Salas-Gomez, M. | Search bioRxiv

bioRxiv2022

Plant autophagosomes mature into amphisomes prior to their delivery to the central vacuole

Autophagosomes are double-membraned vesicles that traffic harmful or unwanted cellular macromolecules to the vacuole for recycling. Although autophagosome biogenesis has been extensively studied, mechanisms of autophagosome maturation, i.e., delivery and fusion with the vacuole, remain largely unknown in plants. Here, we have identified an autophagy adaptor, CFS1, that directly interacts with the autophagosome marker ATG8 and localizes on both membranes of the autophagosome. Autophagosomes form normally in Arabidopsis thaliana cfs1 mutants, but their delivery to the vacuole is disrupted. CFS1s function is evolutionarily conserved in plants as it also localizes to the autophagosomes and plays a role in autophagic flux in the liverwort Marchantia polymorpha. CFS1 regulates autophagic flux by connecting autophagosomes with the ESCRT-I component VPS23, leading to the formation of amphisomes. Disrupting the VPS23-CFS1 interaction affects autophagic flux and renders plants sensitive to starvation stress. Altogether, our results reveal a deeply conserved mechanism of vacuolar delivery in plants that is mediated by amphisomes.

plant biology↗

bioRxiv2021

Robust transcriptional indicators of plant immune cell death revealed by spatio-temporal transcriptome analyses

Recognition of a pathogen by the plant immune system often triggers a form of regulated cell death traditionally known as the hypersensitive response. This type of immune 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? As a consequence, bona fide transcriptional indicators of immune cell death 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 immune cell death 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 immune cell death 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 immune cell death 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 immune cell death-triggering bacteria, opening new avenues for specific and/or high-throughput techniques to study immune cell death processes at a single-cell level.

plant biology↗