Figure 7. Depletion of either Nr1h3 or Rara mimics the Mecp2 knockdown (KD) phenotype during quiescence exit.
(A) ChIP-qPCR analyses of Mecp2 at the promoter-proximal regions of Rara and Nr1h3 in Mecp2fl/fl and Mecp2-cKO livers before and 6 hr post-partial hepatectomy (PHx). (B–D) Either Nr1h3 or Rara KD promotes serum restimulation (SR)-induced quiescence exit in 3T3 cells. (B) Real-time PCR showing lentivirus-mediated KD of either Nr1h3 or Rara. shLuc served as a negative control. (C) Western blotting (WB) of pRb, Cyclin D1, Nr1h3, and Rara in control and Nr1h3 or Rara KD 3T3 cells at the indicated time points. (D) The effect of Nr1h3 or Rara KD on quiescent exit in 3T3 cells determined by Ki67/PI staining followed by flow cytometry. Data are presented as means ± SEM; In (A, B), n = 5; in (D), n = 3. *p<0.05; ***p<0.001; ****p<0.0001 by two-way ANOVA. (E–H) Either Nr1h3 or Rara KD further enhances quiescence exit in Mecp2 cKO livers. (E) Real-time PCR showing adeno-associated virus (AAV)-mediated KD of either Nr1h3 or Rara. shLuc served as a negative control. (F) WB of pRb, Cyclin D1, Nr1h3, and Rara in control and Nr1h3 or Rara KD 3T3 cells at the indicated time points. (G) The effect of Nr1h3 or Rara KD on quiescent exit in Mecp2 cKO livers determined by IF and liver index (H) before and 6 hr post-PHx. Data are presented as means ± SEM. In (E, H), n = 5 mice/group; n.s., not significant; *p<0.05, ****p<0.0001 by two-way ANOVA. (I) Model of the negative regulatory role for Mecp2 in fine-tuning quiescence exit.