Abstract: The early morphogenesis of the embryo and endosperm sets the upper limit of rice grain yield; yet, the cellular dynamics and regulatory mechanisms underlying this critical stage remain largely elusive. Here, we present a single-cell atlas of developing rice seeds based on single-nucleus RNA sequencing of 67,922 high-quality nuclei from rice caryopses. Integration with bulk RNA-seq, in situ hybridization, and promoter-GUS staining enabled systematic cell-type annotation and revealed a distinct population of embryo-endosperm interface (EEI) cells occupying the boundary between the developing embryo and endosperm. Comparative analyses with maize data sets, together with trajectory inference, suggested that a subset of EEI cells shares molecular features with maize embryo-adjacent scutellum cells and shows transcriptional continuity with starchy endosperm cells. Embryo-endosperm interface cells were enriched in transport-related and developmental regulatory genes with known functions in seed development and embryogenesis. Focusing on the EEI-enriched regulator OsBZR4, we found that loss of OsBZR4 altered cellular composition and transcriptional programs during early seed development, disrupted embryonic developmental progression, and reduced the expression of embryonic genes, including OsCDP3.10 and RINO1. Together, our study provides a single-cell resolution framework for understanding early rice seed development, identifies the embryo-endosperm interface as an important cellular domain associated with embryogenesis, and offers a valuable resource for dissecting the molecular basis of seed formation in rice and related cereals.

Key words: embryo-endosperm interface, embryogenesis, endosperm development, rice early seed development, single-nucleus RNA sequencing