Figure 3. Neurotoxic insults increase HDAC2 via stress elements in its promoter.

a, c, Representative pictures of HDAC2 and PGR1 labeling of primary hippocampal neurons treated with (a) H₂O₂ and (c) Aβ-oligomers (n=20-40 neurons per group), scale bar, 10μm. b, d, Quantification of (a) and (c). e, f, Quantitative RT-PCR results showing Hdac2 expression in (e) H₂O₂- and Aβ-treated primary hippocampal neurons, and (f) in the CK-p25 hippocampus (n=7-9 mice each). g, Alignment of the vertebrate glucocorticoid responsive element (GRE) consensus sequence with the GRE in the proximal promoter of mouse Hdac2. h, Quantification and representative WB images of hippocampal extracts of CK-p25 versus control mice (n=3 each). i, Representative images of immunohistochemical labeling of PGR1 and HDAC2 in the CK-p25 hippocampus (n=3-6 slices from 3 mice each); scale bar, 20μm. j, Regression analysis of (i) showing a significant correlation between PGR1 and HDAC2 in CK-p25 (R²=0.686, p≤0.001), but not control mice (R²=0.019, n.s.). k, Quantification and representative WB images of Cdk5cKO and control Cdk5fl/fl forebrain extracts (n=3 each). l, Quantitative PCR results of PGR1-immunoprecipitated chromatin around the GRE in a 1.3 kb-wide Hdac2 promoter region (schematically shown above the graph; TSS, transcriptional start site) in the CK-p25 and control hippocampus (n=3-6 animals each); green lines represent fragments amplified by primer pairs. m, Luciferase activity of CAD cells transfected with the Hdac2 promoter with (orange) or without (blue) GRE (schematic of constructs shown above graph), and treated with H₂O₂ and Aβ_1-42. n, Luciferase activity of CAD cells transfected with Hdac2-GRE in the presence of endogenous GR or of cotransfected GRS211A. In vitro results are from ≥3 independent experiments. *p≤0.05; **p≤0.01; ***p≤0.001, values are mean ± s.e.m.