Functional Research Of SAMHD1 Mutants Associated With Colon Cancer

SAMHD1 is a protein encoded by eukaryotic gene,which is a natural antiviral factor.It has d NTPase activity,which can reduce the level of intracellular d NTPs and inhibit the replication of the virus.SAMHD1 is abundant in organism,and is expressed in many tissues of human body.SAMHD1 can not only inhibit the replication of the virus,but also can be used to down regulation d NTPs in the tumor cells so that inhibit the proliferation of tumor cells.However,in cancer cells,SAMHD1 is usually mutated,this mutation will affect its activity,thereby affecting its anti-tumor effect,and its expression will be reduced to varying degrees.In our study,we choose five single-site mutants in colon cancer.By comparing the mutants and wild type(109aa-626aa),we found that mutants of SAMHD1 can still be expressed in prokaryotes,just soluble in varying degrees.Although the protein expressed in the form of inclusion body still accounts for the majority,however,it is still possible to obtain a sufficient amount of soluble protein for purification and subsequent structural studies by extended culture.Under certain condition,d GTP was added into the purified SAMHD1 and its mutants for a period of time,and the activity of SAMHD1 and its mutants were determined by HPLC.However,the SANHD1 mutation in colon cancer is associated with a loss of d NTPase activity,which is closely related to the loss of tumor cell proliferation.Related reports have shown that SAMHD1 inhibits the transposition of the retrotransposon LINE-1,which is also associated with tumor progression.By transfection of LINE-1 and SAMHD1 and their mutant plasmid in cells,we found that the inhibitory ability of SAMHD1 mutants to LINE-1 was reduced,which may be a factor in tumorigenesis.Previous research on SAMHD1 also showed that the antivirus ability of SAMHD1 can be inhibited by the accessory protein Vpx of HIV-2/SIV.Vpx causes ubiquitination of SAMHD1 by recruitment of the E3 ubiquitin complex and thus is degraded by proteasome.In the study of SAMHD1 and its associated colon cancer mutants,we found that SAMHD1 and its mutants,in addition to D497 Y,are still degraded by Vpx.This may be due to the fact that D497 Y is near to the Vpx interaction region,and the conformational changes caused by its mutation lead to the degradation of Vpx.This paper focuses on the research of colon cancer in five kinds of SAMHD1 mutants,mainly study the mutant SAMHD1 and wild-type SAMHD1 differences in consumption of intracellular d NTPs,differences in enzyme activity is SAMHD1,and the degradation of Vpx differences and retrotransposon LINE-1 influence.Finally,we obtained some data and provided some reference for the further study of the function of SAMHD1 and its related disease mutants.