New insights into uranium stress responses of Arabidopsis roots through membrane- and cell wall-associated proteome analysis.

Uranium (U) is a non-essential and toxic metal for plants. In Arabidopsis thaliana plants challenged with uranyl nitrate, we showed that U was mostly (64-71% of the total) associated with the root insoluble fraction containing membrane and cell wall proteins. Therefore, to uncover new molecular mechanisms related to U stress, we used label-free quantitative proteomics to analyze the responses of the root membrane- and cell wall-enriched proteome. Of the 2,802 proteins identified, 458 showed differential accumulation (≥1.5-fold change) in response to U. Biological processes affected by U include response to stress, amino acid metabolism, and previously unexplored functions associated with membranes and the cell wall. Indeed, our analysis supports a dynamic and complex reorganization of the cell wall under U stress, including lignin and suberin synthesis, pectin modification, polysaccharide hydrolysis, and Casparian strips formation. Also, the abundance of proteins involved in vesicular trafficking and water flux was significantly altered by U stress. Measurements of root hydraulic conductivity and leaf transpiration indicated that U significantly decreased the plant's water flux. This disruption in water balance is likely due to a decrease in PIP aquaporin levels, which may serve as a protective mechanism to reduce U toxicity. Finally, the abundance of transporters and metal-binding proteins was altered, suggesting that they may be involved in regulating the fate and toxicity of U in Arabidopsis. Overall, this study highlights how U stress impacts the insoluble root proteome, shedding light on the mechanisms used by plants to mitigate U toxicity.