While there is tremendous knowledge about the plant immune system, and how the plant defends itself against pathogens, we lack spatial resolution. Plant immune responses are typically recorded via transcriptomics (RNA-seq or qRT-PCR using pooled total RNA/DNA), in vitro assays (Calcium burst in cuvette, kinase phosphorylation in-gel, ROS-burst in cuvette), cell death assays in heterologous system (leaves with infiltrated effectors or pathogens) or with markers using leaves of transiently transformed and expressed proteins. In all cases, the spatial resolution is lost, and the infection is far from natural (the pathogen or its effectors are infiltrated in leaves, or physical damage is done by hand).
To close this knowledge gap, we are using fluorescent markers for a wide array of plant immune pathways in live-imaging experiments. We allow Fusarium oxysporum, an important plant pathogen, to infect the plant by itself and in the tissue it naturally infects, without any infiltration, or dripping onto a certain tissue, by growing plant and pathogen in parallel in one sealed experimental dish. Once the fungus has infected the plants, we live-image to infection process and the immune responses of the plant via the different markers live and in real-time on a microscope. By doing so, we can map which immune responses are triggered at which time point, in which tissues, and in which cell types with cellular resolution, and while simultaneously imaging progression of the infection.
This assay has allowed us to create a map of certain immune pathways, and show how cells in direct contact with the infection site respond, how cells further away respond, and which tissues don't respond.
Further, we have started to analyze the plant's responses in a more natural environment, using 'real world' agricultural soil, rather than just laboratory medium or potting soil.
This spatial immunity approach is new and highly promising to understand the plant immune system on an individual cell level.
