Characterization of mitochondrial dynamics and subcellular localization of ROS reveal that HsfA2 alleviates oxidative damage caused by heat stress in Arabidopsis.

Heat shock transcription factor A2 (HsfA2) participates in multiple stress responses. To provide new insights into the role of HsfA2 in the heat stress (HS) response, in vivo production and localization of reactive oxygen species (ROS) and mitochondrial dynamics were investigated during the onset of cell death induced by an HS (40 degrees C, 10 min) applied after a 2 d recovery at 24 degrees C following a conditioning treatment at 37 degrees C for 1 h. In response to the HS, generated ROS were significantly higher in hsfA2 than in wild-type (WT) protoplasts and did not return to the baseline level when compared with WT protoplasts. The uncontrolled ROS in hsfA2 protoplasts localized not only to mitochondria but also to chloroplasts. Microscopic observations also revealed that, prior to cell death, hsfA2 protoplasts underwent more severe alterations in mitochondrial dynamics than WT protoplasts, including mitochondrial swelling, transmembrane potential loss, and the cessation of mitochondrial movement. The lower cell viability in hsfA2 than in WT protoplasts suggested that--combined with the findings that antioxidants only partially blocked ROS generation and arrested cell death in hsfA2 protoplasts relative to WT protoplasts--ROS participated in HS-induced cell death. Also the disruption of HsfA2 resulted in more severe oxidative stress and more cell death which, together with the more severe alterations in mitochondrial dynamics, could be complemented by introducing a WT copy of HsfA2. These results represent the first subcellular evidence that HsfA2 protects plants against HS-induced oxidative damage, organelle dysfunction, and subsequent cell death.