IDYLYK

In the context of sustainable agriculture limiting pesticides, it is essential to understand the perception of microbe-associated molecular patterns (MAMPs) by surface receptors that regulate plant cell responses to microbes. However, the precise cellular and molecular mechanisms by which receptors are governed on the surface and inside plant cells are not always well understood. This project aims to better describe these mechanisms that could modulate plant-microbe interactions. We will focus on VvLYK receptors of grapevine (Vitis vinifera) involved in the recognition of chitin, the major component of the fungus cell wall.

Grapevine cells expressing VvLYKs fused to fluorescent proteins will be used to i) analyze receptor lateral movement on the surface quantified by Fluorescent Recovery After Photobleaching (FRAP) in presence of chitin, and ii) investigate protoplast membrane topography using correlative Atomic Force Microscope-fluorescence (AFM-fluorescence) with tip bound to chitin allowing the simultaneous membrane mapping and the co-localization of fluorescent receptors. We will also scrutinize protoplasts from leaves of diverse grapevine cultivars that present a range of sensitivity to fungal disease, such powdery mildew caused by Erysiphe necator. As proof of concept, we hypothesize that the abundance or mapping of chitin receptors is one of the factors that could be involved in the difference of sensitivity.

In parallel, the spatio-temporal intracellular fate of receptors from heterocomplexes will be followed after chitin treatment using fluorescent confocal laser scanning and transmission electronic microscopies. Our preliminary findings shown differential internalization of receptors for the same complex that we hypothesize to be a strategy to finely regulate response to microbes.

In addition, we propose to use acoustic AFM (UA-AFM) that offer complementary information on subcellular characteristics before and after MAMP leading to an original analysis of plant cells in condition mimicking microbe interactions.

This multidisciplinary project in line with the HARMI WP.2 Task 2.2 will add to the understanding of the fine regulation of plant immune receptors facing microbes.