REPHASE: REdox regulation of PHAse separation for plant Adaptation to Stressful Environments

ongoing

The proposed project explores the intriguing possibility of reactive oxygen species (ROS) regulating the formation of stress granules (SGs) during heat stress. Heat stress triggers the rapid accumulation of ROS, with H2O2 playing a pivotal role as a signalling molecule that induces oxidative post-translational modifications (Oxi-PTMs) to regulate protein activities. On the other hand, an increase in temperature leads to the formation of enigmatic droplets in the cytosol known as SGs. These membraneless organelles (MLOs) arise through liquid-liquid phase separation (LLPS), enabling the concentration of proteins, mRNA, and metabolites to facilitate cellular adaptation. ROS accumulation and SGs formation have traditionally been viewed as independent events in the heat stress response. However, recent studies have shed light on the significance of Oxi-PTMs as regulators of protein LLPS in plants. Moreover, my preliminary experiments have demonstrated that the chemical blocking of Oxi-PTMs impairs SG formation during heat stress. I propose that ROS accumulation during heat stress induces Oxi-PTMs on proteins, promoting LLPS and facilitating SG formation. The REPHASE project aims to address this hypothesis through two main objectives. Firstly, I will investigate the dynamics of the redox state within SGs during heat stress. Secondly, I will map the landscape of Oxi-PTMs in SGs shedding light on their distribution and functional implications. To achieve these goals, the fluorescent redox biosensor roGFP2-Orp1 and the proteomic OxiTRAP probe YAP1 will be targeted to SGs. Utilizing these innovative methods will enable a comprehensive exploration of the redox biology of SGs, leveraging the state-of-the-art proteomic core and live cell imaging facilities available at the host institute. By bridging the gap between redox signalling and SGs, the REPHASE project promises to provide valuable insights into the mechanisms underlying plant adaptation to environmental stress.