Cellularization is a key event in endosperm development. Polycomb group (PcG) genes, such as Fertilization-Independent Seed 2 ( FIS2 ), are vital for the syncytium-to-cellularization transition in Arabidopsis plants. In this study, we found that OsEMF2a , a rice homolog of the Arabidopsis PcG gene Embryonic Flower2 ( EMF2 ), plays a role similar to that of FIS2 in regard to seed development, although there is limited sequence similarity between the genes. Delayed cellularization was observed in osemf2a , associated with an unusual activation of type I MADS-box genes. The cell cycle was persistently activated in osemf2a caryopses, which was likely caused by cytokinin overproduction. However, the overaccumulation of auxin was not found to be associated with the delayed cellularization. As OsEMF2a is a maternally expressed gene in the endosperm, a paternally inherited functional allele was unable to recover the maternal defects of OsEMF2a . Many imprinted rice genes were deregulated in the defective hybrid seeds of osemf2a (♀)/9311 (♂) (m9). The paternal expression bias of some paternally expressed genes was disrupted in m9 due to either the activation of maternal alleles or the repression of paternal alleles. These findings suggest that OsEMF2a-PRC2-mediated H3K27me3 is necessary for endosperm cellularization and genomic imprinting in rice.

Fertilization Independent Endosperm (FIE) is an essential member of Polycomb Repressive Complex 2 (PRC2) that plays important roles in the developmental regulation of plants. OsFIE1 and OsFIE2 are two FIE homologs in the rice genome. Here, we showed that OsFIE1 probably duplicated from OsFIE2 after the origin of the tribe Oryzeae, but has a specific expression pattern and methylation landscape. During evolution, OsFIE1 underwent a less intensive purifying selection than did OsFIE2. The mutant osfie1 produced smaller seeds and displayed reduced dormancy, indicating that OsFIE1 predominantly functions in late seed development. Ectopic expression of OsFIE1, but not OsFIE2, was deleterious to vegetative growth in a dose-dependent manner. The newly evolved N-terminal tail of OsFIE1 was probably not the cause of the adverse effects on vegetative growth. The CRISPR/Cas9-derived mutant osfie2 exhibited impaired cellularization of the endosperm, which suggested that OsFIE2 is indispensable for early seed development as a positive regulator of cellularization. Autonomous endosperm was observed in both OsFIE2 +À and osfie1/OsFIE2 +À but at a very low frequency. Although OsFIE1-PRC2 exhibited H3K27me3 methyltransferase ability in plants, OsFIE1-PRC2 is likely to be less important for development in rice than is OsFIE2-PRC2. Our findings revealed the functional divergence of OsFIE1 and OsFIE2 and shed light on their distinct evolution following duplication.

Fertilization Independent Endosperm (FIE) is an essential member of Polycomb Repression Complex 2 (PRC2) that plays important roles in the developmental regulation of plants. OsFIE1 and OsFIE2 are two FIE homologs in the rice genome. Here, we showed that OsFIE1 probably duplicated from OsFIE2 after the origin of the tribe Oryzeae, but has a specific expression pattern and methylation landscape. During evolution, OsFIE1 underwent a less intensive purifying selection than did OsFIE2. The mutant osfie1 produced smaller seeds and displayed reduced dormancy, indicating that OsFIE1 predominantly functions in late seed development. Ectopic expression of OsFIE1, but not OsFIE2, was deleterious to vegetative growth in a dosage-dependent manner. The newly evolved N-terminal tail of OsFIE1 was probably not the cause of the adverse effects on vegetative growth. The CRISPR/Cas9-derived mutant osfie2 exhibited impaired cellularization of the endosperm, which suggested that OsFIE2 is indispensable for early seed development as a positive regulator of cellularization. Autonomous endosperm was observed in both OsFIE2+− and osfie1/OsFIE2+− but at a very low frequency. Although OsFIE1-PRC2 exhibited H3K27me3 methyltransferase ability in plants, OsFIE1-PRC2 is likely to be less important for development in rice than is OsFIE2-PRC2. Our findings revealed the functional divergence of OsFIE1 and OsFIE2 and shed light on their distinct evolution following duplication.