Screening and identification of a non-peptide antagonist for the peptide hormone receptor in Arabidopsis

Intercellular signaling mediated by peptide hormones and membrane-localized receptor kinases plays crucial roles in plant developmental processes. Because of their diverse functions, agonistic or antagonistic modulation of peptide signaling holds enormous promise for agricultural applications. Here we established a high-throughput screening system using a bead-immobilized receptor kinase and fluorescent-labeled peptide ligand to identify small molecules that bind peptide hormone receptors in competition with natural ligands. We used the Arabidopsis CLE9-BAM1 ligand-receptor pair to screen a library of ≈30,000 chemicals and identified NPD12704 as an antagonist for BAM1. NPD12704 also inhibited CLV3 binding to BAM1 but only minimally interfered with CLV3 binding to CLV1, the closest homolog of BAM1, demonstrating preferential receptor specificity. Treatment of clv1-101 mutant seedlings with NPD12704 enhanced the enlarged shoot apical meristem phenotype. Our results provide a technological framework enabling high-throughput identification of small non-peptide chemicals that specifically control receptor kinase-mediated peptide hormone signaling in plants.