Abstract
Previous data have suggested that the nuclear translocation of annexin 1 (ANXA1) is involved in neuronal apoptosis after ischemic stroke. As the mechanism and function of ANXA1 nuclear migration remain unclear, it is important to clarify how ANXA1 performs its role as an apoptosis ‘regulator' in the nucleus. Here we report that importazole (IPZ), an importin β (Impβ)-specific inhibitor, decreased ANXA1 nuclear accumulation and reduced the rate of neuronal death induced by nuclear ANXA1 migration after oxygen-glucose deprivation–reoxygenation (OGD/R). Notably, ANXA1 interacted with the Bid (BH3-interacting-domain death agonist) promoter directly; however; this interaction could be partially blocked by the p53 inhibitor pifithrin-α (PFT-α). Accordingly, ANXA1 was shown to interact with p53 in the nucleus and this interaction was enhanced following OGD/R. A luciferase reporter assay revealed that ANXA1 was involved in the regulation of p53-mediated transcriptional activation after OGD/R. Consistent with this finding, the nuclear translocation of ANXA1 after OGD/R upregulated the expression of Bid, which was impeded by IPZ, ANXA1 shRNA, or PFT-α. Finally, cell-survival testing demonstrated that silencing ANXA1 could improve the rate of cell survival and decrease the expression of both cleaved caspase-3 and cleaved poly(ADP-ribose) polymerase. These data suggested that Impβ-dependent nuclear ANXA1 migration participates in the OGD/R-dependent induction of neuronal apoptosis. ANXA1 interacts with p53 and promotes p53 transcriptional activity, which in turn regulates Bid expression. Silencing ANXA1 decreases the expression of Bid and suppresses caspase-3 pathway activation, thus improving cell survival after OGD/R. This study provides a novel mechanism whereby ANXA1 regulates apoptosis, suggesting the potential for a previously unidentified treatment strategy in minimizing apoptosis after OGD/R.
Highlights
The results revealed that Oxygen-glucose deprivation–reoxygenation (OGD/R)-induced cell death and that overexpression of ANXA1 associated with OGD/R promoted neuron death
We elucidated the role of nuclear ANXA1 migration in OGD/R-induced cell apoptosis
We found that following OGD/R, ANXA1 translocated from the cytoplasm to the nucleus and that this migration could be blocked by the importin β (Impβ) inhibitor IPZ
Summary
Ischemia–reperfusion is a well-recognized pathological condition that is characterized by an initial deprivation of blood supply to an area or organ followed by subsequent vascular restoration and concomitant reoxygenation of downstream tissues.[1,2] Ischemia–reperfusion occurs during various complications of vascular diseases, such as stroke and myocardial infarction.[1,3,4,5] Oxygen-glucose deprivation–reoxygenation (OGD/R) is an accepted model for studying ischemia–reperfusion in vitro.[6,7,8] Previous findings have indicated that annexin 1 (ANXA1) is involved in neuronal apoptosis after OGD/R, but the underlying mechanism remains unclear.[9]. Results from our previous study indicated that ANXA1 migration into the nucleus and its regulation of Bid (BH3-interacting-domain death agonist) gene expression were involved in apoptosis.[9] exactly how ANXA1 regulates Bid expression and induces apoptosis after OGD/R remains unclear. Bid can participate in neuronal apoptosis induced by brain ischemia,[14,15] and its expression is regulated by the transcription factor p53.16,17 The genomic loci for both the human and the mouse Bid genes contain p53-binding, DNA-response elements that bind p53 and mediate p53-dependent transactivation of reporter genes.[18,19] Unlike all other known BH3-only proteins, Bid needs to be cleaved by caspase-8 or other proteases into tBid to become activated, whereupon it Received 01.2.16; revised 18.7.16; accepted 29.7.16; Edited by A Verkhratsky. Numerous p53 co-factors have been implicated in cellular processes, and the activation or suppression of these co-factors determines the fate of a cell.[27,28] Whether ANXA1 represents one of the p53 co-factors involved in the regulation of Bid expression and how ANXA1 might interact with p53 to coregulate Bid expression have yet to be determined
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