Fig. 3.

NQO1 binding to the 3′UTR of SERPINA1 mRNA promotes its translation. (A) Biotin-RNA pulldown indicated that NQO1 was capable of binding the 3′ untranslated region (UTR) and the coding region (CR) of SERPINA1 mRNA in two different human cell lines. (B) 3′UTR of SERPINA1 mRNA was cloned downstream of the renilla luciferase (RL) coding sequence, and firefly luciferase (FL) expressed from the same construct served as internal normalization control. After NQO1 silencing, HepG2 cells were transfected with the vector control (psiCHECK2) or with a construct containing SERPINA1 3′UTR. Twenty h later, the ratio of RL/FL activity in siNQO1 cells vs control siRNA cells was calculated using empty vector as reference. (C) NQO1 silencing inhibited RL/FL reporter activity in HepG2 cells. (D) RT-qPCR analysis of RL mRNA levels normalized to FL mRNA levels in the vector control (psi) and in the construct containing SERPINA1 3′UTR (psi-3′UTR). Data represent the mean of three independent experiments ± SEM. ****, P < 0.001.