Sequential onset and concurrent expression of miR-9 genomic loci in single cells contributes to the temporal increase of mature miR-9 in zebrafish neurogenesis

Here, we show that the expression of miR-9 increases spatially and temporarily over zebrafish development. A detailed time course of the expression of 7 pri-miR-9 genomic loci shows that they have distinct temporal and spatial profiles, which may be brought about by different numbers of E/N-boxes in their regulatory regions. Focusing on pairs that are expressed in the same area of the hindbrain region, namely pri-miR-9-1/pri-miR-9-4 and pri-miR-9-1/pri-miR-9-5, we find that they sequentially activated during neurogenesis, concurrent with a known change in Her6 dynamics from noisy to oscillatory. Analysis of expression at the single-cell level shows that although they are sequentially activated, early and late pri-miRs are mostly concurrently transcriptionally active in the same cells. This finding supports the idea that increased mature miR-9 is contributed, at least in part, by overlapping activation of distinct loci. We propose that this may lead to an additive, sharp, increase of mature miR-9 which in turn may lead to rapid changes in Her6 dynamics.