The full-length transcripts and promoter analysis of intergenic microRNAs in Drosophila melanogaster
The full-length transcripts and promoter analysis of intergenic microRNAs in Drosophila melanogaster
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果蝇基因间 microRNA 的全长转录本和启动子分析。
DOI：
10.1016/j.ygeno.2011.02.004
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发表时间：
2011-05-01
期刊：
影响因子：
4.4
通讯作者：
Li, Fei
中科院分区：
文献类型：
--
作者：
Qian, Jinjun;Zhang, Zan;Li, Fei
microRNA（miRNA）转录至今仍不清楚。为了增加miRNA的丰度，我们在果蝇S2细胞中用CuSO 4刺激miRNA的转录，并用dsRNA敲低Drosha酶。获得了bantam、miR-276 a和miR-277的全长转录本、miR-8的5 '端、miR-2b的3'端和miR-10的全长转录本。我们还进行了一系列的miRNA启动子分析，以证明RACE结果的可靠性。荧光素酶报告基因检测证明bantam和miR-276 a启动子成功驱动下游荧光素酶基因的表达。通过在预测的转录因子结合位点引入一个或多个突变来损害启动子活性。染色质免疫沉淀分析证实低磷酸化的RNA聚合酶II和转录因子c-Myc在miRNA启动子处物理结合。对转录因子Mad和Prd的RNA干扰导致bantam、miR-277和miR-2b的表达下调，但对miR-276 a的表达无影响，而Dorsal的RNA干扰则具有相反的效果。(C)2011 Elsevier Inc. All rights reserved.
MicroRNA (miRNA) transcription is still not well understood until now. To increase the miRNA abundance, we stimulated miRNA transcription with CuSO4 and knocked down Drosha enzyme using dsRNA in Drosophila S2 cells. The full length transcripts of bantam, miR-276a and miR-277, the 5'-end of miR-8, the 3'-end of miR-2b and miR-10 were obtained. We also conducted a series of miRNA promoter analysis to prove the reliability of RACE results. Luciferase-reporter assays proved that both bantam and miR-276a promoters successfully drove the expressions of downstream luciferase genes. The promoter activities were impaired by introducing one or multiple mutations at predicted transcription factor binding sites. Chromatin immunoprecipitation analysis confirmed that hypophosphorylated RNA polymerase II and transcription factor c-Myc physically bind at miRNA promoter. RNA interference of transcription factors Mad and Prd led to down-expression of bantam, miR-277 and miR-2b but not miR-276a, whereas RNAi of Dorsal had the opposite effect. (C) 2011 Elsevier Inc. All rights reserved.