Singlet oxygen leads to structural changes to chloroplasts during degradation in the Arabidopsis thaliana plastid ferrochelatase two mutant

Abstract

During photosynthesis, chloroplasts can produce large amounts of reactive oxygen species (ROS), particularly under stressful conditions. Along with other nutrients, chloroplasts also contain 80% of a leafs nitrogen supply. For these reasons, chloroplasts are prime targets for cellular degradation to protect cells from photo-oxidative damage and to redistribute nutrients to sink tissues. Multiple chloroplast degradation pathways have been described and are induced by photo-oxidative stress and nutrient starvation. However, the mechanisms by which damaged or senescing chloroplasts are identified, transported to the central vacuole, and ultimately degraded are not well characterized. Here, we investigated the subcellular structures involved with degrading chloroplasts induced by the ROS singlet oxygen (1O2) in the Arabidopsis thaliana plastid ferrochelatase two (fc2) mutant. Using a three-dimensional serial-block face electron microscopy analysis, we show up to 35% of degrading chloroplasts in fc2 mutants protrude into the central vacuole. While the location of a chloroplast within a cell had no effect on the likelihood of its degradation, chloroplasts in spongy mesophyll cells were degraded at a higher rate than those in palisade mesophyll cells. To determine if degrading chloroplasts have unique structural characteristics allowing them to be distinguished from healthy chloroplasts, we analyzed fc2 seedlings grown under different levels of photo-oxidative stress. A clear correlation was observed between chloroplast swelling, 1O2-signaling, and the state of degradation. Finally, plastoglobule enzymes involved in chloroplast disassembly were shown to be upregulated while plastoglobules increased their association with the thylakoid grana, implicating an interaction between 1O2-induced chloroplast degradation and senescence pathways.