Quantifying Plasmodesmatal Transport with an Improved GFP Movement Assay.

Abstract

Plasmodesmata (PD) are membrane-lined channels that cross the cell wall to connect the cytosol of adjacent plant cells, permitting diverse cytosolic molecules to move between cells. PD are essential for plant multicellularity, and the regulation of PD transport contributes to metabolism, developmental patterning, abiotic stress responses, and pathogen defenses, which has sparked broad interest in PD among diverse plant biologists. Here, we present a straightforward method to reproducibly quantify changes in the rate of PD transport in leaves. Individual cells are transformed with Agrobacterium to express fluorescent proteins, which then move beyond the transformed cell via PD. Forty-eight to 72h later, the extent of GFP movement is monitored by confocal fluorescence microscopy. This assay is versatile and may be combined with transient gene overexpression, virus-induced gene silencing, physiological treatments, or pharmaceutical treatments to test how PD transport responds to specific conditions. We expect that this improved method for monitoring PD transport in leaves will be broadly useful for plant biologists working in diverse fields.