Abstract
Ischemia-induced cell death is a major cause of disability or death after stroke. Identifying the key intrinsic protective mechanisms induced by ischemia is critical for the development of effective stroke treatment. Here, we reported that 14-3-3γ was a selective ischemia-inducible survival factor in cerebral cortical neurons reducing cell death by downregulating Bax depend direct 14-3-3γ/p-β-catenin Ser37 interactions in the nucleus. 14-3-3γ, but not other 14-3-3 isoforms, was upregulated in primary cerebral cortical neurons upon oxygen–glucose deprivation (OGD) as measured by quantitative PCR, western blot and fluorescent immunostaining. The selective induction of 14-3-3γ in cortical neurons by OGD was verified by the in vivo ischemic stroke model. Knocking down 14-3-3γ alone or inhibiting 14-3-3/client interactions was sufficient to induce cell death in normal cultured neurons and exacerbate OGD-induced neuronal death. Ectopic overexpression of 14-3-3γ significantly reduced OGD-induced cell death in cultured neurons. Co-immunoprecipitation and fluorescence resonance energy transfer demonstrated that endogenous 14-3-3γ bound directly to more p-β-catenin Ser37 but not p-Bad, p-Ask-1, p-p53 and Bax. During OGD, p-β-catenin Ser37 but not p-β-catenin Ser45 was increased prominently, which correlated with Bax elevation in cortical neurons. OGD promoted the entry of 14-3-3γ into the nuclei, in correlation with the increase of nuclear p-β-catenin Ser37 in neurons. Overexpression of 14-3-3γ significantly reduced Bax expression, whereas knockdown of 14-3-3γ increased Bax in cortical neurons. Abolishing β-catenin phosphorylation at Ser37 (S37A) significantly reduced Bax and cell death in neurons upon OGD. Finally, 14-3-3γ overexpression completely suppressed β-catenin-enhanced Bax and cell death in neurons upon OGD. Based on these data, we propose that the 14-3-3γ/p-β-catenin Ser37/Bax axis determines cell survival or death of neurons during ischemia, providing novel therapeutic targets for ischemic stroke as well as other related neurological diseases.
Highlights
There is not such clinically effective cytoprotective drug discovered until now
We demonstrated that 14-3-3g was an important early ischemia-inducible protective factor in cerebral cortical neurons
In pure cultured cerebral cortical neurons and cortical neurons in vivo, only the g but not any other isoform of 14-3-3 could be upregulated by oxygen–glucose deprivation (OGD) or permanent middle cerebral artery occlusion (pMCAo)
Summary
There is not such clinically effective cytoprotective drug discovered until now. It is well known that 14-3-3 elicits antiapoptotic effects by interacting with various pro-apoptotic proteins such as Bad, Bax, p53 and Ask-1 in the cytoplasm depending on specific apoptotic cues.[14,15,16] In mammalian brains, six (b, e, Z, g, t, z) of the seven 14-3-3 isoforms are highly expressed In neurological diseases, such as Creutzfeldt–Jakob disease, the elevation of distinct 14-3-3 isoforms in the cerebral–spinal fluid is considered as a biological marker of the disease or predictor of its progression.[17] Malfunction of 14-3-3 is heavily associated with neurodegenerative diseases such as Alzheimer’s disease[18] and Parkinson’s disease[19] the exact biological functions of 14-3-3 isoforms remain to be defined. We have previously reported that 14-3-3g is selectively upregulated by oxygen–glucose deprivation (OGD) in cultured astrocytes and protects them from OGD-induced cell death via binding to p-Bad Ser[112] selectively.[20,21] It is interesting to know whether 14-3-3g could protect ischemic neurons via a similar mechanism or not as 14-3-3g is expressed predominantly in neurons
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