Oilseeds accumulate a large amount of storage lipids, which are used as sources of carbon and energy for seed germination and seedling growth. The storage lipids are accumulated in oil bodies during seed maturation. Oil bodies in seeds are surrounded with three oil-body-membrane protein families, oleosins, caleosins and steroleosins. These proteins are plant-specific and much abundant in seeds. Here we show a unique function of oleosins in preventing fusion of oil bodies and maintaining seed germination. Reverse genetic analysis using oleosin-deficient mutants shows the inverse proportion of oil-body sizes to total oleosin contents. The double mutant ole1 ole2 with the lowest levels of oleosins has irregularly-enlarged oil bodies throughout the seed cells, and hardly germinates. Germination rates are positively associated with oleosin contents, suggesting that the defects of germination are related to the expansion of oil bodies due to oleosin deficiency. Interestingly, freezing treatment followed by imbibition at 4 degrees C inhibits seed germination of single mutants (ole1 and ole2), which germinate normally without freezing treatment. The freezing treatment accelerates the fusion of oil bodies and generates eccentric nuclei in ole1 seeds, which caused seed mortality. Taken together, our findings suggest that oleosins increase the viability of oilseeds by preventing abnormal fusion of oil bodies for overwintering. Knowledge of oleosin contributes a great deal to not only an insight into freezing tolerance of oilseeds, but also creating genetically modified plants for developing a bioenergy and biomass resource.