Plasma membrane CBL Ca2+ sensor proteins function in regulating primary root growth and nitrate uptake by affecting global phosphorylation patterns and microdomain protein distribution.

The collective function of Calcineurin B-like (CBL) Ca2+ sensors and CBL interacting protein kinases (CIPKs) in decoding plasma membrane initiated Ca2+ signals to convey developmental and adaptive responses to fluctuating nitrate availability remained to be determined. Here, we generated a cbl-quintuple mutant in Arabidopsis thaliana devoid of these Ca2+ sensors at the plasma membrane and performed comparative phenotyping, nitrate flux determination, phosphoproteome analyses and studies of membrane domain protein distribution in response to low and high nitrate availability. We observed that CBL proteins exert multifaceted regulation of primary and lateral root growth and nitrate fluxes. Accordingly, we found that loss of plasma membrane Ca2+ sensor function simultaneously affected protein phosphorylation of numerous membrane proteins including several nitrate transporters, proton pumps and aquaporins as well as their distribution within plasma membrane microdomains and identified specific phosphorylation and domain distribution pattern during distinct phases of low and high of nitrate responses. Collectively, these analyses reveal a central and coordinative function of CBL-CIPK mediated signalling in conveying plant adaptation to fluctuating nitrate availability and identify a crucial role of Ca2+ signalling in regulating the composition and dynamics of plasma membrane microdomains.