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Several authors have suggested or inferred that modest changes in microRNA expression can potentiate or impinge on their capacity to mediate gene repression, and that doing so could play a significant role in diseases. Such interpretations are based on several assumptions, namely: (i) changes in microRNA expression correlate with changes in the availability of mature, functional miRISC, (ii) changes in microRNA expression can significantly alter the stoichiometry of miRISC populations with their cognate targets, (iii) and this, in turn, can result in changes in miRISC silencing output. Here, we experimentally challenge those assumptions by quantifying and altering the availability of miRISC across several families of microRNAs. Doing so revealed a surprising fragmentation in the miRISC functional pool, striking differences in the availability of miRNA families and saturability of miRNA-mediated silencing. Furthermore, we provide direct experimental evidence that only a limited subset of miRNAs, defined by a conjuncture of expression threshold, miRISC availability and low target site abundance, is susceptible to competitive effects through microRNA-binding sites.

On the availability of microRNA-induced silencing complexes, saturation of microRNA-binding sites and stoichiometry