D'Orsi Beatrice, Engel Tobias, Pfeiffer Shona, Nandi Saheli, Kaufmann Thomas, Henshall David C, Prehn Jochen H M
Department of Physiology and Medical Physics, Centre for the Study of Neurological Disorders, Royal College of Surgeons in Ireland, Dublin 2, Ireland, and.
Institute of Pharmacology, University of Bern, Bern CH-3010, Switzerland.
J Neurosci. 2016 Apr 20;36(16):4564-78. doi: 10.1523/JNEUROSCI.3780-15.2016.
Bok (Bcl-2-related ovarian killer) is a Bcl-2 family member that, because of its predicted structural homology to Bax and Bak, has been proposed to be a pro-apoptotic protein. In this study, we demonstrate that Bok is highly expressed in neurons of the mouse brain but that bok was not required for staurosporine-, proteasome inhibition-, or excitotoxicity-induced apoptosis of cultured cortical neurons. On the contrary, we found that bok-deficient neurons were more sensitive to oxygen/glucose deprivation-induced injury in vitro and seizure-induced neuronal injury in vivo Deletion of bok also increased staurosporine-, excitotoxicity-, and oxygen/glucose deprivation-induced cell death in bax-deficient neurons. Single-cell imaging demonstrated that bok-deficient neurons failed to maintain their neuronal Ca(2+)homeostasis in response to an excitotoxic stimulus; this was accompanied by a prolonged deregulation of mitochondrial bioenergetics.bok deficiency led to a specific reduction in neuronal Mcl-1 protein levels, and deregulation of both mitochondrial bioenergetics and Ca(2+)homeostasis was rescued by Mcl-1 overexpression. Detailed analysis of cell death pathways demonstrated the activation of poly ADP-ribose polymerase-dependent cell death in bok-deficient neurons. Collectively, our data demonstrate that Bok acts as a neuroprotective factor rather than a pro-death effector during Ca(2+)- and seizure-induced neuronal injury in vitro and in vivo
Bcl-2 proteins are essential regulators of the mitochondrial apoptosis pathway. The Bcl-2 protein Bok is highly expressed in the CNS. Because of its sequence similarity to Bax and Bak, Bok has long been considered part of the pro-apoptotic Bax-like subfamily, but no studies have yet been performed in neurons to test this hypothesis. Our study provides important new insights into the functional role of Bok during neuronal apoptosis and specifically in the setting of Ca(2+)- and seizure-mediated neuronal injury. We show that Bok controls neuronal Ca(2+)homeostasis and bioenergetics and, contrary to previous assumptions, exerts neuroprotective activities in vitro and in vivo Our results demonstrate that Bok cannot be placed unambiguously into the Bax-like Bcl-2 subfamily of pro-apoptotic proteins.
Bok（Bcl-2相关卵巢杀手）是一种Bcl-2家族成员，因其与Bax和Bak预测的结构同源性，被认为是一种促凋亡蛋白。在本研究中，我们证明Bok在小鼠大脑神经元中高表达，但在培养的皮质神经元中，星形孢菌素、蛋白酶体抑制或兴奋性毒性诱导的凋亡过程中，bok并非必需。相反，我们发现bok缺陷型神经元在体外对氧/葡萄糖剥夺诱导的损伤以及在体内对癫痫发作诱导的神经元损伤更敏感。敲除bok也增加了bax缺陷型神经元中星形孢菌素、兴奋性毒性和氧/葡萄糖剥夺诱导的细胞死亡。单细胞成像表明，bok缺陷型神经元在受到兴奋性毒性刺激时无法维持其神经元钙（Ca2+）稳态；这伴随着线粒体生物能量学的长期失调。bok缺陷导致神经元Mcl-1蛋白水平特异性降低，而线粒体生物能量学和钙（Ca2+）稳态的失调通过Mcl-1过表达得以挽救。对细胞死亡途径的详细分析表明，bok缺陷型神经元中聚ADP-核糖聚合酶依赖性细胞死亡被激活。总体而言，我们的数据表明，在体外和体内Ca2+和癫痫发作诱导的神经元损伤过程中，Bok作为一种神经保护因子而非促死亡效应器发挥作用。
Bcl-2蛋白是线粒体凋亡途径的重要调节因子。Bcl-2蛋白Bok在中枢神经系统中高表达。由于其与Bax和Bak的序列相似性，Bok长期以来一直被认为是促凋亡的Bax样亚家族的一部分，但尚未在神经元中进行研究来验证这一假设。我们的研究为Bok在神经元凋亡过程中，特别是在Ca2+和癫痫介导的神经元损伤情况下的功能作用提供了重要的新见解。我们表明Bok控制神经元钙（Ca2+）稳态和生物能量学，与先前的假设相反，在体外和体内发挥神经保护作用。我们的结果表明，Bok不能明确地归入促凋亡蛋白的Bax样Bcl-2亚家族。
