Glutathione S -transferases protect against heat-inhibited pollen germination and pollen tube growth in the pistil by regulating proanthocyanidins and fructose to maintain reactive oxygen species homeostasis

Glutathione S-transferases (GSTs) participate in diverse stress responses, but their function in heat responses remains poorly understood. Two rice varieties differing in heat tolerance, Yongyou15 (YY15) and Y-Liangyou1 (YLY1), were subjected to heat stress during anthesis. YY15 exhibited higher spikelet fertility than YLY1 under heat stress, primarily due to superior pollen germination on the stigma, enhanced pollen tube growth within the pistil, and lower reactive oxygen species (ROS) levels in the pistil. Transcriptome analysis and physiological assessments revealed that GSTs, glutathione (GSH), proanthocyanidins (PAs), and fructose play pivotal roles in mediating the heat tolerance differences between the varieties. Specifically, these factors confer heat tolerance by maintaining ROS homeostasis. Exogenous GST enhancers, myristic acid (MA) and fulvic acid (FA), significantly improved spikelet fertility, GST activity, pollen germination, and pollen tube growth. They also substantially increased GSH content, PA levels, fructose content, and antioxidant capacity, while concurrently reducing ROS levels. Conversely, application of the GST inhibitor ethacrynic acid reversed these beneficial effects. Importantly, exogenous application of FA, PAs, or fructose-either individually or in combination-significantly enhanced spikelet fertility under heat stress. This indicates that GSTs, mediating GSH metabolism, play a crucial role in preventing heat-induced pistil dysfunction through the PA and fructose pathways.