CLE19 suppresses brassinosteroid signaling output via the BSL-BIN2 module to maintain BES1 activity and pollen exine patterning in Arabidopsis.

Abstract

The pollen exine serves as a protective barrier and signaling interface essential for male fertility in flowering plants. Its precise patterning depends on coordinated interactions between microspores and tapetal cells. While the CLAVATA3/EMBRYO SURROUNDING REGION-related 19 (CLE19) peptide has been identified as a microspore-derived "brake" that restricts tapetal activity to maintain exine developmental homeostasis, how CLE19 integrates with hormonal signaling pathways remains poorly understood. Here, we demonstrate that CLE19 attenuates brassinosteroid (BR) signaling output by engaging a defined BSL-BIN2-BES1 signaling cascade. Through quantitative phosphoproteomic analysis, we identified that CLE19 affects the phosphorylation of multiple BR signaling components, including BSL-type phosphatases BSL1/2/3, the GSK3-like kinase BIN2, and the transcription factor BES1. We show that CLE19 is perceived by its receptor PXL1, which directly interacts with BSL-type phosphatases to activate the GSK3-like kinase BIN2, leading to phosphorylation of BES1 at serine residues S219 and S223. Functional analyses using phospho-dead and phospho-mimic BES1 variants confirm that CLE19-dependent phosphorylation controls BES1 nuclear export and degradation, ultimately suppressing BR-responsive transcriptional outputs required for pollen exine patterning. Together, our findings define a peptide-hormone signaling axis that regulates transcription factor activity through post-translational regulation, providing mechanistic insight into how developmental robustness is maintained via intercellular signal integration in plant reproduction.