HvEPF1 acts to prevent cells entering the stomatal lineage, guard mother cell maturation, and substomatal cavity and subsidiary cell formation. Schematic to illustrate the putative mode of action of HvEPF1 in barley stomatal development. Left to right: Undifferentiated epidermal cells at the base of leaves are formed in cellular files. Cells in some files gain the capacity to divide asymmetrically to create small stomatal precursor cells shown here as immature guard mother cells (GMC, green). A developmental step, potentially under the control of the transcription factor MUTE, stimulates guard mother cell maturation (dark green) and division of adjacent epidermal cells to form subsidiary cells (SC, orange). Mature GMCs then divide symmetrically to form pairs of dumbbell-shaped guard cells (red). In the underlying mesophyll layer (M, green shaded regions), a substomatal cavity forms during either the mature GMC or guard cell stage, although the exact developmental staging of this is process is unknown. In the HvEPF1 overexpressing plants, HvEPF1 prevents GMC maturation, perhaps through the suppression of MUTE activity, resulting in arrested GMCs that are unable to differentiate into mature stomatal complexes complete with subsidiary cells, guard cells, and substomatal cavities. Drawn with reference to Brachypodium development in Raissig et al. (2016).