Centromere-size reduction and chromatin state dynamics following intergenomic hybridization in cotton

Centromeres are pivotal for accurate chromosome segregation, yet their regulation and evolutionary dynamics remain poorly understood. Here, we investigate centromeres of the diploid species Gossypium anomalum (Ga, B-genome) that were transferred into tetraploid cotton G. hirsutum (Gh, AD-genome) as either an additional or integrated chromosome, as well as in synthetic allohexaploid (AABBDD) lines. We demonstrate consistent size reduction for all Ga centromeres in the Gh background. Histone modification profiling across 10 marks revealed heightened levels of both active and repressive chromatin marks within the Ga centromeres when transferred into the Gh background, particularly for H3K36me2. The centromeric histone modification perturbation extended into pericentromeric regions, with variable CENH3-binding domains consistently exhibiting a more pronounced increase in histone modification levels compared to stable centromere regions, highlighting the role of histone modification elevation in centromere dynamics. In addition, we observed enhanced chromatin accessibility and the presence of non-B-form DNA motifs, such as A-phased DNA repeats within stable centromere domains that are correlated with centromere stability. Hi-C analysis reveals a reorganized 3D chromatin architecture within the introgression line centromeres, including the formation of new topologically associating domains linked to H3K36me2 dynamics, emphasizing the importance of H3K36me2 in centromere organization. Together, these findings elucidate epigenetic mechanisms underlying centromere composition following intergenomic hybridization and allopolyploid formation, offering insights into centromere evolution in plants and its myriad epigenetic and potentially functional dimensions.