Abstract: Hyperosmolality-triggered physiological drought hinders plant growth and development, leading to a drop in crop yields. Hyperosmolality triggers calcium signaling, and yet how osmotic-induced calcium signaling participates in cellular osmotic response remains enigmatic. To date, several Ca²⁺ channels and transporters have been identified to regulate osmotic-induced calcium signal generation (CaSG) or Ca²⁺ homeostasis. However, there has been no report on their function in calcium signal clearance (CaSC) in plants, especially in crops. Here, we investigated the role of a rice cation/calcium exchanger OsCCX2 in modulating calcium signaling dynamics using two distinct calcium reporters aequorin and GCaMP6s. The results showed that, under osmotic stress conditions, CaSC was significantly delayed in both root and guard cells of ccx2 mutants compared with the wild-type. Further studies revealed that hyperosmotic stress-triggered influxes of sodium (Na⁺), potassium (K⁺), and chloride (Cl⁻) ions were significantly reduced in ccx2 mutants, resulting in a significantly smaller range of osmotic pressure and water potentials (Ψ_w) adjustment. In addition, the stomatal response was impaired, with a faster water loss in ccx2 in response to hyperosmotic stress. Furthermore, the absence of OsCCX2 altered the expression patterns of key osmotic-responsive genes, but their transcriptional activation was unaffected. Collectively, these changes ultimately led to reduced hyperosmotic stress tolerance in the mutants. Additionally, OsCCX2 is likely to be located in the endoplasmic reticulum and plasma membrane, and possess Na⁺/Ca²⁺ exchange activity. To sum up, our findings provide evidence that OsCCX2, as a CaSC regulator, is involved in cell osmotic adjustment, water homeostasis and osmotic stress tolerance in rice, which offers new insight into potential applications in drought-resistant crop improvement.

Key words: calcium signal clearance (CaSC), Oryza sativa L, OsCCX2, osmotic adjustment