Male and female gametophytes meet after winter: Testing the overwintering hypothesis for delayed fertilization in Japanese stone oak

PremiseFertilization delays of nearly a year have been observed in many acorn-producing Fagaceae trees. Despite this long-standing recognition, the underlying mechanisms and adaptive significance of delayed fertilization remain largely unexplored. It has been hypothesized that these prolonged delays may serve as an adaptive strategy, allowing ovules to be fertilized and seeds to develop during more favorable seasons, thereby avoiding the adverse conditions of winter. However, empirical evidence to support this hypothesis is still lacking. MethodsTo test the overwintering hypothesis, we observed the seasonal progression of pollen tube growth and ovule development in Lithocarpus edulis, a species exhibiting two flowering seasons, spring and autumn. We conducted monthly observations of pollen tube growth and ovule development in both spring and autumn flower samples. We employed microtome techniques and scanning confocal microscopy to closely examine the seasonal progression of pollen tube growth and ovule development. ResultsWe found that pollen tubes were arrested at the style-joining site, and ovules remained immature in both spring and autumn samples prior to winter. Following winter, pollen tube regrowth and ovule maturation were synchronized in the subsequent spring in both spring and autumn samples. ConclusionsOur results support the overwintering hypothesis, suggesting that delayed fertilization may serve as an adaptive strategy to ensure fertilization in spring, overcoming the reproductive challenges posed by cold winter conditions. Our study introduces a new avenue for studying the temporal organization of plant reproductive phenology, specifically the alignment of flowering, pollination, fertilization, and fruiting in seasonal environments.