Visualizing physical interactions between guard and epidermal cells during light-induced stomatal opening in 2 angiosperms

Stomatal function results from structural movements of guard cells in which an increase in their volume opens the pore, while a decrease in volume closes the pore. In angiosperms, neighboring epidermal cells are described as having a mechanical advantage over the guard cells, yet we have limited information on how epidermal cells contribute to stomatal pore formation. Here we visualize the structural interactions between guard cells and neighboring epidermal cells in 2 angiosperms, Vicia faba and Tradescantia virginiana, using cryo-SEM to capture guard cell and epidermal cell cross-sectional conformations in both open (light adapted and transpiring) and closed (dark adapted) stomatal states, and 3D confocal imaging in intact leaves to track the shape and movement of guard cells and epidermal cells during light-induced pore opening. We find that the change in guard cell shape as the cells increase in volume does not result in pore formation; only lateral movement of the guard-epidermal cell wall boundary results in the formation of a pore. Although it is often assumed that decreases in epidermal cell volume are required to create space in the epidermis for pore opening to occur, we found no evidence of a decrease in epidermal cell volume in T. virginiana. Instead, guard cell intrusion forces a change in epidermal cell shape, moving epidermal cell volume out of plane and creating space for pore formation to occur.