Regulation Of DAPK Protein And MRNA Expression By Sumo Signaling Pathway And LncRNA

SUMO(Small Ubiquitin Like Molecules)is a kind of ubiquitin polypeptide with a molecular weight of about 11 kDa.In eukaryotic cells,the SUMO process is involved in many physiological and biochemical reactions,including the replication and repair of DNA,the regulation of transcriptional activity and so on.SUMO specific proteases(SENPs)also play an important role in the process of SUMO.It not only catalyzes the maturation of SUMO precursors,but also catalyzes the transformation of SUMO substrates to SUMO,thus maintaining the dynamic balance between SUMO and SUMO in vivoLong non-coding RNAs(LncRNAs)are a class of non-coding RNA molecules with a length of over 200nt.They do not encode proteins,but they regulate the expression of genes on multiple levels in the form of RNA.Death associated protein kinase(DAPK)is a serine threonine kinase,which plays an important role in cell proliferation,autophagy,adhesion and anoikis process.Its function of disease and lack of Alzheimer's disease and atherosclerosis and cancer are closely related.In addition to the methylation of genes,the regulation of the expression of DAPK protein is mainly the pathway of protein degradation.However,the mechanism of sumo's regulation of DAPK has still not been found.In our previous study,we found that:When the SUMO signal pathway is activated,the expression of DAPK protein is downregulated.When SENPs is overexpressed,the expression of DAPK is up-regulated.And using point mutation method,it is found that Flag-SENP6 mainly regulates the expression of DAPK protein through the 1-364 amino acid region of DAPK.On this basis,the main purpose of this study is to study the mechanism of the regulation of the expression of DAPK protein by SUMO signaling pathway.We found that HA-DAPK(1-364)-M mainly through the proteasome degradation pathway.Using the GPS-SUMO system,we discovered the only two potential SUMO conjugation sites on the HA-DAPK(1-364)-M,we mutated 141st and 167th two sites separately or simultaneously.It is found that the regulation of Flag-SENP6 to HA-DAPK(1-364)-M is not through direct combination.In comparison to wild-type HA-DAPK(1-364)-M,no mutants were able to enhance the stability of HA-DAPK(1-364)-M.SUMO signaling pathways can neither through direct combination and does not affect DAPK protein degradation to regulate the expression of exogenous DAPK,and therefore the SUMO regulation must be through the influence of DAPK protein translation and mRNA stability.The mRNA expression of HA-DAPK(1-364)-M can be regulated by cotransfection of Flag-SENP6,HA-DAPK(1-364)-M and His-sumol,His-sumo3,UBC9,HA-DAPK(1-364)-M.Further exploration also found that the SUMO signaling pathway is mainly through the regulation of mRNA expression in the 1-364 region of DAPK.Moreover,the expression of the endogenous DAPK mRNA can be down-regulated and up-regulated after the establishment of the stable cell with the overexpression and knockdown UBC9.So how does the SUMO signaling pathway regulate the mRNA of DAPK?Recently,long noncoding RNAs(LncRNAs)have been shown to regulate gene expression at various levels,Including protein translation and mRNA stability.Then does SUMO regulate the expression of DAPK by adjusting the LncRNAs?To verify this conjecture,we overexpressed the LncRNAs of DAPK screened at the cellular level.The experimental results showed that LncRNA with Lnc-CTSL1-9:1 number could down regulate the expression of DAPK mRNA and protein.This topic mainly explores the mechanism of the regulation of the expression of DAPK protein by SUMO signaling pathway.As a potential drug target,DAPK is being paid more and more attention.Therefore,a comprehensive and in-depth understanding of the protein expression of DAPK and the improvement of its molecular regulatory network will help to research and develop more new drugs related to DAPK protein,which is conducive to the research and treatment of related diseases.