The peripheral-type benzodiazepine receptor is involved in control of Ca2+-induced permeability transition pore opening in rat brain mitochondria
The peripheral-type benzodiazepine receptor is involved in control of Ca2+-induced permeability transition pore opening in rat brain mitochondria
复制标题
DOI：
10.1016/j.ceca.2006.11.004
复制
发表时间：
2007-07-01
期刊：
影响因子：
4
通讯作者：
Reiser, Georg
中科院分区：
文献类型：
--
作者：
Azarashvili, Tamara;Grachev, Dmitry;Reiser, Georg
外周型苯二氮卓受体 (PBR) 是一种 18 kDa 线粒体膜蛋白，在细胞死亡中的功能仍难以捉摸。在这里，我们研究了 PBR 是否参与 Ca2+ 诱导的大鼠离体脑线粒体 (RBM) 的通透性转变孔 (PTP) 打开。 PTP 开放对于导致程序性细胞死亡的线粒体事件非常重要。免疫印迹显示纯化 RBM 中存在单个 18 kDa 抗 PBR 抗体免疫反应带。在 PBR 免疫沉淀物中发现了腺嘌呤核苷酸转运蛋白，它是 PTP 的关键成分。在分离的完整 RBM 中，添加特定的抗 PBR 抗体 [H. Li，Z. Yao，B. Degenhardt，G. Teper，V. Papadopoulos，外周型苯二氮卓受体的胆固醇识别/相互作用氨基酸共识（CRAC）上的胆固醇结合以及 HIV TAT-CRAC 肽对类固醇生成的抑制，Proc。国家。阿卡德。科学。 U.S.A. 98 (2001) 1267-1272]延迟了 Ca2+ 诱导的膜电位耗散 (psi(m)) 并减少了环孢菌素 A 敏感的 Ca2+ 流出，这两者都表明抑制 PTP 打开。此外，尽管 psi(m) 消散，抗 PBR 抗体仍导致 Ca2+ 部分保留在线粒体基质中，并降低 Ca2+ 诱导的 PTP 打开时呼吸频率的激活。在 Ca2+ 负荷阈值时，RBM 会释放促凋亡因子、AIF 和细胞色素 c。抗PBR抗体阻断AIF的释放，但不影响细胞色素c的释放。添加 ATP 能够启动 PTP 关闭，与 psi(m) 恢复和 Ca2+ 重新积累相关。同时，线粒体蛋白磷酸化（从[γ-(32)p]ATP 掺入(12)p）发生，并且抗PBR 抗体能够抑制这些蛋白的磷酸化。内源性 PBR 配体原卟啉 IX 促进线粒体蛋白的 PTP 打开和磷酸化，从而诱导与抗 PBR 抗体相反的作用。这项研究为 PBR 参与 PTP 打开、控制 Ca2+ 诱导的 Ca2+ 外流以及线粒体中 AIF 的释放（程序性细胞死亡启动的重要阶段）提供了证据。 (C) 2006 Elsevier Ltd. 保留所有权利。
The peripheral-type benzodiazepine receptor (PBR) is an 18 kDa mitochondrial membrane protein with still elusive function in cell death. Here, we studied whether PBR is involved in Ca2+-induced permeability transition pore (PTP) opening in isolated rat brain mitochondria (RBM). PTP opening is important in mitochondrial events leading to programmed cell death. Immunoblots revealed a single 18 kDa anti-PBR antibody-immunoreactive band in purified RBM. Adenine nucleotide transporter, a key PTP component, was found in the PBR-immunoprecipitate. In isolated intact RBM, addition of a specific anti-PBR antibody [H. Li, Z. Yao, B. Degenhardt, G. Teper, V. Papadopoulos, Cholesterol binding at the cholesterol recognition/interaction amino acid consensus (CRAC) of the peripheral-type benzodiazepine receptor and inhibition of steroidogenesis by an HIV TAT-CRAC peptide, Proc. Natl. Acad. Sci. U.S.A. 98 (2001) 1267-1272] delayed Ca2+-induced dissipation of membrane potential (psi(m)) and diminished cyclosporine A-sensitive Ca2+ efflux, which are both indicative for the suppression of PTP opening. Moreover, anti-PBR antibody caused partial retention of Ca2+ in the mitochondrial matrix in spite of psi(m) dissipation, and reduced activation of respiratory rate at Ca2+-induced PTP opening. A release of pro-apoptotic factors, AIF and cytochrome c, from RBM was shown at threshold Ca2+ load. Anti-PBR antibody blocked the release of AIF but did not affect the cytochrome c release. Addition of ATP was able to initiate PTP closing, associated with psi(m) restoration and Ca2+ re-accumulation. At the same time mitochondrial protein phosphorylation (incorporation of (12)p from [gamma-(32)p]ATP) occurred and anti-PBR antibody was able to inhibit phosphorylation of these proteins. The endogenous PBR ligand, protoporphyrin IX, facilitated PTP opening and phosphorylation of the mitochondrial proteins, thus, inducing effects opposite to anti-PBR antibody. This study provides evidence for PBR involvement in PTP opening, controlling the Ca2+-induced Ca2+ efflux, and AIF release from mitochondria, important stages of initiation of programmed cell death. (C) 2006 Elsevier Ltd. All rights reserved.