Research Institute of Ophthalmology and Visual Sciences, Affiliated Eye Hospital of Nanchang University, Nanchang, China.
Department of Pharmacology, School of Pharmaceutical Science, Nanchang University, Nanchang, China.
J Cell Biochem. 2018 Jun;119(6):4815-4825. doi: 10.1002/jcb.26680. Epub 2018 Mar 7.
Alzheimer's disease, one of the most common neurodegenerative diseases, is pathologically characterized by Amyloid beta containing plaques and neurofibrillary tangles. Amyloid beta (Aβ) induces neuronal apoptosis through the intracellular Ca increase, subsequent hyperactivation of cyclin-dependent kinase 5 (Cdk5) and mitochondrial abnormality. Recently, Cdk5 was identified as an upstream regulator of mitochondrial fission during neuronal apoptosis, but the underlying mechanism remains unclear. Here, in vitro phosphorylation assays showed that Cdk5 could phosphorylate the recombinant Drp1 at Serine 579. Aβ stimulation increased the phosphorylation level of Drp1 at Serine 579 in mouse cortical neurons. Cdk5 inhibitor roscovitine and knockdown of Cdk5 by a lentiviral vector expressing shRNA targeting Cdk5 (Lenti-Cdk5-shRNA) efficiently prevented Aβ induced Drp1 phosphorylation in neurons. In addition, Aβ stimulation induced markedly mitochondrial fission in neurons. Roscovitine, Lenti-Cdk5-shRNA and expression of phospho-defect mutatant GFP-Drp1-S579A in neurons attenuated Aβ induced mitochondrial fission, whereas expression of phospho-mimetic mutant GFP-Drp1-S579D alone resulted in mitochondiral fission similar to Aβ stimulation. Moreover, Roscovitine and Lenti-Cdk5-shRNA suppressed the cleavage of caspase-3 and protected neurons against Aβ induced neuronal apoptosis.Thus, our data indicate that Drp1 is a direct target of Cdk5, and Cdk5-mediated phosphorylation of Drp1 at Serine 579 regulates Aβ induced mitochondrial fission and neuronal toxicity.
阿尔茨海默病是最常见的神经退行性疾病之一，其病理特征为含有淀粉样β的斑块和神经原纤维缠结。淀粉样β（Aβ）通过细胞内 Ca 增加、随后细胞周期蛋白依赖性激酶 5（Cdk5）的过度激活和线粒体异常诱导神经元凋亡。最近，Cdk5 被鉴定为神经元凋亡过程中线粒体分裂的上游调节因子，但潜在的机制尚不清楚。在这里，体外磷酸化实验表明 Cdk5 可以在丝氨酸 579 处磷酸化重组 Drp1。Aβ 刺激增加了小鼠皮质神经元中 Drp1 丝氨酸 579 的磷酸化水平。Cdk5 抑制剂罗克洛文（roscovitine）和通过表达靶向 Cdk5 的 shRNA 的慢病毒载体（Lenti-Cdk5-shRNA）的敲低 Cdk5 可有效防止 Aβ 诱导的神经元中 Drp1 的磷酸化。此外，Aβ 刺激诱导神经元中线粒体明显分裂。罗克洛文（roscovitine）、Lenti-Cdk5-shRNA 和在神经元中表达磷酸缺陷突变 GFP-Drp1-S579A 减弱了 Aβ 诱导的线粒体分裂，而单独表达磷酸模拟突变 GFP-Drp1-S579D 导致类似 Aβ 刺激的线粒体分裂。此外，罗克洛文（roscovitine）和 Lenti-Cdk5-shRNA 抑制了 caspase-3 的切割，并保护神经元免受 Aβ 诱导的神经元凋亡。因此，我们的数据表明 Drp1 是 Cdk5 的直接靶标，Cdk5 介导的 Drp1 丝氨酸 579 磷酸化调节 Aβ 诱导的线粒体分裂和神经元毒性。
