BAK/BAX 大孔在细胞凋亡过程中促进线粒体突出和 mtDNA 外流,Science

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BAK/BAX 大孔在细胞凋亡过程中促进线粒体突出和 mtDNA 外流
Science ( IF 44.7 ) Pub Date : 2018-02-22 , DOI: 10.1126/science.aao6047
Kate McArthur _{1,

2,

3} , Lachlan W. Whitehead _{1,

2} , John M. Heddleston ₄ , Lucy Li ₁ , Benjamin S. Padman ₅ , Viola Oorschot ₆ , Niall D. Geoghegan _{1,

2} , Stephane Chappaz _{1,

2,

3} , Sophia Davidson ₁ , Hui San Chin ₁ , Rachael M. Lane ₃ , Marija Dramicanin _{1,

2} , Tahnee L. Saunders ₃ , Canny Sugiana ₃ , Romina Lessene _{1,

2} , Laura D. Osellame ₅ , Teng-Leong Chew ₄ , Grant Dewson _{1,

2} , Michael Lazarou ₅ , Georg Ramm _{5,

6} , Guillaume Lessene _{1,

2,

7} , Michael T. Ryan ₅ , Kelly L. Rogers _{1,

2} , Mark F. van Delft _{1,

2} , Benjamin T. Kile _{1,

2,

3}

Affiliation

大逃逸线粒体 DNA (mtDNA) 是一种有效的损伤相关分子模式，如果它到达细胞质或细胞外环境，就会触发先天免疫途径。mtDNA 信号传导与多种疾病有关。然而，mtDNA 释放的机制尚不清楚，迄今为止还没有实时观察到该过程。麦克阿瑟等人。使用活细胞晶格光片显微镜观察内在细胞凋亡过程中的 mtDNA 释放。促死亡蛋白 BAK 和 BAX 的激活导致线粒体外膜中大孔的形成。这些巨大的洞导致线粒体内膜膨胀到细胞质中，导致线粒体突出。这个过程允许线粒体基质的内容物，包括 mtDNA，逃逸到细胞质中。科学，这个问题 p。eaao6047 由于 BAK/BAX 诱导的线粒体疝，凋亡细胞中的线粒体 DNA 被释放。引言细胞死亡途径和损伤相关分子模式 (DAMP) 信号在塑造炎症和免疫反应中的作用引起了人们的极大兴趣。线粒体是内在细胞凋亡途径的核心，这是程序性细胞死亡的经典形式。一些线粒体成分被认为是 DAMP，包括线粒体 DNA (mtDNA)。最近的工作表明，内在 BAK 和 BAX 介导的细胞凋亡的激活导致先天免疫 cGAS/STING 途径的 mtDNA 依赖性触发，从而导致死亡细胞产生 I 型干扰素。凋亡半胱天冬酶级联反应通常会抑制这种 mtDNA 诱导的 cGAS/STING 信号传导，使细胞凋亡“免疫沉默”。基本原理据认为，在细胞凋亡过程中，mtDNA 被释放到细胞质中。除了细胞凋亡外，基质中 mtDNA 的丢失还与 HIV 和登革热感染、钙超载、辐射或炎症性疾病（如系统性红斑狼疮或类风湿性关节炎）有关。然而，尚未实时记录 mtDNA 从线粒体中逃逸的情况。结果结合活细胞晶格光片显微镜、3D 结构照明显微镜、相关光电子显微镜和电子冷冻断层扫描，我们发现在 BAK/BAX 激活和细胞色素 c 丢失后，线粒体网络崩溃，外膜出现大的 BAK/BAX 孔。这些 BAK/BAX 大孔使内膜成为突出的出口，携带线粒体基质成分，包括线粒体基因组。一部分突出的内膜失去了完整性，使 mtDNA 暴露在细胞质中。结论大量文献表明，mtDNA 存在于线粒体外——实际上，也存在于细胞外——在多种情况下。我们的研究提供了其从线粒体中释放的机制描述。细胞凋亡过程中线粒体释放 mtDNA 与 caspase 活性无关，但在正常细胞中，caspase 通过驱动细胞快速崩溃和清除来减弱随后的 cGAS/STING 介导的干扰素反应。线粒体疝可能代表了 mtDNA 逃逸的一般机制。除了 BAK 和 BAX 寡聚化之外，可能还有其他触发因素——例如，其他成孔蛋白（宿主或病原体衍生的）或线粒体应激——导致这种现象的发生。凋亡线粒体疝示意图。插图显示了由晶格光片显微镜、结构照明显微镜以及相关光和电子显微镜捕获的关键阶段。线粒体凋亡由 BAK 和 BAX 介导，这两种蛋白质可诱导线粒体外膜透化，导致细胞色素 c 释放和凋亡半胱天冬酶的激活。在没有活性半胱天冬酶的情况下，线粒体 DNA (mtDNA) 触发先天免疫 cGAS/STING 通路，导致垂死的细胞分泌 I 型干扰素。cGAS 如何获得 mtDNA 尚不清楚。我们使用活细胞晶格光片显微镜检查小鼠胚胎成纤维细胞中的线粒体网络。我们发现在 BAK/BAX 激活和细胞色素 c 丢失后，线粒体网络崩溃，外膜出现大的 BAK/BAX 孔。这些 BAK/BAX 大孔允许线粒体内膜突出到细胞质中，携带线粒体基质成分，包括线粒体基因组。凋亡的半胱天冬酶不能预防疝气，但会分解垂死的细胞以抑制 mtDNA 诱导的先天免疫信号。我们发现在 BAK/BAX 激活和细胞色素 c 丢失后，线粒体网络崩溃，外膜出现大的 BAK/BAX 孔。这些 BAK/BAX 大孔允许线粒体内膜突出到细胞质中，携带线粒体基质成分，包括线粒体基因组。凋亡的半胱天冬酶不能预防疝气，但会分解垂死的细胞以抑制 mtDNA 诱导的先天免疫信号。我们发现在 BAK/BAX 激活和细胞色素 c 丢失后，线粒体网络崩溃，外膜出现大的 BAK/BAX 孔。这些 BAK/BAX 大孔允许线粒体内膜突出到细胞质中，携带线粒体基质成分，包括线粒体基因组。凋亡的半胱天冬酶不能预防疝气，但会分解垂死的细胞以抑制 mtDNA 诱导的先天免疫信号。

