The ELD4-OsPRR95 module represses OsMADS51 in regulating rice heading date.

Heading date is a key agronomic trait that affects crop yield and regional adaptability. In this study, we identified a rice (Oryza sativa) early heading mutant and cloned the causal heading inhibitor gene EARLY HEADING AT LONG DAY 4 (ELD4) using the MutMap method. The eld4 CRISPR mutants and ELD4 RNAi plants flowered earlier than the wild type under natural long-day (NLD) conditions. ELD4 is a zinc finger transcription factor that localizes in the nucleus. Biochemical and genetic evidence suggests that ELD4 physically interacts with the pseudo-response regulator (PRR) protein OsPRR95. The Osprr95 mutant also exhibits an earlier heading phenotype under NLD. ELD4 and OsPRR95 co-regulate heading date by directly binding to the promoter and the first intron of OsMADS51, which promotes heading date. Moreover, electrophoretic mobility shift and luciferase complementation assays showed that ELD4 enhances OsPRR95 binding to the first intron of OsMADS51. These results suggest that the ELD4-OsPRR95 module directly represses OsMADS51 in regulating flowering time. Haplotype analysis reveals that OsPRR95 haplotype 3, which is geographically distributed in the north, exhibits a shorter heading date than haplotype 1 found in the south. This indicates that selection at the OsPRR95 locus has enhanced the regional adaptability of rice.