SYP22 binding with BIN2 modulates vacuolar fusion in guard cells and affects stomatal opening in Arabidopsis

Stomatal movement confers plants the ability to regulate gas exchange, water loss, pathogen defense and photosynthesis in response to diverse environmental signals. Rapid changes in the stomatal aperture are accompanied by adjustment of vacuole volume and morphology in guard cells (GCs). Fusion of small vacuoles is known to be required for stomatal opening. However, the regulation of vacuolar dynamics in GCs represents a largely uncharted area for understanding stomatal function and crop improvement. Here, we combined genetic, cytological and molecular approaches to investigate the regulatory mechanisms of vacuolar fusion during stomatal opening in Arabidopsis thaliana. We report that both the loss function and overexpression of SYP22, a Qa-SNARE that facilitates homotypic vacuole fusion, repress vacuolar fusion in GCs and stomatal opening. BIN2 interacts with SYP22 and decreases the stability of SYP22 via phosphorylation. Further investigation reveals that the loss-of-function mutants of BIN2 and its homologs, bin2-3 bil1 bil2, promote vacuolar fusion and stomatal opening, whereas the gain-of-function mutant, bin2-1, exhibited restricted vacuolar fusion and stomatal opening. Our findings uncover that vacuolar fusion regulated by the BIN2-SYP22 module is required for stomatal opening, providing insights for breeding to improve plant environmental adaptability.