Figure 3.

METTL3 (methyltransferase-like protein 3) regulates fibrinolysis in vitro via Jun proto-oncogene (JUN)-PAI-1 (plasminogen activator inhibitor-1) axis. A, Schematic diagram showed the JUN-binding position in PAI-1 promoter (left) and JUN binding to PAI-1 promoter was determined by ChIP-PCR (right). ACTB (β-actin) was used as a negative control to show immunoprecipitation specificity. Input: sheared chromatin was prepared before immunoprecipitation and used as a positive control for PCR. B and C, The expression of JUN and PAI-1 in shJUN HUVECs were detected by quantitative real-time PCR (qPCR; B) and Western blot (C) (n=3, data are median±SD). D, Fibrin formation and fibrinolysis were shown in shJUN HUVECs and control cells (n=3, data are mean±SEM). E and F, The expression of JUN and PAI-1 in JUN overexpressed HUVECs and control cells were analyzed by Western blot (E) and qPCR (F) (empty vector: EV, n=3, data are mean±SEM). G, Fibrin formation and fibrinolysis were shown in JUN overexpressed and control cells (n=3, data are mean±SEM). H and I, The expression of METTL3 and PAI-1 in shMETTL3 HUVECs infected with WT JUN (shMETTL3+OE JUN) and control lentivirus (shMETTL3+EV) were measured by Western blot (H) and qPCR (I) (n=3, data are mean±SEM). J, Fibrinolysis was shown in shMETTL3 HUVECs infected WT JUN (shMETTL3+OE JUN) and control lentivirus (shMETTL3+EV) (n=3, data are mean±SEM).