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Published on: July 25, 2011
LINC00894 Regulates Cerebral Ischemia/Reperfusion Injury by Stabilizing EIF5 and Facilitating ATF4-Mediated Induction
Yifei Chen1,2, Hengxiang Cui3, Zhuanzhuan Han2
1Department of Emergency and Critical Care Medicine, The Second Affiliated Hospital of Soochow University, No.1055, San Xiang Road, Suzhou, Jiangsu, 215004, China.
Abstract:
The non-coding RNA LINC00894 modulates tumor proliferation and drug resistance. However, its role in brain is still unclear. Using RNA-pull down combined with mass spectrometry and RNA binding protein immunoprecipitation, EIF5 was identified to interact with LINC00894. Furthermore, LINC00894 knockdown decreased EIF5 protein expression, whereas LINC00894 overexpression increased EIF5 protein expression in SH-SY5Y and BE(2)-M17 (M17) neuroblastoma cells. Additionally, LINC00894 affected the ubiquitination modification of EIF5. Adeno-associated virus (AAV) mediated LINC00894 overexpression in the brain inhibited the expression of activated Caspase-3, while increased EIF5 protein level in rats and mice subjected to transient middle cerebral artery occlusion reperfusion (MCAO/R). Meanwhile, LINC00894 knockdown increased the number of apoptotic cells and expression of activated Caspase-3, and its overexpression decreased them in the oxygen-glucose deprivation and reoxygenation (OGD/R) in vitro models. Further, LINC00894 was revealed to regulated ATF4 protein expression in condition of OGD/R and normoxia. LINC00894 knockdown also decreased the expression of glutamate-cysteine ligase catalytic subunit (GCLC) and ATF4, downregulated glutathione (GSH), and the ratio of GSH to oxidized GSH (GSH: GSSG) in vitro. By using RNA-seq combined with qRT-PCR and immunoblot, we identified that fibroblast growth factor 21 (FGF21) and aconitate decarboxylase 1 (ACOD1), as the ATF4 target genes were regulated by LINC00894 in the MCAO/R model. Finally, we revealed that ATF4 transcriptionally regulated FGF21 and ACOD1 expression; ectopic overexpression of FGF21 or ACOD1 in LINC00894 knockdown cells decreased activated Caspase-3 expression in the OGD/R model. Our results demonstrated that LINC00894 regulated cerebral ischemia injury by stabilizing EIF5 and facilitating EIF5-ATF4-dependent induction of FGF21 and ACOD1.
Insights
The long non-coding RNA LINC00894 protects the brain from ischemic injury by stabilizing the EIF5 protein. This interaction promotes the expression of FGF21 and ACOD1, reducing cell death and improving outcomes after stroke.
Area of Science:
- Neuroscience
- Molecular Biology
- Genetics
Background:
- The role of long non-coding RNA LINC00894 in brain function is largely unknown.
- Previous studies indicate LINC00894 influences tumor progression and drug resistance.
Purpose of the Study:
- To investigate the function of LINC00894 in the brain, particularly in the context of cerebral ischemia.
- To elucidate the molecular mechanisms by which LINC00894 exerts its effects.
Main Methods:
- RNA-pull down assays, mass spectrometry, and RNA binding protein immunoprecipitation to identify interacting proteins.
- In vitro cell models (neuroblastoma cells, oxygen-glucose deprivation/reoxygenation) and in vivo animal models (transient middle cerebral artery occlusion/reperfusion).
- Gene knockdown and overexpression techniques, Western blotting, qRT-PCR, RNA-seq, and measurement of glutathione levels.
Main Results:
- LINC00894 directly interacts with and stabilizes the EIF5 protein, affecting its ubiquitination.
- LINC00894 overexpression reduced apoptosis and activated Caspase-3 in both in vitro and in vivo ischemia models, while knockdown had the opposite effect.
- LINC00894 regulates ATF4 expression, which in turn controls the expression of downstream target genes FGF21 and ACOD1, crucial for mitigating ischemic injury.
Conclusions:
- LINC00894 plays a protective role against cerebral ischemia injury.
- The mechanism involves EIF5 stabilization, leading to an EIF5-ATF4-dependent induction of FGF21 and ACOD1, ultimately reducing apoptosis and brain damage.
