OsCPK9-mediated Thr105 phosphorylation activates OsCATC to enhance drought tolerance in rice

Abstract

Introduction: Drought poses a significant environmental challenge, disrupting plant growth and reducing crop productivity. As sensors and effectors in calcium signaling, calcium-dependent protein kinases (CPKs) regulate plant development and environmental adaptation. However, the specific roles and molecular networks of many OsCPKs in rice adaptation to abiotic stress remain to be elucidated.

Objectives: Drought-tolerant rice holds significant potential for enhancing yield stability and global food security. The study aimed to isolate drought-responsive OsCPKs and explore their molecular mechanisms.

Methods: To elucidate the mechanisms underlying OsCPK9-mediated drought tolerance, we integrated assays of Co-immunoprecipitation, yeast two-hybrid, GST pull-down and bimolecular fluorescence complementation to validate the OsCPK9-interacting proteins. In vitro protein phosphorylation analysis was conducted to identify phosphorylated substrates and sites. Rice yield performance was evaluated through three-year field trials.

Results: OsCPK9 overexpressing plants enhanced drought tolerance in rice, accompanied by lower H₂O₂ content and higher catalase activity than wild-type (WT) plants. Conversely, OsCPK9 knockout plants exhibited sensitive to drought stress compared with WT plants. Myristoylation and palmitoylation contributed to plasma membrane localization of OsCPK9, which is required for drought tolerance. OsCPK9 interacts with and phosphorylates Catalase C (OsCATC) at the plasma membrane. Phosphorylation of OsCATC at the highly conserved Thr105 enhanced its self-interaction, promoting oligomer formation and thereby increasing catalytic activity. OsCATC overexpressing plants showed higher catalase activity and improved drought tolerance compared to WT plants. Moreover, OsCPK9 overexpressing plants mitigated a 17.6% average yield loss per plant relative to WT plants.

Conclusion: This study reveals that myristoylation and palmitoylation of OsCPK9 contribute to its plasma membrane localization and drought tolerance. The post-translational regulatory mechanism mediated by OsCPK9 enhances environmental adaptability and provides a promising strategy to improve rice grain yield under drought stress conditions.

Keywords: Drought stress tolerance; H(2)O(2); OsCATC; OsCPK9; Phosphorylation; Rice.