The Role And Mechanism Of PRMT1-mediated Stability Of Histone H4 In Cellular Senescence

Cellular senescence is an irreversible cell cycle arrest process.Multiple factors can trigger senescence including telomere shortening,oncogenic signaling,oxidative stress,and DNA damage.Characteristic features of senescent cells include cell cycle arrest,active DNA damage response?DDR?,enhance senescence-associated secretory phenotype?SASP?,senescence associated?-galactosidase?SA-?-gal?activity,and chromatin structure changes.Chromatin structure change is partially mediated by altering its constitutive components such as histones.For example,late-passage cells senescence displays reduced level of core H3 and H4 histone components of the nucleosome.Furthermore,Nucleosome occupancy decrease has also been demonstrated to be associated with global upregulation of gene expression with aging in yeast and mammalian livers,while ameliorating this loss in yeast extends lifespan.Together,these data show that senescence is accompanied with histones level alteration;however,the role and the mechanism of histones reduction in cellular senescence are far from elucidated.PRMT1 is an important epigenetic modification enzyme in mammalian cells,mediates the asymmetric dimethylation of histone H4 on arginine 3?H4R3me2as?and plays a role in activating gene transcription.Studies over the past decades also linked PRMT1 to aging and cellular senescence.PRMT1 can also methylate DAF-16/FOXO,blocks its phosphorylation by AKT,increases the expression of longevity-related genes,thus leading to life span extension of C.elegans.At the cellular level,PRMT1loss-of-function cells exhibit cell cycle delays,checkpoint activation,genome instability and cellular proliferation inhibition.Our previous study also indicates that depletion of PRMT1 arrests breast cancer cell growth and induces cellular senescence.However,the roles of PRMT1-mediated H4R3me2as in cellular senescence remain largely unknown.We found that knockdown PRMT1 significantly reduced the level of histone H4while inducing cellular senescence.Subsequently,we found that 4 core histones were down-regulated during Ras,Etoposide and H₂O₂ induced cellular senescence,and the downregulation of histone H4 was earlier than the other 3 core histones,indicating that downregulation of histone H4 may be an early marker of aging.Further studies showed that the downregulation of histone H4 was caused by the decrease of its stability.Co-IP confirmed that the ubiquitination level of histone H4 did not change,while the interaction between PA200 and histone H4 was significantly increased during cellular senescence,revealing that the PA200 proteasome mediated the degradation of histone H4.Futhermore,we found that the downregulation of PRMT1-mediated H4R3me2as was earlier than the degradation of histone H4.RNAi targeted PRMT1-3'UTR knockdown of endogenous PRMT1,and overexpression of wild-type PRMT1 inhibited degradation of histone H4,but not enzyme mutation of PRMT1.Co-IP and peptide pull down demonstrated that H4R3me2as antagonize association of PA200 with H4,indicating that PRMT1-mediated H4R3me2as maintains histone H4 stability.Mechanistically,the degradation of histone H4 reduced the nucleosome occupancy,which in turn changed the gene expression network,activated the transcription of the cell cycle inhibitor genes p21,GADD45A,SASP-related genes FGF2,MMP3,IL11,IGFBP7 and anti-apoptotic genes BCL-XL,BCL-W,and resulting in cellular senescence.In addition,we also found that the anti-aging drugs metformin,rapamycin and resveratrol can inhibit the degradation of histone H4,The restorationof histone H4 was significantly earlier than that of cyclinA2 and the reduction of p21 and SA-?-gal positive cells,indicating that histone H4 can be used as a target for anti-aging drugs screening.The present study unveils a novel function of PRMT1-mediated H4R3me2as in modulation of cellular senescence via regulating histone H4 stability.Our findings also points to the value of histone H4 as an early senescence indicator and a potential anti-aging drug screening marker andprovides an important theoretical basis for early identification of aging and anti-aging drugs screening.