期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2015
卷号：112
期号：1
页码：130-135
DOI：10.1073/pnas.1415261112
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceThe question as to how the proton motive force in mitochondria is distributed among the proteins that require a proton gradient for their work is one of the central unresolved questions in mitochondrial physiology and important for the mechanistic insight in the function of mitochondrial proteins. Our results suggest that the local separation of the proteins on the inner mitochondrial membrane makes it impossible for uncoupling protein 4 (UCP4) to uncouple phosphorylation from proton pumping. Nonetheless, UCP4 should be well able to shortcut excessive transmembrane proton gradients to thereby regulate reactive oxygen species production. It explains how the proton transporter may fulfill that function without being a real UCP like UCP1. Because different proteins compete for the proton gradient across the inner mitochondrial membrane, an efficient mechanism is required for allocation of associated chemical potential to the distinct demands, such as ATP production, thermogenesis, regulation of reactive oxygen species (ROS), etc. Here, we used the superresolution technique dSTORM (direct stochastic optical reconstruction microscopy) to visualize several mitochondrial proteins in primary mouse neurons and test the hypothesis that uncoupling protein 4 (UCP4) and F0F1-ATP synthase are spatially separated to eliminate competition for the proton motive force. We found that UCP4, F0F1-ATP synthase, and the mitochondrial marker voltage-dependent anion channel (VDAC) have various expression levels in different mitochondria, supporting the hypothesis of mitochondrial heterogeneity. Our experimental results further revealed that UCP4 is preferentially localized in close vicinity to VDAC, presumably at the inner boundary membrane, whereas F0F1-ATP synthase is more centrally located at the cristae membrane. The data suggest that UCP4 cannot compete for protons because of its spatial separation from both the proton pumps and the ATP synthase. Thus, mitochondrial morphology precludes UCP4 from acting as an uncoupler of oxidative phosphorylation but is consistent with the view that UCP4 may dissipate the excessive proton gradient, which is usually associated with ROS production.
关键词：mitochondrial membrane proteins ; proton diffusion ; direct stochastic optical reconstruction microscopy ; uncoupling ; reactive oxygen species
