Natural phenolic autofluorescence reveals cell wall-specific distribution of hydroxycinnamates in Brachypodium...

p-Coumaroyl-CoA:monolignol transferase has been demonstrated to be involved in the coumaroylation of lignins in Brachypodium distachyon. However, the specific localization of acylation remains poorly documented at the tissue and cell wall levels in this species. Detecting molecules that are sometimes present at levels <1% in the cell wall remains a challenge, especially when suitable antibodies are not always available. In this work, we applied fluorescence microscopy methods, including large-beam excitation scanning at the SOLEIL synchrotron, to detect variations induced by hydroxycinnamic acids within B. distachyon stems. Using principal component analysis (PCA), fluorescence microscopy imaging effectively distinguished genotypes differing subtly in phenolic composition. The results were supported by immunolabeling, which confirmed that p-coumarate co-localizes with S-unit lignin in lignified tissues, while ferulate is broadly distributed. Strong autofluorescence in the mestome sheath and metaxylem pit area indicated a potential functional role for p-coumaric acid in these tissues. Finally, autofluorescence-based imaging and PCA proved to be a robust, non-destructive tool for visualizing lignin and phenolic compounds, offering valuable insights into cell wall specialization and phenolic function.