LOX5 roles in forming diverse aroma profiles of Nicotiana tabacum via multi-omics analysis

As the global cash crops, the formation of distinct aroma profiles in Nicotiana tabacum have garnered significant research interest. Lipids as precursors of aroma compounds in plants, play an essential role in defining the unique aroma characteristics of tobacco. In this study, lipidomics analysis was performed to identify a total of 471 lipid components in leaves of strong fragrance tobacco (SF) and mild fragrance tobacco (MF). Comparative analysis of SF and MF revealed 70 differentially changed lipid compounds, with 26 up-regulated and 44 down-regulated lipids. The up-regulated lipids were predominantly glycoglycerolipids (GL) and glycerophospholipids (GP), while the down-regulated lipids were enriched in sphingolipids (SP) and fatty acyls (FA). Gas chromatography-mass spectrometry (GC-MS) revealed that SF exhibited relatively higher levels of cis-3-hexenal and cis-3-hexenol compared to MF. Transcriptomics analysis identified a total of 905 differential expression genes (DEGs), with 370 up-regulated and 535 down-regulated DEGs. Notably, a robust correlation was observed between the expression of the key gene lipoxygenase 5 (LOX5) and content of phosphatidylcholine (PC) (15:0_18:3), a precursors of linolenic acid. Additionally, a positive correlation was detected between LOX5 expression and the degradation products of linolenic acid, including cis-3-hexenal and cis-3-hexenol. Transient over-expression of LOX5 in tobacco leaves resulted in an enhanced accumulation of these linolenic acid degradation products. Our findings elucidate the mechanism underlying the formation of diverse aroma profiles in tobacco, and provide a theoretical basis for breeding high-quality tobacco crop. • Cis-3-hexenol and cis-3-hexenal were increased in SF relative to MF. • LOX5 expression was correlated with PC (15:0_18:3) content and - cis-3-hexenal and cis-3-hexenol levels. • LOX5 - was involved in shaping the aromatic profile.