1 Department of Biology, Concordia University, Montreal, Quebec, H4B 1R6, Canada.
2 Key Laboratory of Molecular Epigenetics of MOE and Institute of Genetics & Cytology, Northeast Normal University, Changchun 130024, China.
3 Lethbridge Research and Development Centre, Agriculture and Agri-Food Canada, Lethbridge, Alberta, T1J 4B1, Canada.
4 Laboratory of Plant Molecular Genetics & Crop Gene Editing, School of Life Sciences, Linyi University, Linyi 276000, China.

Peptide signaling has emerged as a key component of plant growth and development, including stomatal patterning, which is critical for plant productivity and survival. Although exciting progress has been made in understanding EPIDERMAL PATTERNING FACTOR (EPF) signaling in Arabidopsis, the mechanisms by which EPF peptides control different stomatal patterns and morphologies in grasses is poorly understood. Here, by examining expression patterns, overexpression transgenics, and cross-species complementation, the antagonistic stomatal ligands orthologous to Arabidopsis AtEPF2 and AtSTOMAGEN/AtEPFL9 peptides were identified in Triticum aestivum(wheat) and the grass model organism Brachypodium distachyon. Application of bioactive BdEPF2 peptides inhibited stomatal initiation, but not the progression or differentiation of stomatal precursors in Brachypodium. Additionally, the inhibitory roles of these EPF peptides during grass stomatal development were suppressed by the contrasting positive action of the BdSTOMAGEN peptide in a dose-dependent manner. These results not only demonstrate how conserved EPF peptides that control different stomatal patterns exist in nature but also suggest new strategies to improve crop yield through the utilization of plant-derived antagonistic peptides that optimize stomatal density on the plant epidermis.