Heat stress accelerates the development of hypoxia and hypoxia-induced ferroptosis-like cell death in barley root tip under waterlogging

The root tip is extremely sensitive to waterlogging, where the hypoxic conditions evoke an extensive cell death within a few hours. Using a pharmacological approach, we demonstrated that hypoxia-induced cell death in barley root tips exhibits several key features of ferroptosis, including iron-dependent ROS generation, enhanced lipoxygenase activity and lipid peroxidation. We also observed that calcium influx from the apoplast into the cytoplasm played a critical role in the initiation of ferroptosis-like cell death. In the prevention of hypoxia-induced root death, the most effective were the iron chelator deferoxamine, the lipoxygenase inhibitor naproxen and the Ca 2+ chelator EGTA. Hypoxia-induced lipid peroxidation and cell death started in the root transition zone, including the surrounding meristem and elongation zones, and subsequently spread to nearly the entire root tip. Heat stress accelerated the onset and development of hypoxia in waterlogged barley root tips, consequently also accelerating the progression of root cell death. This waterlogging/hypoxia-induced ferroptosis-like cell death in barley root tips was triggered within 1–2 hours, even at a moderately elevated temperature of 30 °C, indicating that climate changes leading to the co-occurrence of high temperature and heavy rainfall in the future may be detrimental to crop production, including barley. • Our results demonstrate that heat stress accelerated the development of hypoxia, hence also hypoxia-induced cell death in barley root tips under waterlogging. This hypoxia-induced cell death in barley root tips shows several attributes of ferroptosis, like Fe-dependent ROS generation, lipid peroxidation and enhanced LOX activity. In addition, extracellular Ca 2+ influx into the cytoplasm is involved in the initiation of ferroptosis-like cell death in hypoxic root tips.