The CsDof1.8–CsLIPOXYGENASE09 module regulates C9 aroma production in cucumber.

This article investigates the regulatory mechanism underlying the accumulation of nine-carbon (C9) aroma compounds, which are key contributors to the characteristic cucumber (Cucumis sativus L.) fruit flavor. Through comparative analysis of two cucumber inbred lines (Q16 and Q24), the lipoxygenase gene CsLOX09 was identified as a pivotal enzyme responsible for C9 aroma biosynthesis, with higher expression correlating with increased C9 aroma content. Functional validation via CRISPR/Cas9 knockout of CsLOX09 resulted in an 80–99% reduction in C9 aromas, alongside altered levels of related volatile compounds and unsaturated fatty acids. Furthermore, the transcription factor CsDof1.8 was found to directly bind and activate the CsLOX09 promoter, as demonstrated by multiple molecular assays, and its overexpression enhanced CsLOX09 expression and C9 aroma accumulation. This CsDof1.8–CsLOX09 regulatory module elucidates a key transcriptional control point in cucumber fruit aroma biosynthesis, providing a foundation for flavor improvement in cucumber breeding.