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BAK/BAX macropores facilitate mitochondrial herniation and mtDNA efflux during apoptosis

The great escape Mitochondrial DNA (mtDNA) is a potent damage-associated molecular pattern that, if it reaches the cytoplasm or extracellular milieu, triggers innate immune pathways. mtDNA signaling has been implicated in a wide range of diseases; however, the mechanisms of mtDNA release are unclear, and the process has not been observed in real time thus far. McArthur et al. used live-cell lattice light-sheet microscopy to look at mtDNA release during intrinsic apoptosis. Activation of the pro-death proteins BAK and BAX resulted in the formation of large macro-pores in the mitochondrial outer membrane. These massive holes caused the inner mitochondrial membrane to balloon out into the cytoplasm, resulting in mitochondrial herniation. This process allowed the contents of the mitochondrial matrix, including mtDNA, to escape into the cytoplasm. Science, this issue p. eaao6047 Mitochondrial DNA is released from mitochondria in apoptotic cells as a result of BAK/BAX-induced mitochondrial herniation. INTRODUCTION There has been an explosion of interest in the role of cell death pathways and damage-associated molecular pattern (DAMP) signaling in shaping inflammatory and immune responses. Mitochondria are central to the intrinsic apoptosis pathway, the classical form of programmed cell death. Several mitochondrial constituents have been implicated as DAMPs, including mitochondrial DNA (mtDNA). Recent work has shown that activation of intrinsic BAK and BAX–mediated apoptosis results in mtDNA-dependent triggering of the innate immune cGAS/STING pathway, resulting in type I interferon production by dying cells. The apoptotic caspase cascade normally functions to suppress this mtDNA-induced cGAS/STING signaling, rendering apoptosis “immunologically silent.” RATIONALE It is thought that during apoptosis, mtDNA is released into the cytoplasm. In addition to apoptosis, loss of mtDNA from the matrix has been associated with conditions including HIV and dengue infection, calcium overload, irradiation, or inflammatory diseases such as systemic lupus erythematosus or rheumatoid arthritis. However, mtDNA escape from the mitochondria has not been documented in real time. RESULTS Using a combination of live-cell lattice light-sheet microscopy, 3D structured illumination microscopy, correlative light electron microscopy, and electron cryotomography, we found that after BAK/BAX activation and cytochrome c loss, the mitochondrial network broke down and large BAK/BAX pores appeared in the outer membrane. These BAK/BAX macropores allowed the inner membrane an outlet through which it herniated, carrying with it mitochondrial matrix components, including the mitochondrial genome. A subset of the herniated inner membranes lost their integrity, allowing mtDNA to be exposed to the cytoplasm. CONCLUSION An extensive literature suggests that mtDNA is found outside the mitochondria—and, indeed, outside the cell—in a wide range of circumstances. Our study provides a mechanistic description of its release from the mitochondria. mtDNA release from mitochondria during apoptosis occurs irrespective of caspase activity, but in normal cells, caspases attenuate the subsequent cGAS/STING-mediated interferon response by driving rapid cellular collapse and clearance. Mitochondrial herniation might represent a general mechanism of mtDNA escape. In addition to BAK and BAX oligomerization, there may be alternative triggers—for example, other pore-forming proteins (host- or pathogen-derived) or mitochondrial stresses—that lead to the occurrence of this phenomenon. Schematic of apoptotic mitochondrial herniation. Inset images show the key stages as captured by lattice light-sheet microscopy, structured illumination microscopy, and correlative light and electron microscopy. Mitochondrial apoptosis is mediated by BAK and BAX, two proteins that induce mitochondrial outer membrane permeabilization, leading to cytochrome c release and activation of apoptotic caspases. In the absence of active caspases, mitochondrial DNA (mtDNA) triggers the innate immune cGAS/STING pathway, causing dying cells to secrete type I interferon. How cGAS gains access to mtDNA remains unclear. We used live-cell lattice light-sheet microscopy to examine the mitochondrial network in mouse embryonic fibroblasts. We found that after BAK/BAX activation and cytochrome c loss, the mitochondrial network broke down and large BAK/BAX pores appeared in the outer membrane. These BAK/BAX macropores allowed the inner mitochondrial membrane to herniate into the cytosol, carrying with it mitochondrial matrix components, including the mitochondrial genome. Apoptotic caspases did not prevent herniation but dismantled the dying cell to suppress mtDNA-induced innate immune signaling.

更新日期：2018-02-22

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