Fig 2: CREB interacts with KMT5A to regulate PTP1B transcriptional activity in endothelial cells.A CREB and H4K20me1 were enriched at the PTP1B promoter region. B The putative CREB binding site of PTP1B. C PTP1B promoter activity was determined by luciferase reporter assays. D western blot analysis of KMT5A and CREB levels in HUVECs. E, F qPCR analysis of mRNA expression of KMT5A and CREB. G western blot analysis of KMT5A and CREB levels in HUVECs. H, I qPCR analysis of mRNA expression of KMT5A and CREB. J western blot analysis of KMT5A and PTP1B levels in HUVECs. K, L qPCR analysis of mRNA expression of KMT5A and PTP1B. (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, n = 5/group).

Fig 3: Immunoblots showing the hippocampal ERK–CREB–BDNF signaling-pathway-related protein expression levels of the OVX mice treated with AE (100 or 300 mg/kg/d, p.o.). Results are presented as mean ± SD. ### p < 0.001 vs. SHAM group; * p < 0.05, ** p < 0.01, *** p < 0.001 vs. OVX group.

Fig 4: Bortezomib and ONX-0914 differentially affect neural protein expression after 24 h. Cell lysates of primary cortical cultures (DIV14), treated for 24 h with indicated inhibitor concentrations, were analysed via immunoblot for selected proteins (A) and signal intensities were subsequently determined (B). Normalization was ensured via loading same protein amounts and quantitative analysis of the respective Coomassie stained sister gel. Data were expressed in relation to the control condition (100%). Note, protein levels for NF?B, CREB and pCREB showed a tendency towards decreased expression with increasing inhibitor concentrations, whereas this effect was evident earlier and stronger on Bortezomib treatment compared to ONX-0914. Conversely, levels of the apoptosis marker Caspase 3 were significantly increased after Bortezomib treatment at doses of 0.01 and 0.1 µM after 24 h. No significant differences were detected for the protein level of alpha6, ß-actin, AMPA and Synaptophysin 1 as well as the glial marker Iba1 and GFAP after 24 h inhibitor treatment (4B). Activation of CREB signalling was assessed by calculation of the pCREB/CREB ratio (4C). A slight tendency for CREB activation indicated by phosphorylation was observed after 24 h at doses of 0.1 µM for both inhibitors. Presented data are mean ±SEM of 2–3 technical replicates each from 3 independent experiments, B: Student’s t-test, *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001, C: one-sample t-test against hypothetical value 1

Fig 5: mPGES-1/PGE2 regulated MTDH expression via the cAMP/PKA-CREB pathway. The adenylate cyclase (AC) agonist forskolin increased the expression of p-CREB, CREB and MTDH, while the protein kinase A (PKA) inhibitor H89 induced the opposite effects on these proteins (a, b). The relative luciferase activity (firefly/Renilla) in group 1 (cells cotransfected with pCMV-GFP-Puro-01-CREB1, pGL3-MTDH and pRL-TK) was almost 3.43 times that in group 2 (cells cotransfected with pCMV-GFP-Puro-NC, pGL3-MTDH and pRL-TK) (c). **p < 0.01, *p < 0.05.