Arbuscular mycorrhiza (AM) symbiosis represents a fungi-plant association that facilitates the bi-directional exchange of metabolites such as inorganic phosphate by the host. AM fungi form complex feeding structures called arbuscules within the roots of most land plants. This leads to reprogramming events that culminate with the synthesis of a host-derived membrane that surrounds the arbuscule, the peri-arbuscular membrane (PAM). However, the process and dynamics of PAM biogenesis remains poorly characterised. Scientists of the EU-funded ENDOSYM (3-D quantitative modelling of eukaryotic endosymbiosis: a pioneering innovative imaging approach) project set out to develop novel bioimaging techniques that would permit live-cell deep-tissue 4-D imaging of membrane dynamics and high-resolution ultra-structural analyses. Researchers established an AM symbiosis model of rice and Rhizophagus irregularis and applied high resolution multi-photon confocal microscopy (MPCM) as well as ultra-structural tomography. To facilitate live-cell imaging, rice roots were engineered to express a fluorescent form of the mycorrhiza-colonised phosphate transporter, PT1. Additional subcellular fluorescent membrane markers were utilised for MPCM imaging. Results indicated for the first time that PT11 localisation at the PAM of developing arbuscules occurred at distinct foci surrounding the tips of fine arbuscule branches. Temporal imaging of PT11 localisation suggested that it either directly influences arbuscule branching or that the latter dictates the localisation of PT11. With respect to the 3D membrane structures, scientists tested various approaches for membrane preservation. Using high-pressure freezing, they observed novel fungal tubular membrane extensions resembling nanopodia and validated the complex nature of PAM. Overall, researchers demonstrated that AM symbiosis represents an excellent model system to elucidate molecular processes that underpin complex invader-mediated reprogramming of eukaryotic organisms. The developed imaging techniques could apply to other systems to provide vital structural and molecular information.
Symbiosis, Arbuscular mycorrhiza, ENDOSYM, Rhizophagus irregularis, PT11