Study On The Stability Of RNA M~6A Methylase METTL3 Protein Regulated By HSP90-CHIP

Many kinds of chemical modifications of RNA exist in cells,which have been identified more than 150 kinds so far.And the 6-methyladenosine(m~6A)that was one of the most common RNA modifications on m RNA was widely found in all kinds of eukaryotes.The required methyltransferases(Writers),demethylases(Erasers)and binding proteins(Readers)for m~6A have been successively identified.METTL3(methyltransferase like 3)is the catalytic core enzyme in methyltransferase complex.Recent studies have shown that m~6A plays an important role in tumor development.In most of cancers,the METTL3protein is highly expressed and its activity is elevated so that mediating the protein regulation of METTL3 is crucial to the development of cancer.Therefore,mediating the stability regulation of METTL3 protein is crucial for the occurrence and development of cancer.To date,little is known about studies on the post-translational modification of METTL3protein.Therefore,we wanted to know how this modification of ubiquitination regulates the stability of the METTL3 protein.In this study,tumor cells were used as materials to focus on the regulation of METTL3 protein stability by the heat shock protein HSP90 and ubiquitin ligase CHIP complex.The main findings are as follows:1.Various cells were treated under various stress conditions,and it was found that the protein level of METTL3 increased significantly under heat shock(41-42℃).METTL3protein levels were also found to increase significantly when HEK293T cells were treated with proteasome inhibitor MG132.2.The results of Co-Immunoprecipitation(Co-IP)experiments showed that METTL3had physical interactions with HSP90 and HSP70.3.Treatment of various cells with two different HSP90 inhibitors PU-H71 and 17-AAG.Western Blot(WB)test results showed that both inhibitors could lead to a significant decrease in endogenous METTL3 protein levels.4.Through Overexpression,Knockdown and other experiments,it was found that METTL3 physically interacts with CHIP and CHIP could significantly degrade the protein level of METTL3.In summary,the experimental results indicated that HSP90 and CHIP regulate the stability of METTL3 protein together.However,the regulation of modification level for m~6A by HSP90 inhibitors needed further study.In this paper,the stability regulation of METTL3 protein by HSP90 inhibitor may affect the modification level of m~6A.The study provides a basis for more systematic and in-depth functional research in the future and provides a new target for anti-tumor drug research and development.