Study On The Molecular Mechanism Of RCC1 And PALB2 Involved In DNA Damage Repair

Despite constant assaults by endogenous and environmental agents on the DNA,the genes of the human body have not become a bunch of garbled information or degraded.In response to this threat,life have developed several systems to detect DNA damage,send out signals and mediate its repair so that the body can restore the integrity and fidelity of DNA and maintain the genetic stability of the genome.The destruction or uncontrollability of DNA repair pathways could lead to genome instability,therefore,elucidating the mechanism of DNA damage repair is critical to maintaining genome stability and the integrity of genetic information.Cell cycle progression and DNA damage repair work together to help stabilize the genetic material inside the cell.PALB2(Partner And Localizer Of BRCA2)is an important breast cancer inhibitor.As a BRCA2 binding protein,PALB2 colocalizes with BRCA2 in nuclear foci,promotes its localization and stability in DNA damage repair.As a core protein,PALB2 connects BRCA1 and BRCA2 and promotes homologous recombination repair(HR).RCC1(Regulator Of Chromosome Condensation 1)binds to H2 A / H2 B on nucleosomes and participates in chromatin condensation in the late S phase and the early M phase.Many damage repair factors' function in DNA damage response(DDR)and the cell cycle progress depend on Ran-GTPase-regulated nucleocytoplasmic transport(NCT).RCC1 is the only guanine exchange factor of Ran.Combined with Ran,RCC1 promotes the conversion of Ran-GDP to Ran-GTP.As a chromatin concentration regulator,RCC1 is involved in chromatin concentration and the concentration of chromatin is closely related to the process of DNA damage repair.First,we found that PALB2 is a new interacting protein of RCC1 by immunoprecipitation.In order to identify the key domains of interaction between PALB2 and RCC1,we transfected PALB2 wild-type and deletion mutant plasmids into 293 T cells,we found that the interaction between PALB2 and RCC1 depends on P11 domain of PALB2(aa.692-759)by immunoprecipitation,Deletion of P11 domain of PALB2 could abrogate the interaction between PALB2 and RCC1.And we constructed RCC1 partial deletion mutant plasmids D1-D3.Next,we investigated the effects of PALB2,RCC1,and Ran knockdown on acetylated histone expression.After knocking down the expression of PALB2,RCC1 or Ran in U2 OS cells by si RNA,we detected changes in the expression levels of acetylation of lysine at different sites on H2 A and H2 B.Compared with the control group,the expression levels of H2BK12 ac,H2BK15ac,H2BK20 ac were significantly reduced.What's more,after knocking down RCC1 and Ran,H4K16 ac expression level was also significantly reduced.In summary,we initially verified the interaction between RCC1 and PALB2 and the regulatory mechanisms of RCC1,PALB2 and Ran on the expression levels of histones H2BK12 ac,H2BK15ac,H2BK20 ac and H4K16 ac,laid the foundation for further study on how RCC1 and PALB2 synergistically participate in DNA damage repair and their respective functions.