Abstract

Recognition of a pathogen by the plant immune system often triggers a form of regulated cell death traditionally known as the hypersensitive response (HR). This type of cell death occurs precisely at the site of pathogen recognition, and it is restricted to a few cells. Extensive research has shed light into how plant immune receptors are mechanistically activated. However, a central key question remains largely unresolved: how does cell death zonation take place and what are the mechanisms that underpin this phenomenon? Consequently, bona fide transcriptional indicators of HR are lacking, which prevents gaining a deeper insight of its mechanisms before cell death becomes macroscopic and precludes any early or live observation. We addressed this question using the paradigmatic Arabidopsis thaliana-Pseudomonas syringae pathosystem, by performing a spatio-temporally resolved gene expression analysis that compared infected cells that will undergo HR upon pathogen recognition vs by-stander cells that will stay alive and activate immunity. Our data revealed unique and time-dependent differences in the repertoire of differentially expressed genes, expression profiles and biological processes derived from tissue undergoing HR and that of its surroundings. Further, we generated a pipeline based on concatenated pairwise comparisons between time, zone and treatment that enabled us to define 13 robust transcriptional HR markers. Among these genes, the promoter of an uncharacterized AAA-ATPase has been used to obtain a fluorescent reporter transgenic line, which displays a strong spatio-temporally resolved signal specifically in cells that will later undergo pathogen-triggered cell death. In sum, this valuable set of genes can be used to define those cells that are destined to die upon infection with HR-triggering bacteria, opening new avenues for specific and/or high-throughput techniques to study HR processes at a single-cell level.

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