Figure 6

Fluoxetine was found to disassociate the MeCP2-CREB-Bdnf promoter IV complex via the phosphorylation of MeCP2. (a) Illustration shows the effect of MeCP2-CREB-Bdnf promoter IV complex on Bdnf gene expression. As shown in the upper panel, MeCP2 combined with methylated Bdnf promoters to form a repressor complex, which inhibited the CREB binding to the CRE and suppressed Bdnf gene transcription; As shown in the lower panel, the phosphorylated MeCP2 disassociated the CREB from this repressor complex, after which the CREB combined with the CRE and induced Bdnf gene transcription. (b) Immunoblot experiments show the protein levels of MeCP2, pMeCP2, CREB and pCREB among the PSD, PSDV and PSDF groups. (c–f) Quantification of the protein levels and the band intensities of MeCP2 (c), pMeCP2 (d), CREB (e) and pCREB (f) (n = 6 per group, one-way ANOVA followed by Newman-Keuls multiple comparisons). (g) Co-IP of 500 μg of proteins from the hippocampus with nonspecific IgG (IgG), anti-MeCP2 or anti-CREB. Input: 20 μg of protein from the extracts without IP was loaded. (h) Quantification of the Co-IP of the MeCP2/CREB ratio among the PSD, PSDV and PSDF groups (n = 6 per group, one-way ANOVA followed by Newman-Keuls multiple comparisons). (i) ChIP-qPCR assay of Bdnf promoter IV combined with pCREB among PSD, PSDV and PSDF groups (n = 6 per group, one-way ANOVA followed by Newman-Keuls multiple comparisons). Full-length blots/gels are presented in Supplementary Figure 5. *P < 0.05, **P < 0.01 vs. PSDV group. Data are presented as the mean ± SEM. PSD: post stroke depression group; PSDV: PSD with vehicle injection group; PSDF: PSD with fluoxetine injection group.