Histone variant H2A.J involved in persistent DNA damage signaling triggers senescence-associated inflammatory cytokine secretion

Irreparable DNA damage in response to ionizing radiation (IR) triggers prolonged DNA damage response (DDR) and induces premature senescence. Profound chromatin reorganization with formation of senescence-associated heterochromatin foci (SAHF) is an essential epigenetic mechanism for controlling different aspects of the senescence program, including senescence-associated secretory phenotype (SASP). Deciphering the molecular mechanisms provoking continuous DDR leading to radiation-induced senescence, we analyzed acute and long-term dynamics of histone variant H2A.J in human fibroblasts after IR exposure with different doses. During acute DDR most radiation-induced 53BP1-foci resolved within 24h post-IR, whereas only few 53BP1-foci persisted for days and weeks, particularly after high-dose exposure (≥10Gy). Nucleoplasmic H2A.J expression correlated with transient growth retardation after moderate doses and permanent cell-cycle arrest after higher doses. Significantly, H2A.J depletion had no obvious effects on DSB repair kinetics with the induction of transient or permanent cell-cycle arrest, nor on the establishment of senescent state. Using high-resolution imaging techniques, we found that persisting 53BP1-foci after IR develop into DNA segments with chromatin alterations reinforcing senescence (DNA-SCARS), characterized by patchy chromatin structures consistently located in the periphery of SAHFs. Electron microscopic immunogold analysis revealed that H2A.J is preferentially incorporated into these radiation-induced DNA-SCARS during senescence progression, forming huge co-localizing clusters with 53BP1. Strikingly, H2A.J depletion in siRNA-transfected fibroblasts abolished SASP expression, and thereby inhibited the chronic production of inflammatory mediators, but did not affect senescence-associated chromatin re-structuring nor stability of cell-cycle arrest. These findings provide new mechanistic insights into biological phenomena of SASP and suggest that H2A.J inhibition could be a reasonable strategy to ablate SASP, without affecting senescence-associated growth arrest.