Fernandez-Sanz Celia, De la Fuente Sergio, Nichtova Zuzana, Federico Marilen, Duvezin-Caubet Stephane, Lanvermann Sebastian, Tsai Hui-Ying, Xin Yanguo, Csordas Gyorgy, Wang Wang, Mourier Arnaud, Sheu Shey-Shing
Center for Translational Medicine, Department of Medicine, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA.
University of Bordeaux, CNRS, IBGC, UMR 5095, F-33000 Bordeaux, France.
Cells. 2025 Aug 14;14(16):1259. doi: 10.3390/cells14161259.
Mitochondrial fission and fusion appear to be relatively infrequent in cardiac cells compared to other cell types; however, the proteins involved in these events are highly expressed in adult cardiomyocytes (ACM). Therefore, these proteins likely have additional non-canonical roles. We have previously shown that DRP1 not only participates in mitochondrial fission processes but also regulates mitochondrial bioenergetics in cardiac tissue. However, it is still unknown where the DRP1 that does not participate in mitochondrial fission is located and what its role is at those non-fission spots. Therefore, this manuscript will clarify whether oligomeric DRP1 is located at the SR-mitochondria interface, a specific region that harbors the Ca microdomains created by Ca release from the SR through the RyR2. The high Ca microdomains and the subsequent Ca uptake by mitochondria through the mitochondrial Ca uniporter complex (MCUC) are essential to regulate mitochondrial bioenergetics during excitation-contraction (EC) coupling. Herein, we aimed to test the hypothesis that mitochondria-bound DRP1 preferentially accumulates at the mitochondria-SR contacts to deploy its function on regulating mitochondrial bioenergetics and that this strategic position is modulated by calcium in a beat-to-beat manner. In addition, the mechanism responsible for such a biased distribution and its functional implications was investigated. High-resolution imaging approaches, cell fractionation, Western blot, 2D blue native gel electrophoresis, and immunoprecipitations were applied to both electrically paced ACM and Langendorff-perfused beating hearts to elucidate the mechanisms of the strategic DRP1 localization. Our data show that in ACM, mitochondria-bound DRP1 clusters in high molecular weight protein complexes at mitochondria-associated membrane (MAM). This clustering requires DRP1 interaction with β-ACTIN and is fortified by EC coupling-mediated Ca transients. In ACM, DRP1 is anchored at the mitochondria-SR contacts through interactions with β-ACTIN and Ca transients, playing a fundamental role in regulating mitochondrial physiology.
与其他细胞类型相比，线粒体分裂和融合在心脏细胞中似乎相对不常见；然而，参与这些过程的蛋白质在成年心肌细胞（ACM）中高度表达。因此，这些蛋白质可能具有额外的非经典作用。我们之前已经表明，动力相关蛋白1（DRP1）不仅参与线粒体分裂过程，还调节心脏组织中的线粒体生物能量学。然而，不参与线粒体分裂的DRP1位于何处以及它在那些非分裂位点的作用仍然未知。因此，本手稿将阐明寡聚体DRP1是否位于肌浆网-线粒体界面，这是一个特定区域，含有通过肌浆网中兰尼碱受体2（RyR2）释放钙所产生的钙微区。高钙微区以及随后线粒体通过线粒体钙单向转运体复合物（MCUC）摄取钙对于在兴奋-收缩（EC）偶联期间调节线粒体生物能量学至关重要。在此，我们旨在检验以下假设：与线粒体结合的DRP1优先聚集在线粒体-肌浆网接触部位，以发挥其调节线粒体生物能量学的功能，并且这种战略位置受到钙的逐搏调节。此外，还研究了造成这种偏向分布的机制及其功能意义。高分辨率成像方法、细胞分级分离、蛋白质免疫印迹、二维蓝色天然凝胶电泳和免疫沉淀法被应用于电刺激的ACM和Langendorff灌注的跳动心脏，以阐明DRP1战略定位的机制。我们的数据表明，在ACM中，与线粒体结合的DRP1聚集在线粒体相关膜（MAM）处的高分子量蛋白质复合物中。这种聚集需要DRP1与β-肌动蛋白相互作用，并由EC偶联介导的钙瞬变增强。在ACM中，DRP1通过与β-肌动蛋白和钙瞬变相互作用而锚定在线粒体-肌浆网接触部位，在调节线粒体生理学中发挥重要作用。
