Involvement of the LcARF17- and LcRAP2-4-LcLOX7 regulatory modules in the biosynthesis of fresh aroma in litchi aril

Abstract

Fatty acid-derived volatile organic compounds (VOCs), especially C ₆ and C ₉ aldehydes and alcohols, are vital contributors to the fresh aroma of fruits. However, the specific volatiles responsible for this freshness and their biosynthetic regulatory mechanisms remain poorly characterized in litchi (Litchi chinensis Sonn.). In this study, we systematically characterized the VOC profiles of 24 representative litchi cultivars and identified four critical compounds- trans,cis-2,6-nonadien-1-ol, 1-hexanol, (E)-6-nonenal, and (E)-2-hexen-1-ol-as primary determinants of fresh-aroma variation. Weighted gene co-expression network analysis of the transcriptomic data, corroborated by RT-qPCR, revealed a strong positive correlation between the expression of LcLOX7 and the abundance of these key fresh-aroma volatiles. Functional characterization via LcLOX7 overexpression in litchi callus and tomato fruits validated its pivotal role in enhancing the biosynthesis of fatty acid-derived VOCs, specifically C ₉ volatiles. Subsequent in vivo and in vitro assays confirmed the direct transcriptional activation of LcLOX7 by two transcription factors (TF), LcARF17 and LcRAP2-4. The expression patterns of these TFs correlated with the accumulation of key fresh-aroma volatiles across different litchi cultivars and paralleled LcLOX7 expression during fruit ripening. Moreover, overexpression and silencing of LcARF17 or LcRAP2-4 in litchi callus validated their regulatory function in promoting C ₉ volatile synthesis. Our findings collectively support a regulatory model wherein the LcARF17/LcRAP2-4–LcLOX7 module orchestrates the biosynthesis of fresh aroma in litchi fruit. Notably, this study provides the first evidence that ARF transcription factor participates in the formation of fresh fruit aroma, thereby offering novel insights for the molecular breeding of flavor quality in fruit crops.

Acknowledgements

This research was funded by the China Agricultural Research System (CARS-32-05), the National Natural Science Foundation of China (grant numbers 32330092 and 32202447), and the Natural Science Foundation of Guangdong Province (grant numbers 2023A1515012661). We extend our thankful to Researchers Huang X.F. and Yuan Y. from Dongguan Botanical Garden for their valuable contributions to the collection of litchi cultivars.

Author contributions

Jianguo Li, Xingshuai Ma, and Minglei Zhao conceived and designed the study; Zhuoyi Liu performed all of the experiments; Yimeng Wang, Ke Ma, Zidi He, Zhiqi Li, and Hang Zhang provided technical assistance; Zhuoyi Liu, Xingshuai Ma, Minglei Zhao, and Jianguo Li drafted and revised the manuscript.

Data availability

All data generated or analyzed during this study are included in this published article and its supplementary information files.

Conflicts of interest statement

The authors declare no competing interests.

Supplementary material

Supplementary material is available at Horticulture Research online.

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