Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins University School of Medicine, 1550 Orleans Street, Baltimore, MD 21287, USA.
Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins University School of Medicine, 1550 Orleans Street, Baltimore, MD 21287, USA ; Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, 1550 Orleans Street, Baltimore, MD 21287, USA.
FEBS Open Bio. 2014 May 2;4:441-9. doi: 10.1016/j.fob.2014.04.010. eCollection 2014.
Micro RNAs (miRNA) are non-coding RNAs expressed in the cytoplasm as their mature, 21-22-nucleotide short forms. More recently, mature miRNAs have also been detected in the nucleus, raising the possibility that their spatial distribution may be more complex than anticipated. Here we undertook comprehensive systematic analyses of miRNA distribution in several subcellular compartments of human cancer cells. In particular, we focused on the potential presence of miRNAs in the nucleolus, which contains an abundance of small non-coding RNAs. We employed two miRNA expression array platforms and small RNA deep sequencing of small RNAs isolated from cells, nuclei, cytoplasm and the nucleoli. We developed an assay to compare RNAs of isolated nucleoli before and after denaturation and used Northern hybridization to verify the presence of miRNAs in the subcellular compartments. Consistently, we found more than 10 miRNAs associated with the nucleolar preparations. Several miRNAs had greater relative abundance in the nucleolus compared to the other compartments. The nucleolar presence of miRNAs was independent of Dicer and the main activity of the nucleolus, RNA polymerase I transcription, but was dependent on CRM1 previously associated with nucleolar trafficking of small nucleolar RNAs. These results highlight the complexity of miRNA spatial arrangement and regulation.
微小 RNA（miRNA）是在细胞质中表达的非编码 RNA，以其成熟的 21-22 个核苷酸的短形式存在。最近，成熟的 miRNA 也在核内被检测到，这增加了它们的空间分布可能比预期的更复杂的可能性。在这里，我们对几种人类癌细胞亚细胞区室中 miRNA 分布进行了全面系统的分析。特别是，我们专注于 miRNA 存在于核仁中的可能性，核仁中含有大量的小非编码 RNA。我们采用了两种 miRNA 表达谱平台和从小细胞、核、细胞质和核仁中分离的小 RNA 进行的深度测序。我们开发了一种比较分离核仁中 RNA 在变性前后的方法，并使用 Northern 杂交验证了 miRNA 在亚细胞区室中的存在。一致地，我们发现了 10 多种与核仁制剂相关的 miRNA。与其他隔间相比，几种 miRNA 在核仁中的相对丰度更高。miRNA 在核仁中的存在与 Dicer 无关，与核仁的主要活性 RNA 聚合酶 I 转录无关，但与以前与小核仁 RNA 的核仁运输有关的 CRM1 有关。这些结果突出了 miRNA 空间排列和调节的复杂性。
