Calmodulin binds to extracellular sites on the plasma membrane of plant cells and elicits a rise in intracellular calcium concentration.

Calmodulin (CaM) is a highly conserved intracellular calcium sensor. In plants, CaM also appears to be present in the apoplasm, and application of exogenous CaM has been shown to influence a number of physiological functions as a polypeptide signal; however, the existence and localization of its corresponding apoplasmic binding sites remain controversial. To identify the site(s) of action, a CaM-conjugated quantum dot (QD) system was employed for single molecule level detection at the surface of plant cells. Using this approach, we show that QD-CaM binds selectively to sites on the outer surface of the plasma membrane, which was further confirmed by high resolution transmission electron microscopy. Measurements of Ca(2+) fluxes across the plasma membrane, using ion-selective microelectrodes, demonstrated that exogenous CaM induces a net influx into protoplasts. Consistent with these flux studies, calcium-green-dextran and FRET experiments confirmed that applied CaM/QD-CaM elicited an increase in cytoplasmic Ca(2+) levels. These results support the hypothesis that apoplasmic CaM can act as a signaling agent. These findings are discussed in terms of CaM acting as an apoplasmic peptide ligand to mediate transmembrane signaling in the plant kingdom.