Figure 3

TTP binds to the 5’ proximal fragment of BDNF 3’UTR. a. Illustration showing the position of analyzed 3’UTR fragments U1 and U2 in BDNF-S, AREs are marked as on Figure 1a. b. Effects of 1–100 ng of plasmid/96-well encoding for TTP on reporter construct expression carrying BDNF-S, U1 and U2 in HEK-293 cells. TTP encoding plasmid has a moderate effect on U2 containing reporter construct expression only at the highest concentration (100 ng), while U1 is inhibited already at a 10 fold lower TTP encoding plasmid concentration (10 ng), suggesting different affinity of U1 and U2 towards TTP. A well-established target of inhibition by TTP, a 140 bp ARE-rich fragment from TNF-alpha 3’UTR (Luc-TNF-ARE) was included as a positive control, N = 2-3 experiments with 4 repeats per experiment. Error bars indicate SD, ***p < 0.001. c. Electrophoretic mobility shift assay showing band shift of RNA probe U1 with recombinant TTP protein (50 ng and 100 ng) and no band shift of RNA probe U2 under the same conditions. Experiment was repeated twice.