Thomas Maren, Lange-Grünweller Kerstin, Hartmann Dorothee, Golde Lara, Schlereth Julia, Streng Dennis, Aigner Achim, Grünweller Arnold, Hartmann Roland K
Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, 35032 Marburg, Germany.
Int J Mol Sci. 2013 Jun 7;14(6):12273-96. doi: 10.3390/ijms140612273.
The human polycistronic miRNA cluster miR-17-92 is frequently overexpressed in hematopoietic malignancies and cancers. Its transcription is in part controlled by an E2F-regulated host gene promoter. An intronic A/T-rich region directly upstream of the miRNA coding region also contributes to cluster expression. Our deletion analysis of the A/T-rich region revealed a strong dependence on c-Myc binding to the functional E3 site. Yet, constructs lacking the 5'-proximal ~1.3 kb or 3'-distal ~0.1 kb of the 1.5 kb A/T-rich region still retained residual specific promoter activity, suggesting multiple transcription start sites (TSS) in this region. Furthermore, the protooncogenic kinase, Pim-1, its phosphorylation target HP1γ and c-Myc colocalize to the E3 region, as inferred from chromatin immunoprecipitation. Analysis of pri-miR-17-92 expression levels in K562 and HeLa cells revealed that silencing of E2F3, c-Myc or Pim-1 negatively affects cluster expression, with a synergistic effect caused by c-Myc/Pim-1 double knockdown in HeLa cells. Thus, we show, for the first time, that the protooncogene Pim-1 is part of the network that regulates transcription of the human miR-17-92 cluster.
人类多顺反子miRNA簇miR - 17 - 92在造血系统恶性肿瘤和癌症中经常过度表达。其转录部分受E2F调控的宿主基因启动子控制。miRNA编码区上游直接的内含子富含A/T区域也有助于簇的表达。我们对富含A/T区域的缺失分析表明，其强烈依赖c-Myc与功能性E3位点的结合。然而，缺少1.5 kb富含A/T区域5'近端约1.3 kb或3'远端约0.1 kb的构建体仍保留残余的特异性启动子活性，表明该区域存在多个转录起始位点（TSS）。此外，从染色质免疫沉淀推断，原癌基因激酶Pim-1、其磷酸化靶点HP1γ和c-Myc共定位于E3区域。对K562和HeLa细胞中pri-miR - 17 - 92表达水平的分析表明，E2F3、c-Myc或Pim-1的沉默对簇的表达有负面影响，在HeLa细胞中c-Myc/Pim-1双敲低会产生协同效应。因此，我们首次表明原癌基因Pim-1是调控人类miR - 17 - 92簇转录的网络的一部分。
