Identification of Phosphorylation and Protein Interactions of Plant Pattern Recognition Receptors Through Immunoprecipitation.

Plasma membrane-localized pattern-recognition receptors (PRRs) play a crucial role in detecting pathogen-associated molecular patterns (PAMPs). Among them, leucine-rich repeat (LRR) receptor-like kinases (RLKs) are particularly important in plants. However, their structural complexity and membrane association pose significant challenges for biochemical analysis. Many LRR-RLKs exhibit low expression levels in planta, and their hydrophobic surfaces contribute to instability, often requiring detergent-based extraction. These properties complicate co-immunoprecipitation (Co-IP), hindering the identification of interacting proteins and post-translational modifications. Recent advancements in immunoprecipitation technologies have improved the isolation of membrane proteins, enabling the analysis of protein-protein interactions and phosphorylation dynamics. Here, we describe an optimized immunoprecipitation protocol for studying plant plasma membrane receptor-like kinases, facilitating robust characterization of their interactomes and regulatory modifications.