Mitochondrial fusion is required for regulation of mitochondrial DNA replication.

Eduardo Silva Ramos,Johanna H K Kauppila,Jakob D Busch,Arnaud Mourier,Christian Brüser,Elisa Motori,Kjell Hultenby,Inge Kühl,Benedetta Ruzzenente,Nils-Göran Larsson,Assa Yeroslaviz,Bianca H Habermann,Stefan Jakobs

doi:10.1371/journal.pgen.1008085

Abstract

Mitochondrial dynamics is an essential physiological process controlling mitochondrial content mixing and mobility to ensure proper function and localization of mitochondria at intracellular sites of high-energy demand. Intriguingly, for yet unknown reasons, severe impairment of mitochondrial fusion drastically affects mtDNA copy number. To decipher the link between mitochondrial dynamics and mtDNA maintenance, we studied mouse embryonic fibroblasts (MEFs) and mouse cardiomyocytes with disruption of mitochondrial fusion. Super-resolution microscopy revealed that loss of outer mitochondrial membrane (OMM) fusion, but not inner mitochondrial membrane (IMM) fusion, leads to nucleoid clustering. Remarkably, fluorescence in situ hybridization (FISH), bromouridine labeling in MEFs and assessment of mitochondrial transcription in tissue homogenates revealed that abolished OMM fusion does not affect transcription. Furthermore, the profound mtDNA depletion in mouse hearts lacking OMM fusion is not caused by defective integrity or increased mutagenesis of mtDNA, but instead we show that mitochondrial fusion is necessary to maintain the stoichiometry of the protein components of the mtDNA replisome. OMM fusion is necessary for proliferating MEFs to recover from mtDNA depletion and for the marked increase of mtDNA copy number during postnatal heart development. Our findings thus link OMM fusion to replication and distribution of mtDNA.

Highlights

Mammalian mitochondria are dynamic organelles present as long interconnected tubules or individual units that may undergo intracellular transport [1,2]
Loss of outer mitochondrial membrane (OMM) fusion leads to impaired mtDNA maintenance and oxidative phosphorylation (OXPHOS) deficiency To study the link between mitochondrial fusion and mtDNA maintenance, we generated heart dMfn KO mice, which were born at the expected Mendelian ratios (S1A Fig) and showed loss of transcripts encoding MFN1 and mitofusin 2 (MFN2) in heart tissue (S1B Fig)
Transmission electron microscopy analysis of heart tissue revealed that dMfn KO mice exhibited a significant increase in ratio of mitochondrial/ cytoplasmic area associated with aberrant mitochondrial ultrastructure and disruption of the myofibril organization (Fig 1B and 1C)

Summary

Author summary

Mammalian mitochondria contain multiple copies of the mitochondrial genome (mtDNA), which encodes genes that are essential for the oxidative phosphorylation. An important feature of mtDNA is that it is evenly distributed throughout the mitochondrial network. Dynamin-related GTPase proteins help control the size and shape of mitochondria by fusion and fission events and are intimately linked to maintenance and distribution of mtDNA. We demonstrate that rapid mtDNA synthesis in proliferating tissue-culture cells or cardiomyocytes during post-natal heart development requires mitochondrial fusion. The absence of mitochondrial fusion in mouse heart is not associated with mtDNA integrity defects but instead affects the replication of mtDNA. These findings provide direct evidence for the importance of mitochondrial fusion in maintaining mtDNA replication

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