Plant biomass (essentially the cell wall) is the carbon source that will underpin a sustainable bioeconomy. Drought can severely affect plant growth and reduces crop yield and quality, and is a growing problem in agriculture. It has been proposed that modification of the cell wall can play a crucial role in plants surviving a water-deficit stress. What is not understood is how these changes enable drought tolerance at a molecular level. Previously, we demonstrated that multi-dimensional ssNMR can be used to investigate the impact of genetic engineering on the structure and arrangement of cell wall polymers. We used this information to develop iteratively refined models of cell wall nanoarchitecture. Here, we want to use these tools to explore cell wall remodeling when the plant is under water-deficit stress, and understand the mechanism and the roles of cell wall polymers in responding to loss of water. This will provide a framework for developing breeding strategies for biomass crops with improved drought tolerance. We will target two key cell wall components that have been implicated in drought tolerance: acetylation and lignification. In the first aim, we will use wild-type and engineered Arabidopsis with altered pectic methyl- or O-acetyl-ester substitutions to explore the functional roles of these cell wall polysaccharide substitutions under drought stress. In the second aim, we will use our collection of model and bioenergy species (Arabidopsis, sorghum, poplar, switchgrass) which have been all been modified to have reduced lignin via introduction of the bacterial QsuB gene. We will explore the impact of temporal drought treatments on the cell wall architecture and understand the role of the 3D lignin-xylan-cellulose interactions in response to drought. We will also employ cryo-electron tomography to visualize the ultrastructure of the plant cell wall and further validate the ssNMR data. These data will be used to establish a structure-function relationship when refining our cell walls models, which in turn underpin our development of drought-tolerant bioenergy crops.