Fig. 4.

Systematic identification of miR-203 targets by microarray and bioinformatic analyses with the inducible mouse model. (A) Six hours after Dox injection, miR-203 is readily induced in the basal progenitor cells only in the DP but not in the SP control skin. All epidermal cells are marked by H2BGFP; the basement membrane is marked by β4 integrin; miR-203 is marked by red cytoplasmic signals. Scale bars: 10 μm. (B) miR-203 is induced at the physiological level in the basal progenitor cells. The expression of miR-205 is not affected by the induction of miR-203 by Dox treatment. (C) Twenty-four hours after Dox treatment, p63 is downregulated ∼30% at the mRNA level, as measured by qPCR. (D) High enrichment of miR-203 seed sequences is detected only in the 3′UTR of the downregulated genes upon miR-203 induction. Neither 5′UTR nor the CDS is enriched for the miR-203 seed sequences. (E) Perfect matches to 5′ sequences of miR-203, including numbers 1-7, 2-8 and 3-9 nucleotides but not other regions of miR-203, e.g. numbers 16-22, are highly enriched in the 3′UTRs of the downregulated genes. (F) Effectiveness of seed matches on the downregulation of target mRNAs. Changes in the mRNA level after miR-203 induction are plotted with the cumulative distributions. Repression from UTRs containing an 8-mer (2-9) site is most significant, whereas 7-mer sites (2-8, 3-9, 1-7) also show strong repression, compared with all expressed UTRs. (G) Effectiveness of the number of seed matches on the downregulation of target mRNAs. Changes in the mRNA level after miR-203 induction are plotted with the cumulative distributions. Repression from UTRs containing more than two seed matches is strongest, whereas repression from UTRs containing 2 or 1 seed matches is also significant, compared with UTRs without any seed match.