Temperature signals drive grass secondary cell wall thickening

In grasses, stem elongation is driven by intercalary meristems at node-internode junctions, where cells divide, elongate, and in some cell types secondary wall maturation. Cellulose is the predominant polymer in plant cells and the most abundant biopolymer on Earth. It is synthesized at the plasma membrane by multi-protein complexes that include CELLULOSE SYNTHASE A (CESA) proteins. To investigate the spatiotemporal regulation of cellulose deposition during development, we developed a CESA8 luciferase gene expression reporter system in Brachypodium distachyon. High bioluminescence was observed in stem nodes, a specific region of elongating internodes, and the inflorescence, indicating sites of active secondary wall deposition. Within internodes, luminescence followed a distinct pattern, with a "dark zone" directly above the node with minimal signal, followed by a "bright zone" approximately 5 mm above the node where bioluminescence peaked. Histological, biophysical, and transcript analysis confirmed that luminescence intensity correlates with thickened secondary cell walls, increased cellulose crystallinity, and elevated CESA8 transcript levels. Time-lapse imaging revealed that CESA8 expression follows a robust diurnal rhythm governed by thermocycles alone, with peak expression occurring in the early morning. Temperature pulse experiments revealed an immediate but transient response of CESA8 to temperature shifts, which we modeled as an incoherent feed-forward loop. Finally, we found a strong correlation between CESA8 expression and stem elongation, highlighting the role of secondary cell wall thickening in supporting upright growth. These findings provide new insights into the regulation of secondary wall formation and its integration with environmental cues, advancing our understanding of grass stem development. SIGNIFICANCEUnderstanding how grasses build strong stems is essential for improving biomass production and crop resilience. In grasses, stem elongation and secondary cell wall thickening occur in distinct zones, yet the precise timing and regulation of this process remain unclear. To investigate this phenomenon, we developed a real-time imaging system to track the expression of CESA8, a key gene involved in cellulose synthesis. Our findings reveal that secondary wall thickening follows a daily rhythm controlled by temperature rather than light. These insights provide a foundation for optimizing plant architecture in bioenergy crops, improving their efficiency and sustainability.