Figure 7. miR-153 regulates the expression of targets involved in the vesicle exocytosis pathway.

(A) Cells with luciferase reporter constructs containing wild-type (purple bars) or mutant (white bars) Bsn, Pclo, Snap25, Snca, Trak2, or Vamp2 mouse 3'-UTR region were co-transfected with the miR-153 expression plasmid. A miR-153 scrambled expression plasmid served as a control (leftmost bars). 3’-UTR mutations were in the seed sequence for the miR-153 binding site for each gene. HEK293T cells were co-transfected with both the reporter gene and miRNA expression vectors, and luciferase activity was measured 48 hr later. The experiments were performed in triplicate and error bars indicate standard deviation. (B) RT-qPCR analysis of RNA levels for genes (Bsn, Pclo, Snap25, Snca, Trak2, and Vamp2) from miRZip-153 (OE) infected 14 DIV hippocampal neurons (red, magenta and purple bars), relative to neurons infected with miRZip-scramble (OE), a control scrambled miR lentivirus (black bar). Each experiment was performed in triplicate. Error bars indicate standard deviation. *p<0.05, **p<0.01, ***p<0.001.

DOI: http://dx.doi.org/10.7554/eLife.22467.016

Figure 7—figure supplement 1. CBP/p300 is a target of miR-153.

(A) Schematic representation of CBP/p300 gene with miR-153 binding site indicated. (B) Cells containing the luciferase reporter constructs containing wild-type (black bars) or mutated CBP/p300 mouse 3'-UTR region (white bars) were co-transfected with miR-153. A miR-153 scrambled expression plasmid served as a control (left two bars). HEK293T cells were co-transfected with both the reporter gene and miRNA expression vectors, and luciferase activity was measured 48 hr later. The experiments were performed in triplicate and error bars indicate standard deviation. (C) Schematic representation of the feedback loop formed between miR-153 and CBP/p300. Green arrows indicate activation or positive feedback; red arrows indicate repression or negative feedback.

DOI: http://dx.doi.org/10.7554/eLife.22467.017