Fusion-Derived Centromeres Reveal Satellite Repeat Remodeling in Miscanthus sinensis

Centromeres are essential for chromosome segregation, yet their repetitive nature makes them among the most difficult genomic regions to assemble and interpret. In polyploid plants, centromeres provide a unique record of chromosome evolution because centromeric repeats can be reshaped by polyploidization, subgenome differentiation and chromosomal rearrangement. Here, we combined low-coverage repeatome profiling, genome-wide mapping, fluorescence in situ hybridization (FISH), fiber-FISH and higher-order repeats (HORs) analysis to investigate centromere-associated repeats in Miscanthus sinensis (2n = 2x = 38). We identified MsSat1, a 137 bp satellite repeat representing approximately 2.1% of the genome, as a major centromere-associated satellite repeat. MsSat1 signals overlapped with the Saccharum officinarum centromeric retrotransposon probe and formed continuous tandem arrays on DNA fibers, supporting its association with centromeric domains. MsSat1-based clustering further recovered 13 related satellite variants, many of which showed strongly biased distribution between the A and B subgenomes. Among them, MsSat1-G displayed the most striking pattern, with strong B-subgenome enrichment and chromosome 7-specific localization. Detailed analysis revealed that MsSat1-G was embedded within an expanded MsSat1-rich domain on the fusion-derived chromosome 7, where it showed a more heterogeneous HORs architecture than the major MsSat1 array. These findings suggest that chromosome fusion was accompanied by local centromeric satellite amplification and repeat-architecture remodeling. Together, our study reveals that fusion-derived centromeres can preserve distinct satellite signatures and provides cytogenetic and genomic evidence that centromeric repeats record both subgenome differentiation and chromosome restructuring in M. sinensis.