Fig. 8

PABPC1 suppresses NUMB expression by enhancing miRNA-mediated NUMB degradation. A Direct interaction between PABPC1 and AGO2 in HUVECs was measured by Co-IP assay. B The site of NUMB 3’UTR bound by miR-31-5p and miR-146a-5p (upper). Dual-luciferase reporter system was performed to detect the interaction of miR-31-5p and miR-146a-5p with the NUMB 3’UTR (bottom). C, D Western blot (C), and RT-qPCR (D) confirming the degradation abilities of miR-31-5p and miR-146a-5p on NUMB. E–H Dual-luciferase reporter system showing the inhibition degree of NUMB by miR-31-5p and miR-146a-5p in HUVECs transfected with different amounts of miR-31-5p and miR-146a-5p with PABPC1 overexpression (E) or knockdown (F)/TRPM2-AS overexpression (G) or knockdown (H). I-K RT-qPCR (I), western blot (J), and immunofluorescence (K) measuring the activation of the NOTCH1 signaling pathway in the control group and in the PABPC1 overexpression group with/without miR-31-5p/miR-146a-5p treatment. Green fluorescence: corresponding primary antibody stained N1ICD; blue fluorescence: DAPI-stained nuclei. Scale bar: 20 μm. L-N RT-qPCR (L), western blot (M), and immunofluorescence (N) measuring the activation of the NOTCH1 signaling pathway in the control group and in the PABPC1 overexpression group with/without AGO2 knockdown. Data were assessed with unpaired Student’s t test or one-way ANOVA and presented as mean ± SD. * P < 0.05; ** P < 0.01; *** P < 0.001