Histone acetyltransferase P300 deficiency promotes ferroptosis of vascular smooth muscle cells by activating the HIF-1α/HMOX1 axis

Background: E1A-associated 300-kDa protein (P300), an endogenous Histone Acetyltransferase, contributes to modifications of the chromatin landscape of genes involved in multiple cardiovascular diseases. Ferroptosis of vascular smooth muscle cells (VSMCs) is a novel pathological mechanism of aortic dissection. However, whether P300 regulates VSMC Ferroptosis remains unknown.

Methods: Cystine deprivation (CD) and imidazole ketone erastin (IKE) were used to induce VSMC Ferroptosis. Two different knockdown plasmids targeting P300 and A-485 (a specific inhibitor of P300) were used to investigate the function of P300 in the Ferroptosis of human aortic smooth muscle cells (HASMCs). Cell counting kit-8, Lactate Dehydrogenase and flow cytometry with propidium iodide staining were performed to assess the cell viability and death under the treatment of CD and IKE. BODIPY-C11 assay, immunofluorescence staining of 4-hydroxynonenal and malondialdehyde assay were conducted to detect the level of lipid peroxidation. Furthermore, co-immunoprecipitation was utilized to explore the interaction between P300 and HIF-1α, HIF-1α and P53.

Results: Compared with normal control, the protein level of P300 was significantly decreased in HASMCs treated with CD and IKE, which was largely nullified by the Ferroptosis inhibitor ferrostatin-1 but not by the Autophagy Inhibitor or Apoptosis inhibitor. Knockdown of P300 by short-hairpin RNA or inhibition of P300 activity by A-485 promoted CD- and IKE-induced HASMC Ferroptosis, as evidenced by a reduction in cell viability and aggravation of lipid peroxidation of HASMCs. Furthermore, we found that hypoxia-inducible factor-1α (HIF-1α)/heme oxygenase 1 (HMOX1) pathway was responsible for the impacts of P300 on Ferroptosis of HASMCs. The results of co-immunoprecipitation demonstrated that P300 and P53 competitively bound HIF-1α to regulate the expression of HMOX1. Under normal conditions, P300 interacted with HIF-1α to inhibit HMOX1 expression, while reduced expression of P300 induced by Ferroptosis inducers would favor HIF-1α binding to P53 to trigger HMOX1 overexpression. Furthermore, the aggravated effects of P300 knockdown on HASMC Ferroptosis were largely nullified by HIF-1α knockdown or the HIF-1α Inhibitor BAY87-2243.

Conclusion: Thus, our results revealed that P300 deficiency or inactivation facilitated CD- and IKE-induced VSMC Ferroptosis by activating the HIF-1α/HMOX1 axis, which may contribute to the development of diseases related to VSMC Ferroptosis.

Ferroptosis; HIF-1α; HMOX1; P300; P53; Vascular smooth muscle cells.