In planta exploitation of leaf apoplastic compounds: a window of opportunity for spatiotemporal studies of apoplastic metabolites, hormones and physiology

Processes in the leaf apoplast are relevant for development, cell wall rheological properties, plant nutrition, sink-source portioning, microbe-host plant-interactions or intercellular information exchange and signaling and are therefore regulated or influenced by the composition of the leaf apoplastic solute. In contrast to the traditional methods for the extraction of apoplastic solutes that are more or less destructive, we propose a new method that allows extraction of leaf apoplastic solutes (i) non-invasively and, thus, (ii) over time. Moreover, the method has (iii) a high spatial resolution that allows identification of solute-microdomains in the leaf apoplast. The method was established for Arabidopsis thaliana and Vicia faba leaves but should also be applicable to other plants species with similar leaf morphologies. It is based on the infiltration of an aqueous extraction solution into the apoplast followed by its recovery seconds later, both through the stomata. By this, the apoplast (and its solutes) of an identical leaf can be sampled on successive days with negligible symplastic contamination. A spatiotemporal mapping of leaf apoplastic ion and metabolite patterns within the identical leaf opens a window of opportunity for understanding apoplast biology. As for example, the existence of apoplastic abscisic acid gradients within a leaf in response to salinity was witnessed in this study, as was the unsuspected accumulation of kaempferol glycosides in the leaf apoplast. The presented method is relevant for plant developmental biologists, phytopathologists, plant physiologists, plant nutritionists and others that need to integrate apoplast biology into their research approaches.