# Comparative proteomic profiling of receptor kinase signaling reveals key trafficking components enforcing plant stomatal development

## TL;DR

This study maps the proteins involved in receptor kinase signaling during plant stomatal development, highlighting the role of endocytosis in regulating this process.

## Contribution

The study identifies specific endocytosis components that regulate ERECTA receptor kinase internalization during stomatal development.

## Key findings

- ERECTA interacts with TOO MANY MOUTHS and endosomal trafficking machinery via clathrin-mediated endocytosis.
- PICALM subfamily proteins synergistically regulate ERECTA internalization and stomatal patterning.
- Disruption of PICALMs impairs ERECTA endocytosis and leads to stomatal clustering.

## Abstract

Receptor kinases are pivotal for growth, development, and environmental response of plants. Yet, their regulatory mechanisms and spatial dynamics remain underexplored. The ERECTA-family receptor kinases coordinate diverse developmental processes, including stomatal development. To understand the proteomic landscape of the ERECTA-mediated signaling pathways, we report comparative analyses of the ERECTA interactome and proximitome by epitope-tagged affinity-purification (ET-AP) and TurboID-based proximity labeling (TbID-PL) mass spectrometry, respectively. While ET-AP recovered receptor complex components (e.g., TOO MANY MOUTHS), TbID-PL effectively captured transient associations with the components of endosomal trafficking, i.e., clathrin-mediated endocytosis machinery. We further identify that specific subfamily members of phosphatidylinositol-binding clathrin assembly proteins (PICALMs) interact with and synergistically regulate ERECTA internalization. Mutations in PICALMs impair ERECTA endocytosis and lead to excessive stomatal clustering by dampening the downstream signaling output. Together, we provide a proteomic atlas of the ERECTA signaling network and demonstrate that timely removal of receptor kinase by the endocytosis machinery is essential for active signal transduction enforcing stomatal patterning.

Proteomic mapping uncovers endocytosis adaptors that drive receptor internalization for robust stomatal development.