The Role of MicroRNAs in the Regulation of Gene Expression

在哺乳动物细胞中，RNA干扰（RNAi）可以由合成的双链RNA介导， 称为小干扰RNA（siRNAs），通过介导完全互补的 mRNA转录物。microRNAs（miRNAs）是一种内源性小分子RNA， 用部分互补区域抑制mRNA，但也可能减少转录 程度.由于miRNAs被认为主要与3个未翻译的 区域（UTR）的转录本，但很少有实验数据支持这一点，我们寻求 为了研究模拟miRNAs的错配siRNAs是否会影响同源mRNA水平， 目标地点。我们的研究发现，靶向3个UTR的错配siRNA 两种内源性转录物的表达比靶向转录物的表达更能降低mRNA水平。 编码区。我们的研究结果证明了靶点位置的重要性， 与RNAi相关的小RNA的内源性mRNA，这些研究发表在FEBS上 2006年的信。为了解决改变的miRNA表达是否有助于癌症过程， Caplen博士（遗传学分支基因沉默科（GSS）科长）参与了 一项由CCR（实验室）的Curt Harriss博士小组进行的研究。人类致癌作用， (LHC)，CCR，NCI）与爱荷华州大学的Carlo Croce博士合作。Carlo Croce博士 其他人先前已经表明，不同类型的癌症与独特的 microRNA模式，这些模式可能是有价值的诊断和确定 预后在这项合作研究中（Yanaihara等人，2006 Cancer Cell 9：189-198）， 来自104对原发性肺癌的差异microRNA表达数据， 邻近正常组织证实了2009年报道的肺癌中microRNA的失调， 早期的论文，并发现了一种新的与预后不良相关的microRNA标记， hsa-miR-155。为了进一步研究miRNA表达在癌症中的作用， 我们自己的基于miRNA微阵列的分析在GSS中使用商业阵列平台， （Ambion/ABI），并已开始广泛分析miRNA表达乳腺癌细胞 线本研究发现了一些在不同乳腺癌组织中差异表达的miRNAs， 癌细胞系后续研究的重点是表征假定的目标， 这些差异表达的miRNAs。最后，我们研究了一个基因组区域，人类 染色体8 q24，通常与伯基特淋巴瘤和一些实体瘤相关 由于基因重排和/或缺失和扩增的结果， 存在以前未鉴定的miRNAs。许多研究表明，miRNAs可能 聚集在已知的基因组不稳定区域内，我们最近对此进行了审查， 特别强调小鼠基因组的特定主题领域（Huppi等人，研讨会 癌症生物学）。在与娜塔莉亚沃尔福夫斯基和鲍勃斯蒂芬斯博士（ABCC， NCI-Fredrick/SAIC Frederick Inc. NCI，NIH）12种可能的miRNA前体序列， 鉴定为定位于人chr. 8 q24区域内，其中7个通过实验 验证进一步的分子和遗传学研究正在调查这些新的miRNAs 在人类和小鼠系统中。

In mammalian cells, RNA interference (RNAi) can be mediated by synthetic duplex RNAs, termed small interfering RNAs (siRNAs, by mediating the cleavage of completely complementary mRNA transcripts. MicroRNAs (miRNAs) are endogenous small RNAs that assist in translationally repressing mRNAs with regions of partial complementarity, but may also reduce transcript levels. Since miRNAs are thought to predominantly interact with the 3 untranslated regions (UTRs) of transcripts but little experimental data exists to support this, we sought to ask if mismatched siRNAs mimicking miRNAs affect cognate mRNA levels as a function of target site location. Our studies found that mismatched siRNAs targeting the 3 UTRs of two endogenous transcripts yield a greater reduction in mRNA levels than those targeting the coding region. Our findings demonstrated the importance of target site location within endogenous mRNAs for small RNAs associated with RNAi and these studies were published in FEBS Letters in 2006. To address if altered miRNA expression can contribute to the cancer process Dr. Caplen, (Section head, Gene Silencing Section (GSS), Genetics Branch), became involved in a study conducted by Dr. Curt Harriss group at CCR (Lab. of Human Carcinogenesis, (LHC), CCR, NCI) in collaboration with Dr. Carlo Croce, University of Iowa. Dr. Carlo Croce and others have previously shown that different types of cancer are associated with unique microRNA patterns, and that these patterns may be valuable in diagnosing and ascertaining prognosis. In this collaborative study (Yanaihara et al., 2006 Cancer Cell 9: 189-198), differential microRNA expression data from 104 pairs of primary lung cancers matched against adjacent normal tissue confirmed the deregulation of microRNAs in lung cancer reported in earlier papers, and uncovered a new microRNA marker associated with poor prognosis, hsa-miR-155. To further study the role of miRNA expression in cancer we have now established our own miRNA microarray based analysis within GSS using a commercial array platform (Ambion/ABI) and have begun extensive analysis of the miRNA expression breast cancer cell lines. This work has identified a number of miRNAs differential expressed in different breast cancer cell line. Follow up studies are focused on characterizing the putative targets of these differentially expressed miRNAs. Finally, we have examined a genomic region, human chromosome 8q24, commonly associated with Burkitts lymphoma and some solid tumors as a result of genetic rearrangements and/or deletions and amplifications for evidence of the presence of previously unidentified miRNAs. A number of studies have suggested that miRNAs may be clustered within known areas of genomic instability and we have recently reviewed this particular subject area with a special emphasis on the mouse genome (Huppi et al., Seminars in Cancer Biology). In collaboration with Drs. Natalia Volfovsky and Bob Stephens (ABCC, NCI-Fredrick/SAIC Frederick Inc. NCI, NIH) 12 possible miRNA precursor sequences were identified as mapping to within the human chr. 8q24 region, of which, 7 were experimentally verified. Further molecular and genetic studies investigating these novel miRNAs are on going in both human and mouse systems.