SUMOylation Of Lin28A Promotes Tumorigenesis By Enhancing Its Inhibition Of Let-7 Biogenesis

MicroRNAs(miRNAs)are a class of small non-codingRNAs of?22-nucleotide(nt)that mediate target mRNA degradation and block translation by binding to complementary sequences in 3'untranslated region(3'UTR).The let-7 family is the most famous tumor suppressor miRNAs,for example,they are capable of inhibiting the expression levels of oncogenes such as c-Myc,Ras,HMGA2.In addition,they also play an important role in cell differentiation and normal development of the organism.Lin28A is an evolutionarily highly conservedRNA-binding protein mainly involved in post-transcriptional regulation of the let-7 family.Lin28A recruits TUT4/TUT7 to polyuridylate pre-let-7(let-7 precursor)by binding to a specific sequence on the terminal loop of pre-let-7,preventing processing cleavage of pre-let-7by DICER which simultaneously leads to degradation of pre-let-7,thereby ultimately inhibiting the maturation of let-7.A large number of studies have shown that the expression of Lin28A decreases with cell differentiation;however,Lin28A is reactivated in various malignant tumors such as lung cancer,breast cancer,prostate cancer and ovarian cancer,causing down-regulation of let-7 expression,which is highly positively correlated with the malignancy degree of the tumors and the poor prognosis of the tumor patients.Post-translational modifications(PTMs)are important mechanisms regulating protein activity and function.It has been found that Lin28A can undergo methylation,phosphorylation and acetylation,which can change the intracellular localization,stability or activity of Lin28A.As an important reversible PTM,SUMO modification(SUMOylation)has been extensively studied in recent years.In this study,we used the methods of Ni²⁺-NTA pull down,co-immunoprecipitation and prokaryotic expression system to identify that Lin28A could be modified in vitro and in vivo,and its main modification site is at the position of K15.By employing RIP,RNA pulldown,EMSA and surface plasmon resonance assays,we found that SUMO1 modification of Lin28A enhanced the binding ability of Lin28A to pre-let-7.Furthermore,using in vitro uridylation assays and in vitro processing experiments,we demonstrated that SUMO1modification of Lin28A promoted uridylation of pre-let-7 by the uridine transferase TUT4,impeding DICER-processing cleavage of pre-let-7,thereby inhibiting let-7biosynthesis.We also found that hypoxia promoted SUMO1 modification of Lin28A,while tumor chemotherapy drugs including cisplatin and paclitaxel reduced SUMO1modification of Lin28A,suggesting that SUMO1 modification of Lin28A may play an important role in tumorigenesis and tumor therapy.Finally,functional studies at the cellular and animal levels were carried out by constructing stable cell lines,indicating that SUMO1 modification of Lin28A does promote the proliferation,migration and invasion of tumor cells by inhibiting the production of let-7.In summary,the results revealed the function and molecular mechanism of SUMO1 modification of Lin28A,which can enhance its binding ability to pre-let-7 and inhibit the production of let-7,thereby promoting tumor progression.Our study provided a new strategy for tumor diagnosis,prognosis and targeted therapy.