Stroke by inducing HDAC9-dependent deacetylation of HIF-1 and Sp1, promotes TfR1 transcription and GPX4 reduction, thus determining ferroptotic neuronal death

Abstract

Background: The inhibition of histone deacetylase 9 (HDAC9) represents a promising druggable target for stroke intervention. Indeed, HDAC9 is overexpressed in neurons after brain ischemia where exerts a neurodetrimental role. However, mechanisms of HDAC9-dependent neuronal cell death are not yet well established. Methods: Brain ischemia was obtained in vitro by primary cortical neurons exposed to glucose deprivation plus reoxygenation (OGD/Rx) and in vivo by transient middle cerebral artery occlusion. Western blot and quantitative real-time polymerase chain reaction were used to evaluate transcript and protein levels. Chromatin immunoprecipitation was used to evaluate the binding of transcription factors to the promoter of target genes. Cell viability was measured by MTT and LDH assays. Ferroptosis was evaluated by iron overload and 4-hydroxynonenal (4-HNE) release. Results: Our results showed that HDAC9 binds to hypoxia-inducible factor 1 (HIF-1) and specificity protein 1 (Sp1), two transcription activators of transferrin 1 receptor (TfR1) and glutathione peroxidase 4 (GPX4) genes, respectively, in neuronal cells exposed to OGD/Rx. Consequently, HDAC9 induced: (1) an increase in protein level of HIF-1 by deacetylation and deubiquitination, thus promoting the transcription of the pro-ferroptotic TfR1 gene; and (2) a reduction in Sp1 protein levels by deacetylation and ubiquitination, thus resulting in a down-regulation of the anti-ferroptotic GPX4 gene. Supporting these results, the silencing of HDAC9 partially prevented either HIF-1 increase and Sp1 reduction after OGD/Rx. Interestingly, silencing of the neurodetrimental factors, HDAC9, HIF-1, or TfR1 or the overexpression of the prosurvival factors Sp1 or GPX4 significantly reduced a well-known marker of ferroptosis 4-HNE after OGD/Rx. More important, in vivo , intracerebroventricular injection of siHDAC9 reduced 4-HNE levels after stroke by preventing: (1) HIF-1 and TfR1 increase and thus the augmented intracellular iron overload; and (2) a reduction of Sp1 and its target gene GPX4. Conclusions: Collectively, results obtained suggest that HDAC9 mediates post-traslational modifications of HIF-1 and Sp1 that, in turn, increases TfR1 and decreases GPX4 expression, thus promoting neuronal ferroptosis in in vitro and in vivo models of stroke.

Selenium Nanoparticles in Protecting the Brain from Stroke: Possible Signaling and Metabolic Mechanisms

“…Epigenetic effects of selenium sources in protecting brain cells from stroke may involve histone deacetylase 9 (HDAC9), which leads to an increase in HIF-1 and a reduction in Sp1 protein levels by deacetylation and deubiquitination. This effect of HDAC9 results in the downregulation of the anti-ferroptotic GPX4 gene, encoding the important selenium-containing protein glutathione peroxidase 4 [ 78 , 79 ]. Inhibition of HDAC9, including through selenium sources, may serve as one of the strategies for protecting brain cells.…”

Targeting epigenetic and posttranslational modifications regulating ferroptosis for the treatment of diseases

“… 301 AIS Acetylation TfR1/GPX4 HDAC9 promotes neuronal ferroptosis through increasing HIF-1 by deacetylation and deubiquitination, thus promoting the transcription of TfR1 and reducing Sp1 protein levels by deacetylation and ubiquitination, thus resulting in a down-regulation of GPX4 in in vitro and in vivo models of stroke. 302 ICH ncRNA ACSL4 LncRNA H19 protects against intracerebral hemorrhage injuries via regulating microRNA-106b-5p/ACSL4. 329 ICH ncRNA COX2/PGE2 miR-137 inhibit ferroptosis in oxyHb-treated SH-SY5Y cells via COX2/PGE2 pathway.…”

“… 19 , 266 – 292 Recently, accumulating evidence has revealed the roles of epigenetic regulation of ferroptosis via ncRNA expression, methylation, ubiquitination, and acetylation in AIS. 293 – 302 Upregulation of the circRNA Carm1 inhibits ferroptosis through its binding to microRNA-3098-3p to downregulate ACSL4 in OGD/R-treated HT22 cells. 293 Upregulation of LncRNA PVT1 promotes ferroptosis through miR-214-mediated suppression of TFR1 and P53 expression.…”

“… 301 HDAC9 promotes neuronal ferroptosis through deacetylation and deubiquitination, dependently increasing HIF-1 expression and thus increasing the transcription of TfR1 and decreasing the Sp1 protein level by deacetylation and ubiquitination, leading to downregulation of GPX4 in in vitro and in vivo models of stroke. 302 …”

Acetylation and Phosphorylation in the Regulation of Hypoxia-Inducible Factor Activities: Additional Options to Modulate Adaptations to Changes in Oxygen Levels

“…Few studies have investigated the role of HDAC9 in hypoxia [104]. Similar to the other class IIa members of the HDACs family, HDAC9 has been shown to interact with HIF1A, deacetylating it and maintaining its activities [105]. Additional mechanisms have also been proposed.…”