microRNAs (miRs) are small regulatory non-coding RNAs that post-transcriptionally regulate gene expression. miR-9 is a miR that is predominantly expressed in the central nervous system, where it regulates cell fate transitions during neurogenesis in Xenopus, zebrafish and mouse embryos. In this study, Nancy Papalopulu and colleagues spatially and temporally map the expression of miR-9 precursors (pri-miR-9-1 to pri-miR-9-7) during zebrafish neurogenesis. Using quantitative single molecule fluorescent in situ hybridisation (smFISH), the authors find that pri-miR-9 expression occurs in a step-wise, additive manner, whereby pri-miR-9-3 to -5 are expressed in the hindbrain at early stages and pri-miR-9-1 is added at later stages of differentiation. Perturbing pri-miR-9-1 function by CRISPR/Cas-9 mutagenesis decreases late neuronal differentiation and fails to attenuate Hairy-related 6 (Her6) expression, oscillations of which regulate neuronal cell fate transitions. Mathematical modelling supports this notion, suggesting that an adaptive network containing Her6 is unresponsive to linear increases in miR-9, but responds to sharp incremental increases in miR-9 expression. Together, these data enhance our understanding of miR-9/Her6 dynamics. Specifically, they show how sequential pri-miR-9 precursor expression temporally regulates miR-9 levels and thus impacts Her6 dynamics during neurogenesis.
