Mapping Mitochondrial Diversity in the Aging Brain and Immune System

The brain and immune system are fueled by energy flow through mitochondria – dynamic, multifunctional organelles that convert energy and act as intracellular signaling hubs within neural, glial, and immune cells. Converging lines of research indicate that mitochondrial alterations likely arise with aging and play a causal role in the pathogenesis of Alzheimer’s disease (AD). However, progress in testing this hypothesis with a sufficient level of rigor and specificity has been hindered by the lack of approaches to address the diversity of mitochondrial phenotypes among different tissues and cell types. To profile multiple AD-relevant aspects of mitochondrial biology at a sufficient level of biological specificity, we have developed a simple computational approach that groups all known mitochondrial genes into functional pathways, measures their relative expression using omics data (RNA sequencing), and yields a fine-grained, interpretable picture of mitochondrial phenotypes – termed mitotypes. In this proposal, we deploy our mitotyping approach at single-cell resolution in two organ systems implicated in AD pathogenesis: the brain and immune system. In Aim 1, we propose to generate sensitive mitotype profiles for >1.4M brain cortical cells (neurons, glia, other cell types) across 381 older adults. This will establish cell type-specific mitochondrial recalibrations associated with age, among women and men, and allow the discovery of specific mitotype signatures associated with cognitive reserve and AD. In Aim 2, we deploy the same approach in >0.9M circulating immune cells from 175 adults ages 20-80, establishing sex-specific immune mitotypes associated with age, cognitive reserve, and AD. Computationally, brain and immune mitotypes are entirely compatible, therefore allowing us to harmonize datasets from both cohorts to identify potential shared abnormal mitochondrial features across the brain and immune cells in persons with cognitive impairment, dementia, and/or AD. Mechanistically, among the various domains of mitochondrial biology, previous research has specifically, but not exclusively, implicated defects in the mitochondrial oxidative phosphorylation (OxPhos) in AD pathogenesis. Therefore, in Aim 3 we use i) our Cellular Lifespan system to dissect the effects of targeted genetic and pharmacological OxPhos perturbations on mitotype signatures in living human cells over time, ii) a co-culture system to examine how a mitochondrial OxPhos impairment in one cell population influences the mitotype of another cell population, revealing new principles of mitotype crosstalk or “contagion” between cells; and iii) a unique cohort of patients with genetically-defined, bioenergetically-profiled mitochondrial OxPhos defects to examine, in the human brain-body system, how OxPhos defects influence other domains of mitochondrial biology as reflected in circulating immune cell mitotypes. Finally, in Aim 4 we develop an online platform, MitotypeExplorer 2.0, which makes mitotyping accessible to the growing community of neuroscientists, immunologists, and other interdisciplinary investigators interested in uncovering the role of age-related mitochondrial impairments and recalibrations in human health and disease.