ZEP1 orchestrates template choice and crossover pathways to ensure meiotic genome integrity in rice.

Meiotic crossovers (COs) are tightly regulated to ensure chromosome segregation while limiting aberrant recombination. Transverse filament (TF) proteins of the synaptonemal complex (SC) regulate class I crossoverts, yet how the SC coordinates homologous recombination (HR) to maintain faithful recombination remains unclear. Here, we show that loss of rice TF protein ZEP1 does not uniformly enhance HEI10-marked class I COs; instead, ZEP1 null mutants display asynapsis, multivalent formation, and chromosome fragmentation. These defects depend on double-strand breaks (DSBs) and genetically place ZEP1 function after strand invasion. ZEP1 interacts with anti-crossover factors MEICA1, FIGNL1, and RMI1, and is required for their enrichment at the synaptonemal complex; its loss causes persistent DMC1/RAD51 signals, indicating dysregulated strand invasion. Genetic interactions further support impaired recombination intermediate homeostasis, with ZMM removal partially alleviating chromosome abnormalities, while MUS81 becomes increasingly essential. We propose that the synaptonemal complex functions as a structural hub, where ZEP1 concentrates anti-crossover modules that restrain invasion and coordinate recombination intermediate processing to promote HR repair. Furthermore, ZEP1 dosage may provide a constrained lever to tune CO outcomes in a background-dependent manner.