The Regulation Of5'UTR Of LINE1on Tumor-associated MiRNAs And Their Target Genes

In recent years, more and more non-coding sequence play important roles. The roles of non-coding sequence in the regulation of physiological and pathological, especial in tumorigenesis, were investigated frequently. Most of non-coding sequences are repeat sequences, which most of them are mobile elements, including transposon and retrotransposon. Regarding retrotransposons, long interspersed nuclear elements (LINEs) is the major elements in human genome and LINE1is the moverwhelming one among them, LINE1is composed of ORFK,ORF2and non-coding region5'UTR,3'UTR. Non-coding sequence5'UTR harbors a bidirectional promoter, forward promoter from Obp, and reverse promoter from400bp, the sequence between400bp and680bp play important role in activity of promoter. So it is implicated that its transcripts will form a double string RNA, which can process a siRNA in vivo. LINE1appears frequently in the form of5'truncated, and its promoter is hypermethylation in most situations and so as to make it silence, whereas, in the developing germ cells, early embryonic tissues and tumor cells, LINE1is activated. It suggested that LINE1activation may involve a common mechanism of tumorigenesis.Non-coding RNA, especially miRNA, attract more attentions. Through inducing mRNA degradation and complementary combination of target mRNA, cause translational repression or other forms of regulation mechanism of inhibiting the expression of target genes. To date, more than9500miRNAs have been found and named, which involved in cell growth, differentiation and cell cycle regulation and so on,50%of them are located in chromosomal regions associated with cancer or fragile regions, it is considered to be the ideal targets for tumor detection and therapy as an oncogene or tumor suppressor genes, miRNA will open a new avenue for understanding tumorigensis. The purpose of this paper is to combine the two, research the regulation of5'UTR of LINE1on tumor-associated miRNAs and their target genesBased on the characters of5'UTR of LINE1, we hypothesized that the transcripts derived from both promoters of5'UTR may contribute to siRNA and miRNA, which give rise to many genes response subsequently. To elucidate the function of5'UTR of LINE1, the investigation was categorized three parts as follows:1. The impact of LINE15'UTR on tumor-associated miRNA expressionWith RNAs prepared by transcription in vitro, HEK293cells were transfected by the double-strand RNA, the sense strand RNA and the antisense strand RNA of LINE15'UTR respectively. The alteration of known tumor-associated miRNA were detected by real time quantitiative PCR.2. The impact of LINE15'UTR on tagert genes of tumor-associated miRNABased on the condidate miRNA identified above, the target genes regulated by these miRNAs were investigated furthermore in transfected HEK293cells for their expression by real time quantitiative PCR.3. The mechanism investigation of the streamline of LINE15'UTR-miRNA-target genes by detecting the promoters methylation of traget genes.With transfected HEK293cells, the methylation in CpG island of promoters of target genes was detected by the approach of restrictive enzyme with methylation sensitivity combining real time quantitiative PCR.The study found that5'UTR of LINE1is significant to the expression of the miRNA and its target gene, detection of methylation fount that the affection of5'UTR of LINE1to expression of target gene of miRNA may be transcriptional lever not epigenetic lever.