Time-resolved multiomics profiling reveals chromatin O-GlcNAc modification promotes senescence-associated transcriptional program

O-GlcNAc modification acts as a fundamental cellular signal across a broad range of physiological and pathological contexts; however, its mechanistic impact on cellular senescence, particularly in coordinating the senescence-associated transcriptional landscape, has not been systematically elucidated. Here, we applied an integrative, time-resolved chemical genomics strategy to map the dynamic landscape of O-GlcNAc chromatin-associated proteins (OCPs) during the process of oncogene-induced senescence (OIS) in primary human fibroblasts. OIS progression is marked by the sustained accumulation of chromatin O-GlcNAc modifications, which drives cellular commitment to OIS. Dynamic multiomics profiling revealed that 1,987 senescence-associated OCPs undergo extensive shifts in genomic occupancy across diverse epigenetic chromatin states and exhibit bimodal, sequential regulatory activities within the 3,466-gene senescence transcriptome. O-GlcNAc serves as an important epigenetic determinant, facilitating the formation of dual-function complexes (TF-SWI/SNF and NuRD) that enable the activation of senescence-associated secretory phenotype (SASP) genes at promoters while enforcing the repression of cell cycle regulators at enhancers. Additionally, we identified O-GlcNAc-modified JUN and GATAD2A as key regulators of OIS phenotypes in both in vitro and in vivo models of senescence-driven tumorigenesis. Our findings reveal the dynamic regulation and chromatin organization principles of O-GlcNAc-related epigenetic factors, offering valuable insights into cellular senescence and potential therapeutic strategies.