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. 2019 Apr 2;9:225. doi: 10.3389/fonc.2019.00225

Figure 4.

Figure 4

BAG3 knockdown destabilizes IL6 mRNA via its 3′UTR. (A) Nascent RNA was labeled and isolated, newly synthesized IL6 mRNA was analyzed using real-time RT-PCR in control or BAG3 knockdown PDAC cells. (B,C) Actinomycin D was added for the indicated period to block RNA synthesis, and IL6 mRNA was analyzed using real-time RT-PCR in control or BAG3 knockdown BxPC3 (B) or SW1990 (C) cells. (D,E) Amanitin was added for the indicated period to block RNA synthesis, and IL6 mRNA was analyzed using real-time RT-PCR in control or BAG3 knockdown BxPC3 (D) or SW1990 (E) cells. (F,G) Relative IL6 mRNA expression at 4 h of Actinomycin D (F) or Amanitin (G) was normalized by that of 2 h. (H) Control or BAG3 knockdown BxPC3 cells were transfected with the indicated luciferase reporter vector and a Renilla reporter vector. Luciferase activity was measured 2 days after transfection and Renilla activity was measured to normalize luciferase activity. N.S., not significant; *P < 0.01.