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首页> 外文期刊>Folia histochemica et cytobiologica >L1-ORF1p and Ago2 are involved in a siRNA-mediated regulation for promoter activity of L1-5’UTR
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L1-ORF1p and Ago2 are involved in a siRNA-mediated regulation for promoter activity of L1-5’UTR

机译：L1-ORF1p和Ago2参与siRNA介导的L1-5’UTR启动子活性的调控

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Introduction. Long interspersed nuclear elements-1 (L1), as the only one self-active retrotransposon of the mobile element, was found to be generally activated in tumor cells. The 5‘UTR of L1 (L1-5’UTR) contains both sense and antisense bidirectional promoters, transcription products of which can generate double-strand RNA (dsRNA). In addition, L1-ORF1p, a dsRNA binding protein encoded by L1, is considered to engage in some RNA-protein (RNP) formation. Ago2, one of the RISC components, can bind to dsRNA to form RISC complex, but its role in L1 regulation still remains unclear. Due that the 5‘UTR of L1 (L1-5’UTR) contains both sense and antisense bidirectional promoters, so the activities in both string were identified. A dsRNA-mediated regulation of L1-5’UTR, with the feedback regulation of L1-ORF1p as well as other key molecules engaged (Ago1–4) in this process, was also investigated. Material and methods. Genomic DNA was extracted from HEK293 cells and subjected to L1-5’UTR preparation by PCR. Report gene system pIRESneo with SV40 promoter was employed. The promoter activities of different regions in L1-5’UTR were identified by constructing these regions into pIRESneo, which SV40 region was removed prior, to generate different recombinant plasmids. The promoter activities in recombinant plasmids were detected by the luciferase expression assay. Western blot and co-immunoprecipitation were employed to identify proteins expression and protein-protein interaction respectively. Results. Ago2 is a member of Agos family, which usually forms a RISC complex with si/miRNA and is involved in post- transcriptional regulation of many genes. Here L1-ORF1p and Ago2 conducts a regulation as a negative feedback for L1-5'UTR sense promoter. L1-ORF1p could form the immune complexes with Ago1, Ago2 and Ago4, respectively. Conclusions. L1-5’UTR harbors both sense and antisense promoter activity and a dsRNA-mediated regulation is responsible for L1-5’UTR regulation. Agos proteins and L1-ORF1p were engaged in this process.
机译：介绍。长散布的核元素-1（L1），作为活动元素的唯一一个自我反转录转座子，通常在肿瘤细胞中被激活。 L1的5′UTR（L1-5′UTR）包含有义和反义双向启动子，其转录产物可产生双链RNA（dsRNA）。此外，L1-ORF1p（一种由L1编码的dsRNA结合蛋白）被认为参与了某些RNA-蛋白（RNP）的形成。 Ago2是RISC的一种成分，可以与dsRNA结合形成RISC复合体，但其在L1调控中的作用仍不清楚。由于L1的5'UTR（L1-5'UTR）包含有义和反义双向启动子，因此可以识别出两个字符串中的活性。还研究了dsRNA介导的L1-5’UTR调控，以及L1-ORF1p以及该过程中参与的其他关键分子（Ago1-4）的反馈调控。材料与方法。从HEK293细胞中提取基因组DNA，并通过PCR进行L1-5'UTR制备。使用具有SV40启动子的报告基因系统pIRESneo。通过将这些区域构建到pIRESneo中来鉴定L1-5’UTR中不同区域的启动子活性，该区域先去除了SV40区域，然后生成了不同的重组质粒。通过荧光素酶表达测定法检测重组质粒中的启动子活性。采用蛋白质印迹和免疫共沉淀分别鉴定蛋白质表达和蛋白质-蛋白质相互作用。结果。 Ago2是Agos家族的成员，该家族通常与si / miRNA形成RISC复合体，并参与许多基因的转录后调控。在这里，L1-ORF1p和Ago2进行调节，作为L1-5'UTR有义启动子的负反馈。 L1-ORF1p可以分别与Ago1，Ago2和Ago4形成免疫复合物。结论。 L1-5’UTR同时具有有义和反义启动子活性，而dsRNA介导的调控则负责L1-5’UTR调控。 Agos蛋白和L1-ORF1p参与了这个过程。

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