Water stress is an increasing problem in agriculture, given the impact of climate change. Discovering how abiotic stress signals like abscisic acid (ABA) control plant adaptive responses to fluctuating water stress is vital for future-proofing crops. Our team recently discovered an ABA-regulated response to transient water stress called Xerobranching (Orman et al  Curr Biol), where root branching ceases in response to fluctuating soil moisture. Preliminary results suggest that ABA regulates Xerobranching by closing inter-cellular pores, termed plasmodesmata (PD), in root cells - disrupting delivery of the branching signal auxin to lateral root 'stem cells'. To discover how ABA controls root responses to fluctuating water stress we propose to characterise key genes, signals and mechanisms underpinning Xerobranching exploiting novel bioassays, hormone reporters and a cell-scale Raman water-imaging technique (Pascut et al  Nature Comms). Our proposal seeks to build on these promising results and techniques to discover how roots respond to transient water stress by addressing 4 objectives: OBJ1 exploits Raman imaging to map hydraulic fluxes across root tissues exposed to Xerobranching stimulus. OBJ2 uses hormone and Raman imaging approaches to reveal whether ABA co-mobilises with water fluxes during Xerobranching responses. OBJ3 addresses if ABA triggers closure of inter-cellular pores termed plasmodesmata. Finally, OBJ4 will discover whether plasmodesmata closure disrupts movement of branching signal auxin, which is needed to 'prime' lateral root 'stem cells' in inner root tissues. The knowledge generated about the new signals, genes and their regulatory pathways will underpin on going efforts to re-engineer root systems architecture and future proof crops. The expertise, resources and tools that have been assembled for this project at Nottingham with our international collaborators uniquely position us to successfully complete this project.