LncR12304.1-miR1507c-CsNADH-GOGAT ceRNA module regulates amino acid biosynthesis in tea plant (Camellia sinensis)

Abstract

Glutamate synthase (GOGAT) is crucial for nitrogen metabolism and amino acid biosynthesis in tea plants, yet the post-transcriptional regulation of GOGAT remains unclear. This study identified miR1507c as a direct interactor of CsNADH-GOGAT, confirmed by DLR assays and 5′ RLM-RACE. Notably in tobacco, the relative luciferase activity in plants overexpressing CsNADH-GOGAT and co-expressing miR1507c + CsNADH-GOGATm3 (mutant) were significantly higher than in those co-expressing miR1507c + CsNADH-GOGAT. Overexpression of miR1507c also significantly suppressed the expression of CsNADH-GOGAT and endogenous NtNADH-GOGAT homologs. Leveraging lncRNA sequencing, we screened lncR12304.1 as a ceRNA that regulates CsNADH-GOGAT by competitively binding to miR1507c. Cytoplasmic co-localization (validated by FISH) and direct interaction (confirmed by DLR assays) between lncR12304.1 and miR1507c were established. RNA pull down-qPCR further demonstrated miR1507c binding to both lncR12304.1 and CsNADH-GOGAT. The regulatory axis lncR12304.1–miR1507c– CsNADH-GOGAT was substantiated in vivo: (i) in tea roots/shoots under varying nitrogen treatments and following miR1507c suppression using Antagomir, and (ii) in tobacco via transient co-overexpression. Collectively, our results demonstrate the establishment of this ceRNA network and its role in regulating glutamate and theanine biosynthesis.

Acknowledgements

This study was supported by the Yunnan Provincial Basic Research Programs (202201AT070253, 202501AS070039), Yunnan Provincial Basic Research Agriculture (202401BD070001-044, 202401BDO70001-007), the Open Fund of State Key Laboratory of Tea Plant Biology and Utilization (NKLTOF20240110), and the Anhui Provincial Natural Science Foundation (2408085MC077).

Author contributions

Zhiwei Liu (Writing-original draft, Data curation, Formal analysis), Yan Ma (Supervision), Teng Wang (Formal analysis), Qiuyue Chen (Project administration), Dihan Yang (Software), Yiqing Guan (Investigation), Tianyuan Yang (Writing-review & editing), and Ming Zhao (Writing-review & editing, Methodology, Funding acquisition)

Data availability

All relevant data in this study are provided in the article and its supplementary files. The raw data of lncRNA in this study had been submitted to NCBI Sequence Read Archive (ID: PRJNA1304500).

Conflicts of interest statement

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Supplementary material

Supplementary material is available at Horticulture Research online.

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