Shao Bryanna, Killion Mason, Oliver Ashton, Vang Chia, Zeleke Faben, Neikirk Kit, Vue Zer, Garza-Lopez Edgar, Shao Jian-Qiang, Mungai Margaret, Lam Jacob, Williams Qiana, Altamura Christopher T, Whiteside Aaron, Kabugi Kinuthia, McKenzie Jessica, Koh Alice, Scudese Estevão, Vang Larry, Marshall Andrea G, Crabtree Amber, Tanghal Janelle I, Stephens Dominique, Koh Ho-Jin, Jenkins Brenita C, Murray Sandra A, Cooper Anthonya T, Williams Clintoria, Damo Steven M, McReynolds Melanie R, Gaddy Jennifer A, Wanjalla Celestine N, Beasley Heather K, Hinton Antentor
Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN, 37232, USA.
Department of Internal Medicine, University of Iowa, Iowa City, IA, 52242, USA.
bioRxiv. 2023 Oct 19:2023.05.20.541602. doi: 10.1101/2023.05.20.541602.
The Sorting and Assembly Machinery (SAM) Complex is responsible for assembling β-barrel proteins in the mitochondrial membrane. Comprising three subunits, Sam35, Sam37, and Sam50, the SAM complex connects the inner and outer mitochondrial membranes by interacting with the mitochondrial contact site and cristae organizing system (MICOS) complex. Sam50, in particular, stabilizes the mitochondrial intermembrane space bridging (MIB) complex, which is crucial for protein transport, respiratory chain complex assembly, and regulation of cristae integrity. While the role of Sam50 in mitochondrial structure and metabolism in skeletal muscle remains unclear, this study aims to investigate its impact. Serial block-face-scanning electron microscopy (SBF-SEM) and computer-assisted 3D renderings were employed to compare mitochondrial structure and networking in -deficient myotubes from mice and humans with wild-type (WT) myotubes. Furthermore, autophagosome 3D structure was assessed in human myotubes. Mitochondrial metabolic phenotypes were assessed using Gas Chromatography-Mass Spectrometry-based metabolomics to explore differential changes in WT and -deficient myotubes. The results revealed increased mitochondrial fragmentation and autophagosome formation in -deficient myotubes compared to controls. Metabolomic analysis indicated elevated metabolism of propanoate and several amino acids, including ß-Alanine, phenylalanine, and tyrosine, along with increased amino acid and fatty acid metabolism in -deficient myotubes. Furthermore, impairment of oxidative capacity was observed upon ablation in both murine and human myotubes, as measured with the XF24 Seahorse Analyzer. Collectively, these findings support the critical role of Sam50 in establishing and maintaining mitochondrial integrity, cristae structure, and mitochondrial metabolism. By elucidating the impact of -deficiency, this study enhances our understanding of mitochondrial function in skeletal muscle.
分选与组装机制（SAM）复合体负责在线粒体内膜中组装β-桶状蛋白。SAM复合体由三个亚基Sam35、Sam37和Sam50组成，通过与线粒体接触位点和嵴组织系统（MICOS）复合体相互作用，连接线粒体内外膜。特别是Sam50可稳定线粒体膜间隙桥接（MIB）复合体，这对于蛋白质运输、呼吸链复合体组装以及嵴完整性的调节至关重要。虽然Sam50在骨骼肌线粒体结构和代谢中的作用尚不清楚，但本研究旨在探究其影响。采用连续块面扫描电子显微镜（SBF-SEM）和计算机辅助3D渲染技术，比较小鼠和人类Sam50缺陷型肌管与野生型（WT）肌管的线粒体结构和网络。此外，还评估了人类肌管中的自噬体3D结构。使用基于气相色谱-质谱联用的代谢组学技术评估线粒体代谢表型，以探索WT和Sam50缺陷型肌管中的差异变化。结果显示与对照组相比，Sam50缺陷型肌管中的线粒体碎片化和自噬体形成增加。代谢组学分析表明，Sam50缺陷型肌管中丙酸盐和几种氨基酸（包括β-丙氨酸、苯丙氨酸和酪氨酸）的代谢升高，同时氨基酸和脂肪酸代谢增加。此外，用XF24 Seahorse分析仪测量发现，在小鼠和人类肌管中敲除Sam50后均观察到氧化能力受损。总的来说，这些发现支持了Sam50在建立和维持线粒体完整性、嵴结构以及线粒体代谢中的关键作用。通过阐明Sam50缺陷的影响，本研究加深了我们对骨骼肌线粒体功能的理解。
