CaM4 Functions as a Positive Regulator of the SOS Pathway in Response to Salt Stress

Salt poses a major environmental threat to plant growth and development. In Arabidopsis thaliana (Arabidopsis), salt overly sensitive (SOS) is a major Ca²⁺-activated salt-responsive signalling pathway defined by four main components: SOS1, SOS2, SOS3 and SOS3-like calcium-binding protein 8 (SCaBP8). Previously, we reported that a calmodulin (CaM) isoform, CaM4, functions in salt resistance by reestablishing the ion balance in Arabidopsis. Here, we showed that CaM4 directly binds to SOS2 (core element of the SOS pathway) both in vivo and in vitro and stimulates its kinase activity under salt stress. This finding was supported by the observation of CaM4 aggregation in the plasma membrane (PM) in response to NaCl. Genetic evidence has revealed that the overexpression of an activated SOS2 protein (SOS2^T168D or SOS2^DF) partially rescued the salt-sensitive phenotype of cam4 plants. Thus, SOS2 is considered a CaM4 target under saline conditions. Biological analyses have indicated that CaM4 activates SOS1 (a PM Na⁺/H⁺ antiporter) through SOS2 and enhances the interaction between SOS2 and SOS1, leading to the expulsion of excess Na⁺ from plant cells. A deficiency in CaM4 further reduced SOS2 kinase activity in scabp8sos3 double mutant plants. Thus, CaM4, together with SOS3 and SCaBP8, coregulate SOS2 kinase activity in Arabidopsis to promote its adaptation to salt stress. Collectively, our results show that CaM4 is a positive regulator of the SOS signalling pathway, which promotes plant salt tolerance.