Fig. 3

Pri-miR-8 expression is independent of miPEP-8 control/activity. a schematic representation of constructs tested on miR-8 expression and activity levels. Arrows locate the primers used in the qPCR experiments determining miPEP and pri-miR relative expression levels. b the characterized pri-miR-8 produces a mature miR-8. S2 cells were transfected with a vector expressing pri-miR-8. Left: detection of the over-expression level of pri-miR-8 by qPCR. Right: detection of the over-expression level of mature miR-8 using the same RNA samples, n = 11 c miPEP-8 lacks repressive activity towards miR-8 expression. Left: level of miPEP-8 over-expression. Middle and right panels: quantification of pri-miR-8 and mature miR-8 in miPEP-8 over-expressing cells compared to control transfected cells (ctrl). n = 13 for the ctrl and 14 for miPEP-8. d, e Insensitivity of miR-8 sensor to miPEP-8 over-expression in S2 transfected cells (n = 16) (d) or in wing imaginal discs when miPEP-8 is expressed under the ptc-GAL4 promoter (e). In d, a miR-8 construct (n = 12) [17] was used as a positive control repressing the miR-8 luciferase sensor [20]. Of note, pri-miR-8 (n = 21) also repressed the miR-8 luciferase sensor. In e, first panel to the left: ptc GAL4 crossed with a UAS mCherry. Second panel to the left: expression pattern of the GFP miR-8 sensor alone. Scale bars (white) indicate 100 μm. A repressive activity is observed with miR-8 expressed in the ptc domain but not with miPEP-8. A representative disc is shown out of ten analyzed