MiRNA-328 Targets Histone H2AX And Regulates Lung Cancer Cells Apoptosis

Objective: Mi RNAs are small non coding RNAs(22 nucleotides)that bind partially complementary sites in the 3' untranslated regions(3?UTR)of target genes m RNA and regulate gene expression mostly at post-transcriptional level.It was known that miRNAs are widely involved in a variety of physiological and pathological processes,and abnormal expression of miRNAs is closely related to tumor development.Increasing reports indicated that miRNAs play a key role,in cell growth,differentiation,and apoptosis.H2 AX is one member of the histone H2 A family and holds an important physiological role of apoptotic regulation.It is related with production or development of tumor and sensitivity of tumors to drugs,H2 AX is knowns as a novel tumor suppressor.A large number of studies have shown that abnormal expression of miRNAs are closely related to tumor development.This topic is to investigate the function and role of the miRNA-328 in lung cancer,and its relationship with histone H2 AX,revealing the molecular mechanism of epigenetic regulation of lung cancer apoptosis.Methods: Firstly,this project mainly used gene silencing,gene chips and other technologies to reveal apoptosis-related genes and miRNAs expression profiles in lung cancer cells.Through analysis m RNA and miRNA expression data and function assays,we try to demonstrate the genes and miRNAs directly regulated by H2 AX in lung cancer cells,and make clear the molecular mechanism of epigenetic regulation of H2 AX in apoptosis.To explain the miRNA-328 relativity with lung cancer,we used four database GSE15008,GSE24709,GSE31568 and GSE61741 from the GEO(Gene Expression Omnibus)in the national center for biotechnology information(NCBI)to statistics gene expression of miRNA-328 in the lung cancer patients and normal control group.To explore the molecular mechanism of miRNA-328 involved in lung cancer cells apoptosis,three different concentrations(100 ?M,10 ?M,1 ?M)of etoposide(VP16)were used to treat lung cancer cells and the miRNA-328 expression was detected.In addition,we respectively transfected miRNA-328 mimics and miRNA-328 inhibitor into A549 and H1650 cells to determine whether miRNA-328 has a role in the apoptotic regulation lung cancer cell.Next,we used Target Scan,mi Randa and mi Rbase to identify the potential downstream targets of miRNA-328.In order to indicate that miRNA-328 functional targeting gene is histone H2 AX,we have used the luciferase report assay,q RT-PCR and western blot methods.Finally,to investigated whether miRNA-328 functions in cell apoptosis via targeting H2 AX and examine the role of H2 AX in lung cancer cell apoptosis,we have transfected a vector carrying H2 AX coding sequence which lacked the 3'UTR containing the mi R-328 binding sites,into A549 and H1650 cells.Results: In this study,we analyzed the GEO database at NCBI and found that miRNA-328 expression is generally associated with lung cancer.Then we examined the role of mi R-328 in apoptosis of lung cancer cells.Mi R-328 was reduced upon apoptotic induction with etoposide(VP16)for lung cancer cells.Further investigation revealed that the histone H2 AX is a direct and functional target of miRNA-328.Mi RNA-328 regulates apoptosis of lung cancer cells through targeting H2 AX.Our results showed that mi R-328 may act as an oncogene in the lung cancer cells.Conclusion: Our results illuminate that mi R-328 functions as an apoptosis silencer to regulate lung cancer cells apoptosis through targeting histone H2 AX and may become a critical therapeutic target in lung cancer.