Abstract
The zinc finger protein (ZFP) transcription factor family plays an important role in regulating plant growth, development, and response to abiotic stress. In this study, we aimed to determine the role of GmZAT4, a C2H2-type transcription factor, in abiotic stress tolerance. The complete coding sequence of the GmZAT4 gene was isolated from soybean root RNA, which shows highest expression level compared with leaf, flower and other tissues. Using multiple sequence alignment and conserved domain analysis, we showed that GmZAT4 is a typical C2H2-type transcription factor which is comprised of two C2H2 domains, including a highly conserved QALGGH motif, and implied the regulation of abiotic stress tolerance in plant. A phylogenetic tree revealed that the soybean GmZAT4 gene clustered with ZAT4 from Glycine soja and AZF1, AZF2, and AZF3 from Arabidopsis thaliana. The mRNA expression levels of GmZAT4 were determined in two soybean cultivars by quantitative reverse transcription (qRT)-PCR and compared. The results showed higher expression (up to 60, 25 and 4 times, respectively) in the drought-tolerant type (Jinda 74) compared to the drought-sensitive soybean cultivar (Jinda 53) following treatment with 18% PEG, 150 mM NaCl, or 100 µM abscisic acid (ABA). GmZAT4 was ectopically over-expressed in A. thaliana to determine its role in abiotic stress tolerance. GmZAT4 overexpression enhanced the tolerance of A. thaliana to treatment with 20% PEG and 150 mM NaCl, and improved the germination rate following treatment with 1 µM or 2 µM ABA. The expression profiles of marker genes in the ABA signaling pathway, such as RD29A, RD29B, ABI, and RAD, indicated that GmZAT4 enhanced the abiotic stress tolerance of Arabidopsis. These results suggest that the C2H2-type ZFP encoded by GmZAT4 plays an important role in PEG and NaCl stress tolerance and ABA responses in soybean and A. thaliana.
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Acknowledgements
We would like to thank Professor Donald Grierson, University of Nottingham, UK, for discussion and helping with the manuscript. We also thank Prof. Guiquan Li for donating the plant material and Prof. Yuguo Wang for giving advice for the experiments. We would like to thank Science and Technology Project of Shanxi Province (20150311007-1) and Research Project Supported by Shanxi Scholarship Council of China (2017069).
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ZS, SH, HL and YH designed the experiments and drafted the manuscript. ZS and KH collected plant materials and cloned the gene. RL, BG and LW transformed the Arabidopsis and analysis the data. YH modified the manuscript.
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13205_2019_1673_MOESM1_ESM.pdf
Figure S1. Transgenic GmZAT4-transformed lines identified by gDNA-PCR and RT-PCR. (A) WT, OE1, and OE2 at 4 weeks. (B) CDS fragments of GmZAT4 amplified from vector (control, CK), transgenic lines (lanes 1–6), WT (lane 7), and negative control (lane 8) using gDNA-PCR. (C) RT-PCR of GmZAT4 gene expression in WT and transgenic lines (lanes 1–6). Lines #1 and # 3 (designated OE1 and OE3, respectively) were selected from the six transgenic lines and used for further analysis. (PDF 96 KB)
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Sun, Z., Liu, R., Guo, B. et al. Ectopic expression of GmZAT4, a putative C2H2-type zinc finger protein, enhances PEG and NaCl stress tolerances in Arabidopsis thaliana. 3 Biotech 9, 166 (2019). https://doi.org/10.1007/s13205-019-1673-0
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DOI: https://doi.org/10.1007/s13205-019-1673-0