OSCA1 is an osmotic specific sensor: a method to distinguish Ca²⁺ -mediated osmotic and ionic perception

Genetic mutants defective in stimulus-induced Ca²⁺ increases have been gradually isolated, allowing the identification of cell-surface sensors/receptors, such as the osmosensor OSCA1. However, determining the Ca²⁺ -signaling specificity to various stimuli in these mutants remains a challenge. For instance, less is known about the exact selectivity between osmotic and ionic stresses in the osca1 mutant. Here, we have developed a method to distinguish the osmotic and ionic effects by analyzing Ca²⁺ increases, and demonstrated that osca1 is impaired primarily in Ca²⁺ increases induced by the osmotic but not ionic stress. We recorded Ca²⁺ increases induced by sorbitol (osmotic effect, OE) and NaCl/CaCl₂ (OE + ionic effect, IE) in Arabidopsis wild-type and osca1 seedlings. We assumed the NaCl/CaCl₂ total effect (TE) = OE + IE, then developed procedures for Ca²⁺ imaging, image analysis and mathematic fitting/modeling, and found osca1 defects mainly in OE. The osmotic specificity of osca1 suggests that osmotic and ionic perceptions are independent. The precise estimation of these two stress effects is applicable not only to new Ca²⁺ -signaling mutants with distinct stimulus specificity but also the complex Ca²⁺ signaling crosstalk among multiple concurrent stresses that occur naturally, and will enable us to specifically fine tune multiple signal pathways to improve crop yields.