Journal: Cellular and Molecular Life Sciences

Article Title: Upregulation of HDAC9 in hippocampal neurons mediates depression-like behaviours by inhibiting ANXA2 degradation

doi: 10.1007/s00018-023-04945-y

Figure Lengend Snippet: HDAC9 deacetylates ANXA2 to inhibit its ubiquitination-mediated degradation. A Real-time quantitative PCR analysis of ANXA2 in mice exposed to chronic restraint stress (CRS) or not (CON), and in HDAC9 conditional knockout (CKO) mice (n = 6 for each group). B Co-immunoprecipitation analysis showed that ANXA2 specifically binding to HDAC9 in hippocampus of CRS mice. C Immunoprecipitation analysis showed that decreased levels of ANXA2 were pulled down with an antibody against Acetyl-Lys in hippocampus of CRS mice. D Immunoprecipitation analysis showed that decreased levels of ANXA2 were pulled down with an antibody against ubiquitin in hippocampus of CRS mice

Article Snippet: Membranes were incubated with primary antibodies against the following: HDAC1 (1:5000, 66085-1-Ig, Proteintech, Wuhan, China), HDAC2 (1:5000, 12922-3-AP, Proteintech, Wuhan, China), HDAC4 (1:1000, 66838-1-Ig, Proteintech, Wuhan, China), HDAC5 (1:500, A01230-6, BOSTER, Wuhan, China), HDAC9 (1:1000, AP1109a, Abcepta, Guangzhou, China), β-tubulin (1:3000, 10094-1-AP, Proteintech, Wuhan, China), GAPDH (1:5000, 10494-1-AP, Proteintech), and ANXA2 (1:5000, 60051-1-lg, Proteintech).

Techniques: Ubiquitin Proteomics, Real-time Polymerase Chain Reaction, Knock-Out, Immunoprecipitation, Binding Assay

Journal: Cellular and Molecular Life Sciences

Article Title: Upregulation of HDAC9 in hippocampal neurons mediates depression-like behaviours by inhibiting ANXA2 degradation

doi: 10.1007/s00018-023-04945-y

Figure Lengend Snippet: Downregulation of ANXA2 rescues depression-like behaviours and spine loss induced by chronic restraint stress (CRS) or HDAC9. A Western blot analysis of ANXA2 expression in the hippocampus of CRS mice and control (CON) mice (n = 5 mice per group; p = 0.0013). B , C Timeline of adeno-associated virus (AAV) injection, behavioural tests, and western blotting. D Western blotting of ANXA2 in the hippocampus of mice that received stereotaxic administration of AAV-sgCON or AAV-sgANXA2 (n = 5 mice per group; p = 0.0018). E Immobility time in the FST. p = 0.0027 for sgCON vs. sgCON + CRS, p = 0.0106 for sgCON + CRS vs. sgANXA2 + CRS. F Immobility time in the TST; p = 0.0058 for sgCON vs. sgCON + CRS, p = 0.0494 for sgCON + CRS vs. SgANXA2 + CRS). G Sucrose consumption in the sucrose preference test (SPT; p = 0.0009 for sgCON vs. sgCON + CRS, p = 0.0018 for sgCON + CRS vs. SgANXA2 + CRS). H Total distance travelled in the OFT. For all the behavioural tests ( E – H ), n = 8 in each of the four groups. I Representative images of dendritic segments of hippocampal CA1 neurons in mice injected with AAV-eGFP (sgCON), mice exposed to CRS and injected with AAV-sgANXA2 (sgA2 + CRS), and mice injected with AAV-sgANXA2 and AAV-HDAC9 (sgA2 + HDAC9). J Stubby spine density in dendrites of CA1 neurons. K Thin spine density in dendrites of CA1 neurons. L Mushroom spine density in dendrites of CA1 neurons. M Total spine density in dendrites of CA1 neurons. In panels J – M , n = 15 spines from each group. N Whole-cell voltage-clamp recordings of sEPSCs in CA1 neurons of mice injected with AAV-eGFP (sgCON), mice exposed to CRS and injected with AAV-sgANXA2 (sgA2 + CRS), and mice injected with AAV-sgANXA2 and AAV-HDAC9 (sgA2 + HDAC9). O The amplitude of sEPSCs in hippocampal neurons from the three groups (n = 11–15 neurons; three mice per group). P The frequency of sEPSCs in hippocampal neurons from the three groups (n = 11–15 neurons; three mice per group). * p < 0.05, ** p < 0.01, *** p < 0.001. Two-tailed t test for A and D ; one-way ANOVA with Tukey’s post hoc test for others. Adjustments were made for multiple comparisons tests. Error bars represent the standard error of the mean

Article Snippet: Membranes were incubated with primary antibodies against the following: HDAC1 (1:5000, 66085-1-Ig, Proteintech, Wuhan, China), HDAC2 (1:5000, 12922-3-AP, Proteintech, Wuhan, China), HDAC4 (1:1000, 66838-1-Ig, Proteintech, Wuhan, China), HDAC5 (1:500, A01230-6, BOSTER, Wuhan, China), HDAC9 (1:1000, AP1109a, Abcepta, Guangzhou, China), β-tubulin (1:3000, 10094-1-AP, Proteintech, Wuhan, China), GAPDH (1:5000, 10494-1-AP, Proteintech), and ANXA2 (1:5000, 60051-1-lg, Proteintech).

Techniques: Western Blot, Expressing, Control, Virus, Injection, Two Tailed Test