, Manavella PA, Ré DA, Alves-Ferreira M, Baldwin IT, Bonaventure G, Chan RL (2011) HAHB10, a sunflower HD-Zip II transcription factor, participates in the induction of flowering and in the control of phytohormone-mediated responses to biotic stress. J Exp Bot 62:1061–1076CrossRefGoogle Scholar
- Eini O, Yang N, Pyvovarenko T, Pillman K, Bazanova N, Tikhomirov N, Eliby S, Shirley N, Sivasankar S, Tingey S, Langridge P, Hrmova M, Lopato S (2013) Complex regulation by Apetala2 domain-containing transcription factors revealed through analysis of the stress-responsive TdCor410b promoter from durum wheat. PLoS ONE 8:e58713CrossRefGoogle Scholar
- Javelle M, Vernoud V, Depège-Fargeix N, Arnould C, Oursel D, Domergue F, Sarda X, Rogowsky PM (2010) Overexpression of the epidermis-specific homeodomain-leucine zipper IV transcription factor Outer Cell Layer1 in maize identifies target genes involved in lipid metabolism and cuticle biosynthesis. Plant Physiol 154:273–286CrossRefGoogle Scholar
- Kaur J, John F, Adholeya A, Velivelli S, El-Mounadi Kaoutar, Natalya N, Thomas C, Shah D (2016) Expression of apoplast-targeted plant defensin MtDef4.2 confers resistance to leaf rust pathogen Puccinia triticina but does not affect mycorrhizal symbiosis in transgenic wheat. Transgenic Res 26:37–49CrossRefGoogle Scholar
- Kovalchuk N, Smith J, Bazanova N, Pyvovarenko T, Singh R, Shirley N, Ismagul A, Johnson A, Milligan AS, Hrmova M, Langridge P, Lopato S (2012a) Characterization of the wheat gene encoding a grain-specific lipid transfer protein TdPR61, and promoter activity in wheat, barley and rice. J Exp Bot 63:2025–2040CrossRefGoogle Scholar
- Kovalchuk N, Wu W, Eini O, Bazanova N, Pallotta M, Shirley N, Singh R, Ismagul A, Eliby S, Johnson A, Langridge P, Lopato S (2012b) The scutellar vascular bundle-specific promoter of the wheat HD-Zip IV transcription factor shows similar spatial and temporal activity in transgenic wheat, barley and rice. Plant Biotechnol J 10:43–53CrossRefGoogle Scholar
- Lu P, Porat R, Nadeau JA, O’Neill SD (1996) Identification of a meristem L1 layer-specific gene in Arabidopsis that is expressed during embryonic pattern formation and defines a new class of homeobox genes. Plant Cell 8:2155–2168Google Scholar
- Page RDM (1996) TreeView: an application to display phylogenetic trees on personal computers. Comp Appl Biosci 12:357–358Google Scholar
- Roderick SL, Chan WW, Agate DS, Olsen LR, Vetting MW, Rajashankar KR, Cohen DE (2002) Structure of human phosphatidylcholine transfer protein in complex with its ligand. Nat Struct Biol 9:507–511Google Scholar