Jones Edith, Gaytan Norma, Garcia Iraselia, Herrera Alan, Ramos Manuel, Agarwala Divya, Rana Maahrose, Innis-Whitehouse Wendy, Schuenzel Erin, Gilkerson Robert
Department of Biology, The University of Texas Rio Grande Valley, 1201 West University Drive, Edinburg, TX, 78539-2999, USA.
Department of Biomedical Sciences, The University of Texas Rio Grande Valley, Edinburg, TX, 78539-2999, USA.
Cell Mol Life Sci. 2017 Apr;74(7):1347-1363. doi: 10.1007/s00018-016-2421-9. Epub 2016 Nov 17.
As an organellar network, mitochondria dynamically regulate their organization via opposing fusion and fission pathways to maintain bioenergetic homeostasis and contribute to key cellular pathways. This dynamic balance is directly linked to bioenergetic function: loss of transmembrane potential across the inner membrane (Δψ ) disrupts mitochondrial fission/fusion balance, causing fragmentation of the network. However, the level of Δψ required for mitochondrial dynamic balance, as well as the relative contributions of fission and fusion pathways, have remained unclear. To explore this, mitochondrial morphology and Δψ were examined via confocal imaging and tetramethyl rhodamine ester (TMRE) flow cytometry, respectively, in cultured 143B osteosarcoma cells. When normalized to the TMRE value of untreated 143B cells as 100%, both genetic (mtDNA-depleted ρ) and pharmacological [carbonyl cyanide m-chlorophenyl hydrazone (CCCP)-treated] cell models below 34% TMRE fluorescence were unable to maintain mitochondrial interconnection, correlating with loss of fusion-active long OPA1 isoforms (L-OPA1). Mechanistically, this threshold is maintained by mechanistic coordination of DRP1-mediated fission and OPA1-mediated fusion: cells lacking either DRP1 or the OMA1 metalloprotease were insensitive to loss of Δψ , instead maintaining an obligately fused morphology. Collectively, these findings demonstrate a mitochondrial 'tipping point' threshold mediated by the interaction of Δψ with both DRP1 and OMA1; moreover, DRP1 appears to be required for effective OPA1 maintenance and processing, consistent with growing evidence for direct interaction of fission and fusion pathways. These results suggest that Δψ below threshold coordinately activates both DRP1-mediated fission and OMA1 cleavage of OPA1, collapsing mitochondrial dynamic balance, with major implications for a range of signaling pathways and cellular life/death events.
作为一个细胞器网络，线粒体通过相反的融合和裂变途径动态调节其组织，以维持生物能量稳态并参与关键的细胞途径。这种动态平衡与生物能量功能直接相关：内膜跨膜电位（Δψ）的丧失会破坏线粒体的裂变/融合平衡，导致网络碎片化。然而，线粒体动态平衡所需的Δψ水平以及裂变和融合途径的相对贡献仍不清楚。为了探究这一点，分别通过共聚焦成像和四甲基罗丹明酯（TMRE）流式细胞术在培养的143B骨肉瘤细胞中检测线粒体形态和Δψ。当以未处理的143B细胞的TMRE值作为100%进行归一化时，TMRE荧光低于34%的遗传（线粒体DNA缺失的ρ）和药理学[羰基氰化物间氯苯腙（CCCP）处理]细胞模型均无法维持线粒体的相互连接，这与融合活性长OPA1异构体（L-OPA1）的丧失相关。从机制上讲，这个阈值是由动力蛋白相关蛋白1（DRP1）介导的裂变和OPA1介导的融合的机制协调维持的：缺乏DRP1或OMA1金属蛋白酶的细胞对Δψ的丧失不敏感，而是保持一种必然融合的形态。总的来说，这些发现证明了由Δψ与DRP1和OMA1的相互作用介导的线粒体“临界点”阈值；此外，DRP1似乎是有效维持和加工OPA1所必需的，这与裂变和融合途径直接相互作用的越来越多的证据一致。这些结果表明，低于阈值的Δψ协同激活DRP1介导的裂变和OMA1对OPA1的切割，破坏线粒体动态平衡，对一系列信号通路和细胞生死事件具有重要意义。
