Ablation of Sam50 is associated with fragmentation and alterations in metabolism in murine and human myotubes

Abstract

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 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 and computer‐assisted 3D renderings were employed to compare mitochondrial structure and networking in Sam50‐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 Sam50‐deficient myotubes. The results revealed increased mitochondrial fragmentation and autophagosome formation in Sam50‐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 Sam50‐deficient myotubes. Furthermore, impairment of oxidative capacity was observed upon Sam50 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 Sam50‐deficiency, this study enhances our understanding of mitochondrial function in skeletal muscle.

MICOS Complex Loss Governs Age-Associated Murine Mitochondrial Architecture and Metabolism in the Liver, While Sam50 Dictates Diet Changes

“…Previously, we have performed a rigorous 3D analysis of mitochondrial structure and networking in Sam50 -deficient human and murine myotubes, showing that the deletion of Sam50 results in smaller and less complex mitochondria (52). To further investigate SAM50’s role in mitochondrial structure, we knocked out Sam50 in fibroblasts and used mito-mCherry confocal to investigate structure (Figures 5N-O’’).…”

“…Previously, we found that Sam50 is responsible for establishing and maintaining mitochondrial integrity, cristae structure, and mitochondrial metabolism (52). To identify the clinical implications of dysregulated SAMM50 gene expression in humans, we modeled genetically regulated gene expression (GReX) of SAMM50 across 85,615 individuals in the Vanderbilt biobank, BioVU (Table 1).…”

“…SAM50 depletion over time affects the protein contents of all large respiratory complexes containing subunits encoded by the mitochondria, suggesting a link between SAM50 and cristae structural integrity, respiratory complex assembly, and mitochondrial DNA (mtDNA) maintenance (107). Our previous study have shown that Sam50 plays a significant role in the maintenance of both the mitochondrial network and the structure of cristae and MICOS, especially in the context of liver injury (52). Mice with Sam50 deletion or liver-specific deletion produced by acetaminophen, an efficient antipyretic and analgesic, experience mtDNA release, which activates the cGAS-STING pathway and causes liver inflammation (81).…”

“…Sam50 maintains the formation of the cristae via interaction with mitofilin and CHCHD6, which are parts of the mitochondrial contact site and cristae organizing system (MICOS) (51). Furthermore, Sam50 aids in the maintenance of the mitochondrial network (50), establishing and maintaining mitochondrial integrity and mitochondrial metabolism, with its loss conferring volumetric losses in mitochondria structure (52). Recently, Sam50 has been linked to NAFLD pathogenesis (49), but the interconnection with aging liver mitochondrial structure remains poorly elucidated.…”