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Characterization of a new allelic mutant of DWARF3 in rice and analysing its function and stability in the presence of strigolactone

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Abstract

Strigolactones (SLs) are important intrinsic growth regulators that control plant architecture by coordinating shoot and root development. Recent studies demonstrate that SL signals act via targeting the degradation protein DWARF53 (D53) family of chaperonin-like proteins. This process requires DWARF14 (D14) as strigolactones signal receptor and DWARF3 (D3) forming Skp-Cullin-F-box (SCF) complex as ubiquitin E3 ligase. Although the interactions of these signal components can be expected, where and how the SLs signalling occur within cells in a tissue-specific manner is still uncertain. In this study, we characterize a rice high-tillering dwarf mutant, ext.-M1B, displaying resistance to synthetic strigolactone mixture rac-GR24. Through genetic analysis, we find that ext.-M1B is a new allelic mutant of D3 with a nucleotide mutation resulting in a truncated protein of wide-type D3. We demonstrate that the mutation affects neither gene expression level nor the protein sub-cellular localization, whereas it disrupts the perception of SLs signal in ext.-M1B mutant. Moreover, we find that overexpression of D3 in wild type background causes no significant phenotype, but suppression of D3 by RNA interfering results in a clear phenocopy of SL mutants. By expressing fluorescent D3 fusion protein in rice, we first show that D3 is stable consistently in the nucleus with or without strigolactone treatment. Taken together, our data indicates that D3 encoding an F-box protein in nucleus, as a stable signal component response to strigolactone regulating rice shoot architecture.

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Acknowledgements

This research was supported by Sichuan Provincial Science and Technology Platform project and Sichuan Science and Technology Achievements Transformation Project, China. (2014GPTZ0010).

Author’s contributions

P.L., S.W. and Y. L designed most of the experiments and directed the project; Y.L. performed the entire experiments. X. H and H. W carried out the field experiments and investigations. F.L. performed the expression analysis, tissue localization; S.L., A.Z., J.Z., H.L., Q. D, and L.W. also contributed to the experimental design; Y.L., S.W., and P.L. analysed the data and wrote the paper.

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    1. Rice Research Institute, Sichuan Agricultural University, 211 Huimin Road, Wenjiang, Sichuan, 611130, China

      Yueyang Liang, Shiquan Wang, Xiaoxi Huang, Haipeng Wang, Fenlong Liu, Shuangcheng Li, Jun Zhu, Qiming Deng, Huainian Liu, Aiping Zheng, Lingxia Wang & Ping Li

    2. State Key Laboratory of Hybrid Rice, Sichuan Agricultural University, Chengdu, 611130, China

      Yueyang Liang, Shiquan Wang, Shuangcheng Li, Jun Zhu, Qiming Deng, Huainian Liu, Aiping Zheng, Lingxia Wang & Ping Li

    3. Key Laboratory of Crop Genetic Resources and Improvement, Sichuan Agricultural University, Ministry of Education, Ya’an, Sichuan, 625014, China

      Yueyang Liang, Shiquan Wang, Shuangcheng Li, Jun Zhu, Qiming Deng, Huainian Liu, Aiping Zheng, Lingxia Wang & Ping Li

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    1. Yueyang Liang
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    Yueyang Liang and Shiquan Wang contributed equally to this work.

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    Fig. S1

    Mutation caused frame shift results in a predicted stop codon in mutant. Comparison of DNA sequence and relative coding amino acid between wild-type and ext.-M1B showing the nucleotide mutation from A to CC leads to corresponding amino acid changed after site 334. The red asterisk indicates the predicted stop codon (JPEG 873 kb)

    Fig. S2

    Transcriptional level of D14 is increased in mutant. Expression of D3 and D14 in one-week-old rice seedlings of wild-type and ext.-M1B were determined by qRT-PCR using rice Actin as the internal reference. Values are means ± SEM (n = 3); **p < 0.01, determined by Student’s t-test (JPEG 118 kb)

    Table S1

    Primers used in this study (DOCX 22 kb)

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    Liang, Y., Wang, S., Huang, X. et al. Characterization of a new allelic mutant of DWARF3 in rice and analysing its function and stability in the presence of strigolactone. Mol Breeding 37, 39 (2017). https://doi.org/10.1007/s11032-017-0640-x

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