Abstract
Key Message
Plasma membrane-localized AtAVT6D importing aspartic acid can be targeted to develop plants with enhanced osmotic and nitrogen-starvation tolerance. AtAVT6D promoter can be exploited as a stress-inducible promoter for genetic improvements to raise stress-resilient crops.
Abstract
The AtAVT6 family of amino acid transporters in Arabidopsis thaliana has been predicted to export amino acids like aspartate and glutamate. However, the functional characterization of these amino acid transporters in plants remains unexplored. The present study investigates the expression patterns of AtAVT6 genes in different tissues and under various abiotic stress conditions using quantitative Real-time PCR. The expression analysis demonstrated that the member AtAVT6D was significantly induced in response to phytohormone ABA and stresses like osmotic and drought. The tissue-specific expression analysis showed that AtAVT6D was strongly expressed in the siliques. Taking together these results, we can speculate that AtAVT6D might play a vital role in silique development and abiotic stress tolerance. Further, subcellular localization study showed AtAVT6D was localized to the plasma membrane. The heterologous expression of AtAVT6D in yeast cells conferred significant tolerance to nitrogen-deficient and osmotic stress conditions. The Xenopus oocyte studies revealed that AtAVT6D is involved in the uptake of Aspartic acid. While overexpression of AtAVT6D resulted in smaller siliques in Arabidopsis thaliana. Additionally, transient expression studies were performed with the full-length AtAVT6D promoter and its deletion constructs to study the effect of ACGT-N24-ACGT motifs on the reporter gene expression in response to abiotic stresses and ABA treatment. The fluorometric GUS analyses revealed that the promoter deletion construct-2 (Pro.C2) possessing a single copy of ACGT-N24-ACGT motif directed the strongest GUS expression under all the abiotic conditions tested. These results suggest that Pro.C2 can be used as a stress-inducible promoter to drive a significant transgene expression.
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Acknowledgements
The authors are grateful to BITS Pilani, K. K. Birla Goa Campus, Goa, India for providing infrastructural and logistic support. We thank Dr. Yi Chen for Xenopus oocyte experiment. PD is thankful to BITS Pilani, CSIR and EMBO for financial assistance. This study was supported by SERB Project EMR/2016/002470 sanctioned by the government of India to SM and RM. AJM was funded by the UK Biotechnology and Biological Sciences Research Council (BBSRC) Institute Strategic Program Grants ‘Molecules from Nature’ (BB/P012523/1) and ‘Plant Health’ (BB/P012574/1) and the John Innes Foundation.
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The details of funding are mentioned in the Acknowledgements section as-<br>This study was supported by SERB Project EMR/2016/002470 sanctioned by the government of India to SM and RM. AJM was funded by the UK Biotechnology and Biological Sciences Research Council (BBSRC) Institute Strategic Program Grants ‘Molecules from Nature’ (BB/P012523/1) and ‘Plant Health’ (BB/P012574/1) and the John Innes Foundation.
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PD planned, designed, performed experiments, analyzed the data, and wrote the manuscript. AM supervised with the yeast and Xenopus studies. RA helped with the promoter studies. RM, and SM supervised the planning, designing of the experiments, writing of the manuscript and gave final approval for publication. All authors read and approved the manuscript. RM agrees to serve as the author responsible for contact and ensures communication.
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Dhatterwal, P., Mehrotra, S., Miller, A.J. et al. Effect of ACGT motif in spatiotemporal regulation of AtAVT6D, which improves tolerance to osmotic stress and nitrogen-starvation. Plant Mol Biol 109, 67–82 (2022). https://doi.org/10.1007/s11103-022-01256-x
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DOI: https://doi.org/10.1007/s11103-022-01256-x