Mitochondrial diversity probed in mouse cerebellum elucidates cell type-specific fine-tuning of mitochondrial biology

Mitochondria house a variety of cellular functions, including catabolic and anabolic pathways, apoptosis and Ca2+ handling. These functions critically depend on nuclear-encoded proteins given that mitochondrial DNA only encodes for 13 proteins, which are incorporated into the respiratory chain. While mitochondria differ in morphology and functions among tissues in vivo, mitochondrial diversity among cell types is less well understood – especially in heterogeneous tissues such as the nervous system. Here, I present an in vivo tool for the characterization of cell type-specific mitochondria in mouse. Via the MitoTag mouse model, mitochondria from the cell type of interest are tagged in a Cre recombinase-dependent manner with GFP, which is localized to the outer mitochondrial membrane (GFP-OMM). This tagging allows for the immunocapture of organelles and their subsequent investigation through functional assays and omics-based screenings. We applied the MitoTag approach to the cerebellum and profiled the mitochondrial proteome of Purkinje cells, granule cells and astrocytes. Among these cell types, we found 196 proteins differentially enriched, of which 19 candidates were independently confirmed as cell type-enriched mitochondrial ‘markers’. Further analysis revealed functional specializations that we corroborated in independent assays using immunocaptured mitochondria. Specifically, astrocytic mitochondria superiorly oxidized long-chain fatty acids, while neuronal mitochondria demonstrated enhanced Ca2+ uptake via the mitochondrial calcium uniporter in granule cells and enhanced contact sites with the endoplasmic reticulum via regulator of microtubule dynamics protein 3 in Purkinje cells. In studies across species, I confirmed that neural mitochondrial diversity is conserved in the nervous system of mammals, aves and amphibian. Hence, we used neuronal and astrocytic mitochondrial ‘markers’ to show mitochondrial pathology in mouse models and human cases of Alzheimer’s disease and amyotrophic lateral sclerosis. The MitoTag approach enables mitochondrial research in a defined cellular context in vivo. Future applications will reveal the cell type-specific fine-tuning of mitochondria in many contexts, such as development, aging and diseases, as well as their contribution to the selective vulnerability of certain cell types.