Experimental procedures to identify and validate specific mRNA targets of miRNAs.
Experimental procedures to identify and validate specific mRNA targets of miRNAs.
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DOI：
10.17179/excli2015-319
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发表时间：
2015
期刊：
影响因子：
4.6
通讯作者：
Yalowich JC
中科院分区：
文献类型：
--
作者：
Elton TS;Yalowich JC
功能成熟的microRNA（miRNAs）是一类小的单链非编码RNA分子，是基因表达的重要转录后调节因子，在许多生理和病理过程中发挥着重要作用。由于单个miRNA的生物学作用将由它们所调控的mRNA决定，因此miRNA/mRNA靶相互作用的鉴定和验证对于我们理解控制生物过程的调控网络至关重要。我们公布了预测算法的组合使用，实验支持的miRNA/mRNA相互作用的策展数据库的检查，在特定的目标mRNA中编目的miRNA结合位点的手动序列检查，并审查已发表的文献作为一个可靠的做法，识别和优先生物学上重要的miRNA/mRNA靶对。一旦选择了一个优选的miRNA/mRNA靶对，我们建议通过满足四个明确定义的实验标准来验证功能性miRNA/mRNA靶对的真实性。本文综述了我们目前对miRNA生物学、miRNA/mRNA靶点预测算法、验证的miRNA/mRNA靶点数据库的了解，并概述了几种验证miRNA/mRNA靶点的实验方法。此外，作为利用这些方法的一个例子，人类内皮糖蛋白的案例研究。
Functionally matured microRNAs (miRNAs) are small single-stranded non-coding RNA molecules which are emerging as important post-transcriptional regulators of gene expression and consequently are central players in many physiological and pathological processes. Since the biological roles of individual miRNAs will be dictated by the mRNAs that they regulate, the identification and validation of miRNA/mRNA target interactions is critical for our understanding of the regulatory networks governing biological processes. We promulgate the combined use of prediction algorithms, the examination of curated databases of experimentally supported miRNA/mRNA interactions, manual sequence inspection of cataloged miRNA binding sites in specific target mRNAs, and review of the published literature as a reliable practice for identifying and prioritizing biologically important miRNA/mRNA target pairs. Once a preferred miRNA/mRNA target pair has been selected, we propose that the authenticity of a functional miRNA/mRNA target pair be validated by fulfilling four well-defined experimental criteria. This review summarizes our current knowledge of miRNA biology, miRNA/mRNA target prediction algorithms, validated miRNA/mRNA target data bases, and outlines several experimental methods by which miRNA/mRNA targets can be authenticated. In addition, a case study of human endoglin is presented as an example of the utilization of these methodologies.