Functional and evolutionary significance of human microRNA seed region mutations [M12]

MicroRNAs (miRNAs) are small non-coding RNAs that play a central role in the regulation of gene expression at the post transcriptional and/or translational level thus impacting various biological processes. Dysregulation of miRNAs could affect processes associated with progression of a variety of diseases including cancer. Majority of miRNA targeting in animals involves a 7-nt “seed region” mapping to positions 2-8 at the molecule’s 5' end. The importance of this 7 nt sequence to miRNA function is evidenced by the fact that the seed region sequence of many miRNAs is highly conserved within and between species. In this study, we computationally and experimentally explore the functional significance of sequence variation within the seed region of human miRNAs. Our results indicate that change of a single nt within the 7-nt seed region changes the spectrum of targeted mRNAs significantly meanwhile further nt changes have little to no additional effect. This high functional cost of even a single nucleotide change within the seed region of miRNAs explains why the seed sequence is highly conserved among many miRNA families both within and between species and could help clarify the likely mechanisms underlying the evolution of miRNA regulatory control. mRNAs were collected from 3 M12 miRNA treated and 3 negative control miRNA treated HEY ovarian cancer cell samples. mRNA expression was captured on Affymetrix U133 Plus 2 chips. To compare mRNA expression pattern between the M12 treated cells and the negative control treated cells, present/absent calls were generated using MAS5, while signals were calculated using GCRMA and then log2 transformed. Expression of differentially expressed genes or down regulated miRanda-mirSVR predicted miRNA target genes was compared between miRNA treated samples.

微小RNA（microRNAs, miRNAs）是一类小型非编码RNA，在转录后及/或翻译水平的基因表达调控中发挥核心作用，进而影响多种生物学过程。微小RNA的表达失调可影响包括癌症在内的多种疾病的发生进展相关生物学过程。在动物体内，大多数微小RNA的靶向作用涉及一段位于其5'端2-8号位点的7核苷酸“种子区域（seed region）”。诸多微小RNA的种子区域序列在物种内部及跨物种间均高度保守，这一事实印证了该7核苷酸序列对微小RNA功能的重要性。本研究通过计算与实验手段，探究人类微小RNA种子区域内序列变异的功能意义。本研究结果显示，7核苷酸种子区域内单个核苷酸的改变即可显著改变靶向mRNA的谱型，而后续额外的核苷酸改变则几乎不会产生额外影响。即便微小RNA种子区域内单个核苷酸的改变都会带来如此高的功能代价，这也解释了为何诸多微小RNA家族的种子序列在物种内部及跨物种间均保持高度保守，同时还有助于阐明微小RNA调控网络进化背后的潜在机制。研究人员从3例经M12微小RNA处理及3例经阴性对照微小RNA处理的HEY卵巢癌细胞样本中提取mRNA。mRNA表达谱通过Affymetrix U133 Plus 2芯片进行检测。为比较M12处理组与阴性对照组细胞的mRNA表达模式，研究人员采用MAS5算法生成存在/缺失检测结果，同时使用GCRMA算法计算信号强度并进行log2转换。研究人员对微小RNA处理样本间的差异表达基因，或经miRanda-mirSVR预测的下调微小RNA靶基因的表达水平进行了比较。