The WUSCHEL-related homeobox transcription factor CsWOX3 negatively regulates fruit spine morphogenesis in cucumber (Cucumis sativus L.)

Abstract

Cucumber (Cucumis sativus L.) is a widely cultivated crop with rich germplasm resources, holding significant nutritional value. It also serves as an important model for studying epidermal cell fate and sex determination. Cucumbers are covered with multicellular and unbranched trichomes, including a specific type called spines found on the surface of the fruit. The presence and density of these fruit spines determine the visual quality of cucumber fruits. However, the key regulatory genes and mechanisms underlying cucumber fruit spine development remain poorly understood. In this study, we identified a WUSCHEL-related homeobox (WOX) family gene CsWOX3, which functioned as a typical transcriptional repressor and played a negative role in fruit spine development. Spatial-temporal expression analysis revealed that CsWOX3 exhibited a relatively high expression level in the cucumber female floral organs, particularly in the fruit exocarp. Knockout of CsWOX3 using CRISPR/Cas9 resulted in a significant 2-to-3-fold increase in the diameter of fruit spines base, while overexpression led to a 17% decrease in the diameter compared to the wild-type. A SQUAMOSA PROMOTER BINDING PROTEIN-LIKE transcription factor CsSPL15 could directly bind and activate the expression of CsWOX3, thereby suppressing the expression of downstream auxin-related genes, such as CsARF18. Additionally, the RING-finger type E3 ubiquitin ligase CsMIEL1-like interacted with the HD domain of CsWOX3, which might result in the ubiquitination and subsequent alteration in protein stability of CsWOX3. Collectively, our study uncovered a WOX transcription factor CsWOX3 and elucidated its expression pattern and biological function. This discovery enhances our comprehension of the molecular mechanism governing cucumber fruit spine morphogenesis.

Acknowledgements

We would like to express our special thanks to Prof. Xiangdong Li from Shandong Agricultural University for providing the TRSV-VIGS vectors. This study was supported by the National Natural Science Foundation of China (31830080), the National Key Research and Development Program ‘Strategic Science and Technology Innovation Cooperation’ Key Special Project (2023YFE0206900), the Project of Yazhouwan Scientific and Technological Administration of Sanya (SYND-2021-18 and SYND-2022-20), Joint Research Institute of China Agricultural University in Aksu, and the Construction of Beijing Science and Technology Innovation and Service Capacity in Priority Subjects (CEFF-PXM2019_014207_000032).

Author contributions

X.L. and H.R. designed the studies and contributed to the original concept of the project; S.X. performed the research and drafted the manuscript; Y.W., S.Y., S.F., and K.S. contributed to experimental design, data interpretation, and manuscript editing; F.W., M.A., Y.Q., M.W., and M.F. contributed to phenotypic data collection in the field; H.R. managed the project; X.L. and Z.L. participated in data analysis and manuscript writing.

Data availability

All relevant data can be found within the manuscript and its supplemental materials.

Conflict of interest statement

The authors declare that there is no conflict of interest.

Supplementary data

Supplementary data is available at Horticulture Research online.

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