Abstract

Within the roots of most land plants beneficial arbuscular mycorrhiza fungi (AMF) form complex tree-shaped feeding structures called arbuscules. Monumental cellular re-differentiation and reprogramming in the inner root cortex result in the de novo synthesis of a host-derived membrane that surrounds the arbuscule, the peri-arbuscular membrane (PAM). This functional symbiosome interface facilitates nutrient exchange between fungus and plant. The extent to which fungal and host membranes are tailored for this plant-fungal dialogue and our understanding of how different PAM -specific proteins are retained within the PAM remain unknown. This is partly due to the low resolution of live-cell imaging of inner cortical rice root cell layers using conventional confocal laser scanning microscopy (CLSM). The aim of this project is to 1.) develop innovative imaging approaches to investigate ultra-structural modifications to plant and fungal membranes during arbuscule differentiation and 2.) to pioneer high resolution live-cell bioimaging to investigate dynamic changes to PAM -specific proteins during arbuscule development. High pressure frozen (HPF) Transmission Electron Microscopy (TEM) of rice roots colonized by Rhizophagus irregularis has uncovered exciting fungal membrane structures that resemble Extracellular Vesicles (EVs) at the AMF -plant interface that are capable of passage through the fungal cell wall into the peri-arbuscular space. Multi-photon confocal microscopy (MPCM) was developed for rice roots to allow deep-tissue time-lapse imaging of fluorescently-tagged PAM proteins during arbuscule development. Results indicate that separate spatio-temporal mechanisms control trafficking of functionally diverse PAM proteins during arbusucle development.