Figure 3: Expression patterns of pri-miR-21-1, the primary transcript of miR-21.

Expression of pri-miR-21-1 in the heart (a–e) and the head vasculature (f–j) at 53 hpf. The WT (a,b,f and g), gata1 morphants (c,d,h and i) and tnnt2 morphants (e,j) were stimulated by epinephrine (Epi) from 51.5 to 53 hpf (b,d,e,g,i and j). Without Epi stimulation, pri-miR-21-1 was expressed only in the valve-forming endocardium (a), not in the head vasculature (f). Elimination of blood cells from the circulation by knocking down gata1 did not affect the expression (c,h). After stimulation by Epi, expression of pri-miR-21-1 became slightly stronger at the AV canal (b). (g) Expression in the head became evident in the metencephalic artery/middle cerebral vein (yellow arrowheads), the primary head sinus (green arrowheads) and the primordial mid-brain channel (black arrowheads). This ectopic induction was not affected in the gata1 morphants (i), but the absence of heartbeat cancelled it (j). (k) Relative expression of the mature miR-21 was measured by qRT–PCR using RNA isolated from the whole bodies. In the WT, Epi induced ~1.7-fold, whereas the induction in the tnnt2 morphant was minimal. In the gata1 morphants, miR-21 was induced at a relatively lower extent than the WT (~1.4-fold). Scale bars, 50 μm (a–e), 100 μm (f–j).