A genetic screen for dominant chloroplast reactive oxygen species signaling mutants reveals life stage-specific singlet oxygen signaling networks

Abstract

AbstractPlants employ intricate molecular mechanisms to respond to abiotic stresses, which often lead to the accumulation of reactive oxygen species (ROS) within organelles such as chloroplasts. Such ROS can produce stress signals that regulate cellular response mechanisms. One ROS, singlet oxygen (1O2), is predominantly produced in the chloroplast during photosynthesis and can trigger chloroplast degradation, programmed cell death (PCD), and retrograde (organelle-to-nucleus) signaling. However, little is known about the molecular mechanisms involved in these signaling pathways or how many different signaling1O2pathways may exist. TheArabidopsis thaliana plastid ferrochelatase two(fc2) mutant conditionally accumulates chloroplast1O2, makingfc2a valuable genetic system for studying chloroplast1O2-initiated signaling. Here, we have used activation tagging in a new forward genetic screen to identify eight dominantfc2activation-tagged (fas) mutations that suppress chloroplast1O2-initiated PCD. Whilefc2 fasmutants all block1O2-triggered PCD in the adult stage, only twofc2 fasmutants block such cellular degradation at the seedling stage, suggesting that life-stage-specific1O2-response pathways exist. In addition to PCD,fasmutations generally reduce1O2-induced retrograde signals. Furthermore,fasmutants have enhanced tolerance to excess light, a natural mechanism to produce chloroplast1O2. However, general abiotic stress tolerance was only observed in onefc2 fasmutant (fc2 fas2). Together, this suggests that plants can employ general stress tolerance mechanisms to overcome1O2production but that this screen was mostly specific to1O2signaling. We also observed that salicylic acid (SA) and jasmonate (JA) stress hormone response marker genes were induced in1O2-stressedfc2and generally reduced byfasmutations, suggesting that SA and JA signaling is correlated with active1O2signaling and PCD. Together, this work highlights the complexity of1O2signaling by demonstrating that multiple pathways may exist and introduces a suite of new1O2signaling mutants to investigate the mechanisms controlling chloroplast-initiated degradation, PCD, and retrograde signaling.