Target repression induced by endogenous microRNAs: large differences, small effects

MicroRNAs are small RNAs that regulate protein levels. It is commonly assumed that the expression level of a microRNA is directly correlated with its repressive activity – that is, highly expressed microRNAs will repress their target mRNAs more. Here we investigate the quantitative relationship between endogenous microRNA expression and repression for 32 mature microRNAs in Drosophila melanogaster S2 cells. In general, we find that more abundant microRNAs repress their targets to a greater degree. However, the relationship between expression and repression is nonlinear, such that a 10-fold greater microRNA concentration produces only a 10% increase in target repression. The expression/repression relationship is the same for both dominant guide microRNAs and minor mature products (so-called passenger strands/microRNA* sequences). However, we find examples of microRNAs whose cellular concentrations differ by several orders of magnitude, yet induce similar repression of target mRNAs. Likewise, microRNAs with similar expression can have very different repressive abilities. We show that the association of microRNAs with Argonaute proteins does not explain this variation in repression. The observed relationship is consistent with the limiting step in target repression being the association of the microRNA/RISC complex with the target site. These findings argue that modest changes in cellular microRNA concentration will have minor effects on repression of targets. Two replicates of S2-DRSC cells under normal conditions微小核糖核酸（microRNAs）是一类能够调控蛋白质水平的小型RNA分子。学界普遍认为，微小核糖核酸的表达水平与其抑制活性呈直接正相关：即表达量越高的微小核糖核酸，对其靶信使核糖核酸（mRNA）的抑制作用越强。本研究针对黑腹果蝇（Drosophila melanogaster）S2细胞中的32种成熟微小核糖核酸，探究了内源性微小核糖核酸表达水平与抑制活性之间的定量关联。总体而言，我们发现表达丰度更高的微小核糖核酸对靶标的抑制作用更强。然而，二者间的关联并非线性：微小核糖核酸浓度每提升10倍，靶标抑制效果仅提升10%。无论是占主导的向导链微小核糖核酸，还是少量成熟产物（即所谓的过客链/microRNA*序列），其表达与抑制的关联模式均一致。但我们也观测到部分特例：部分微小核糖核酸的细胞浓度相差数个数量级，却对靶标mRNA产生了相近的抑制效果。同样地，表达水平相近的微小核糖核酸，其抑制能力也可能存在显著差异。本研究证实，微小核糖核酸与阿戈纳特蛋白（Argonaute proteins）的结合无法解释上述抑制活性的差异。我们观测到的关联模式与靶标抑制过程中的限速步骤相符：即微小核糖核酸与RNA诱导沉默复合物（RNA-induced silencing complex，RISC）形成的复合物与靶标位点的结合为限速环节。上述研究结果表明，细胞内微小核糖核酸浓度的小幅变化，对靶标抑制效果的影响较为有限。正常培养条件下的S2-DRSC细胞两份生物学重复样本。