Effect Of Deficiency Of XPC On Autophagy Of Bladder Cancer Cells

Background and objectives:Xeroderma pigmentosum genome C is the earliest damage identification factor in nucleotide excision repair,playing an important role in DNA damage response and maintaining genetic stability of cells.Autophagy is a highly conserved process that maintains the steady state of the biological process of life in the cell evolution process.Autophagy plays a very key role in normal organism development dealing with the threat of the environment and degrading proteins and organelles.Autophagy also plays an important role in the process of DNA damage response,and the two are closely related.In our early study we found that XPC may be involved in many other processes such as cell cycle: chromotin remodeling and DNA damage response pathway.Whether XPC plays an important role in autophagy has not been reported.This study is aimed to explore the effect of XPC protein deficiency on autophagy in human bladder cancer cells.Methods:We used the already constructed stable XPC deficient cell model in our team;We used chloroquine to inhibit the degradation of autophagosomes so as to better observe the autophagosome accumulation;By using immunoblotting technique we observed the expression of autophagy characterized protein LC3 II before and after the interference of XPC;Fluorescence microscopy was used to visualize the GFP-LC3 puncta after transiently transfecting GFP-LC3 expression vector into T24 cells and HEK293 cells;CCK-8 was used to observe the cell proliferation;Western blot technique was used to detect the expression of proteins involved in autophagy and apoptosis signaling pathway.Results:1.After the interference of XPC,the accumulation of autophagosomes decreased,the proportion of cells with significantly aggregated GFP-LC3 punctas decreased;2.In the cisplatin induced autophagy,we found that after the interference of XPC,the accumulation of autophagosomes decreased,the proportion of cells with significantly aggregated GFP-LC3 punctas decreased;3.After treated with cisplatin,the trend of the abundance of LC3 II was altered by the interference of XPC in HEK293 cells : HEK293 NC cells responded slowly and rose and declined gradually,while HEK293 sh XPC cells responded much more quickly reaching to a peak and then declined gradually;4.Treating bladder cancer T24 cells constantly with cisplatin,the trend of the abundance of LC3 II was altered by the interference of XPC: T24 NC cells responded slowly and rose gradually,while T24 shXPC cells responded much more quickly reaching to a peak and then declined gradually;5.The HEK293 sh XPC cells wre more sensitive to cisplatin,proteins associated wirh DNA damage response pathway including p-ATM(S1981)and p53 were up-regulated;Bladder cancer T24 cells were more resistant to cisplatin after the interference of XPC.P-ATM(S1981)was up-regulated while p53 was down-regulated;Conclusions:With XPC deficiency,the autophagy and the cisplatin induced autophagy of HEK293 cells and bladder cancer T24 cells decrease,HEK293 cells become more sensitive to cisplatin while T24 cells become more resistant;To explore the molecular mechanisms of the sensitivity differences between the two kinds of cells caused by the autophagic degradation after the interference of XPC,we find that the induced state of p53 may play a key role in apoptosis.