Heat-Triggered Dormancy Release in Low-ROS Pollen Grains Reveals a Conserved Reproductive Reserve

Pollen development and fertilization are considered the most heat-sensitive stages of plant reproduction. While heat stress severely impairs pollen germination and tube growth, the physiological diversity within a single flower’s pollen load suggests that subpopulations may exhibit differential climate resilience. In this study, we tested the hypothesis that this heterogeneity reflects a dormancy-based reserve mechanism that preserves fertilization under heat stress. Using flow cytometry and fluorescence-activated cell sorting in Arabidopsis thaliana and Solanum lycopersicum (MicroTom), we resolved pollen subpopulations by reactive oxygen species (ROS) status and examined their behavior under increasing heat stress. In both species, ROS-defined metabolic state was tightly associated with pollen size: high-ROS pollen was larger and readily germination-competent, whereas low-ROS pollen was smaller and showed low basal germination, consistent with dormancy. Heat stress preferentially depleted the high-ROS fraction, whereas the low-ROS fraction persisted and, under heat stress, increased metabolic activity and size. By isolating low-ROS and high-ROS pollen, we further show that a brief heat treatment suppresses germination of active high-ROS pollen but promotes germination of dormant low-ROS pollen. These findings provide direct evidence that heat can release dormancy in low-ROS pollen and support a conserved model in which dormant pollen serves as a heat-resilient reproductive reserve.