The Mechanism Of GAPDH Involved In DNA Double-strand Break Repair

DNA is the carrier of life genetic information.The integrity and stability of DNA molecules are very important for maintaining normal life activities.However,the large number of DNA molecules in human cells and the vulnerability of DNA molecular structure make it easy to be damaged by a variety of endogenous and exogenous factors.If DNA damage is not repaired,it will lead to very serious diseases,such as cancer,immune deficiency and neurodegenerative diseases.Therefore,it is of great significance to study the specific mechanism and regulation process of DNA repair pathway for understanding the pathogenesis of diseases and formulating corresponding diagnosis,treatment and prevention measures.In this study,we found that the absence of glyceraldehyde-3-phosphate dehydrogenase(GAPDH)led to high sensitivity and accumulation of DNA double strand breaks(DSBs)damage induced by ionizing radiation(IR),and IR treatment promoted the nuclear transport of GAPDH.Using DSB repair report system,we found that the repair efficiency of HRR and NHEJ in GAPDH deficient U2 OS cells decreased significantly.By detecting DSB repair related proteins,the results showed that knockdown of GAPDH resulted in a significant decrease of RAD51 protein.Further studies showed that the decrease of RAD51 level was due to the decrease of protein stability.The RAD51 protein is regulated by the level of acetylation in the cell.We found that GAPDH can bind to Histone deacetylase 1(Histone deacetylase 1,HDAC1).The effect of HDAC1 on RAD51 protein was detected.The results showed that overexpression of HDAC1 increased the level of RAD51 protein,but did not affect its transcription level,overexpression of HDAC1 in GAPDH deficient cells could restore the level of RAD51 protein.In addition,the detection of HDAC1 and its target protein indicates that the loss of GAPDH reduces the enzyme activity of HDAC1.This part of the experiment shows that GAPDH can promote HDAC1 to target RAD51 by regulating the activity of HDAC1,thereby improving the stability of RAD51 protein.On the other hand,we studied the mechanism of the effect of GAPDH on HDAC1 activity.The activity of HDAC1 is regulated by other proteins in cells,among which maspin is the inhibitory protein of HDAC1.By analyzing the three-dimensional structure of the protein,we found that GAPDH and Maspin collide with the binding pocket of HDAC1,so there is a potential competitive binding mechanism — GAPDH can compete with Maspin to bind HDAC1,thus releasing HDAC1 activity.Furthermore,we found that the binding of HDAC1 to GAPDH was enhanced and the binding of HDAC1 to Maspin was decreased after IR treatment,thus the activity of HDAC1 was activated.Combined with the above experimental results,we conclude that under the treatment of IR,GAPDH enters the nucleus and competes with Maspin to bind HDAC1,which activates HDAC1 activity.HDAC1 enhances the targeting of RAD51 and then enhances the stability of RAD51 protein and promotes DSB repair.In conclusion,we found that GAPDH is involved in the repair of DSB.Because GAPDH is highly expressed in a variety of cancers,GAPDH is a potential anti-tumor target.Our results provide a new theoretical basis for targeting GAPDH as a tumor therapeutic target.