Departments of Molecular and Clinical Cancer Medicine, University of Liverpool, Ashton Street, Liverpool, L69 3GE, UK.
Molecular and Clinical Pharmacology, University of Liverpool, Ashton Street, Liverpool, L69 3GE, UK.
Cell Death Dis. 2019 Jul 8;10(7):521. doi: 10.1038/s41419-019-1759-y.
The endoplasmic reticulum (ER) with its elaborate network of highly curved tubules and flat sheets interacts with several other organelles, including mitochondria, peroxisomes and endosomes, to play vital roles in their membrane dynamics and functions. Previously, we identified structurally diverse chemicals from different pharmacological classes, which induce a reversible reorganisation of ER membranes. Using apogossypol as a prototypic tool compound, we now show that ER membrane reorganisation occurs at the level of ER tubules but does not involve ER sheets. Reorganisation of ER membranes prevents DRP-1-mediated mitochondrial fission, thereby antagonising the functions of several mitochondrial fission-inducing agents. Previous reports have suggested that ER membranes mark the constriction sites of mitochondria by localising DRP-1, as well as BAX on mitochondrial membranes to facilitate both mitochondrial fission and outer membrane permeabilisation. Following ER membrane reorganisation and subsequent exposure to an apoptotic stimulus (BH3 mimetics), DRP-1 still colocalises with the reorganised ER membranes but BAX translocation and activation, cytochrome c release and phosphatidylserine externalisation are all inhibited, thereby diminishing the ability of BH3 mimetics to induce the intrinsic apoptotic pathway. Strikingly, both ER membrane reorganisation and its resulting inhibition of apoptosis could be reversed by inhibitors of dihydroorotate dehydrogenase (DHODH), namely teriflunomide and its active metabolite, leflunomide. However, neither genetic inhibition of DHODH using RNA interference nor metabolic supplementation with orotate or uridine to circumvent the consequences of a loss of DHODH activity rescued the effects of DHODH inhibitors, suggesting that the effects of these inhibitors in preventing ER membrane reorganisation is most likely independent of their ability to antagonise DHODH activity. Our results strengthen the hypothesis that ER is fundamental for key mitochondrial functions, such as fusion-fission dynamics and apoptosis.
内质网（ER）拥有错综复杂的高度弯曲小管和扁片网络，与其他几种细胞器相互作用，包括线粒体、过氧化物酶体和内体，在它们的膜动力学和功能中发挥着至关重要的作用。此前，我们从不同的药理学类别中鉴定出了结构多样的化学物质，这些物质会诱导 ER 膜的可逆重排。我们现在使用阿朴戈司泊（apogossypol）作为典型的工具化合物来展示 ER 膜重排发生在 ER 小管的水平，但不涉及 ER 片层。ER 膜的重排阻止了 DRP-1 介导的线粒体裂变，从而拮抗了几种诱导线粒体裂变的试剂的功能。之前的报告表明，ER 膜通过将 DRP-1 以及 BAX 定位在线粒体膜上来标记线粒体的缢缩部位，从而促进线粒体裂变和外膜通透化。在 ER 膜重排以及随后暴露于凋亡刺激物（BH3 模拟物）后，DRP-1 仍然与重排的 ER 膜共定位，但 BAX 易位和激活、细胞色素 c 释放和磷脂酰丝氨酸外化都受到抑制，从而降低了 BH3 模拟物诱导内在凋亡途径的能力。引人注目的是，ER 膜重排及其随后对凋亡的抑制作用都可以被二氢乳清酸脱氢酶（DHODH）抑制剂逆转，即特立氟胺及其活性代谢物来氟米特。然而，使用 RNA 干扰对 DHODH 的遗传抑制，或通过补充乳清酸或尿嘧啶来避免 DHODH 活性丧失的后果，都不能挽救 DHODH 抑制剂的作用，这表明这些抑制剂在防止 ER 膜重排中的作用很可能独立于它们拮抗 DHODH 活性的能力。我们的结果强化了内质网对于关键的线粒体功能（如融合裂变动力学和凋亡）至关重要的假设。
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