Cellular Mechanisms of Mitochondrial-Organelle Crosstalk in Cancer Metabolism

Abstract

Mitochondria function as central hubs of cellular metabolism, integrating energy production via the tricarboxylic acid cycle and oxidative phosphorylation with diverse biochemical processes essential for cellular homeostasis. Beyond their metabolic roles, mitochondria-derived metabolites and signals have been shown to regulate fundamental cellular processes, including proliferation, signal transduction, cell fate, and cell death. Dysregulation of these functions is increasingly recognized as a contributor to various diseases.

To meet cellular metabolic demands, robust quality control mechanisms are essential for eliminating dysfunctional mitochondria. Mitophagy is an essential process that removes damaged or old mitochondria through an autophagic turnover via lysosomal degradation. My research uncovered that PINK1-driven mitophagy extends beyond quality control, also replenishing the cellular iron pool to maintain cellular energetics required for cancer growth. These findings underscore the importance of inter-compartmental regulation in maintaining subcellular metabolic balance, laying the foundation for subsequent investigations.

While organelle compartmentalization is critical for cellular metabolic homeostasis, the spatial and temporal organization of metabolic pathways remains largely unexplored. Endoplasmic reticulum (ER)-mitochondria contact sites (ERMCS) is one of the most abundant mitochondria-organelle interactions. Although ERMCS directly regulate metabolite transport, lipid transfer, and signaling, our understanding of their regulation and roles in cellular metabolism are unclear. We investigated the role of ERMCS in mitochondrial function using a machine learning-assisted imaging screen of FDA-approved compounds, identifying fedratinib as an ERMCS inducer. Fedratinib treatment activates a BRD4-dependent gene expression program, leading to mitochondrial structural alterations, including ER envelopment, cristae displacement, loss of membrane potential, and altered lipid and calcium levels in ER-associated mitochondria. By employing a fluorescent split-GFP ERMCS reporter and volumetric electron microscopy, we generated high-resolution 3D visualizations of ERMCS formation and its effects on mitochondrial ultrastructure. Proteomic analyses revealed enrichment of fatty acid oxidation and proline biosynthesis pathways in ER-associated mitochondria, establishing a mechanistic link between ERMCS and subcellular metabolic specialization. Using fedratinib as a potent ERMCS inducer, we further explored roles of ERMCS in cellular function. Stearoyl-CoA desaturase (SCD) is an endoplasmic reticulum-associated enzyme that catalyzes the synthesis of monounsaturated fatty acids. We demonstrated that SCD regulates ERMCS formation by modulating membrane fluidity. Inhibiting SCD reduced ERMCS, highlighting the importance of lipid metabolism in maintaining these structures. Additionally, we found an essential role for mitochondrial respiration through the redox-active lipid, ubiquinone, in modulating ERMCS. We also linked ERMCS to amino acid metabolism, particularly the lysine and proline pathways. A metabolic CRISPR screen identified key regulators of fedratinib sensitivity and resistance, revealing that ERMCS induction enhances sensitivity to cuproptosis. These data demonstrate how ERMCS reprogram cellular metabolism and identify potential therapeutic opportunities.

This dissertation examines the mechanisms governing inter-organellar metabolic communication, with a focus on mitochondrial interactions with other organelles and their implications for health and disease. It provides a comprehensive analysis of mitochondria-organelle communication, uncovering novel regulatory mechanisms and therapeutic targets. Despite the established link between organelle dysfunction and numerous diseases, the role of inter-organellar communication remains poorly understood. This work identifies an FDA-approved drug that significantly increases ERMCS, offering a valuable tool for studying the metabolic rewiring associated with ERMCS induction. The compound enables temporal and reversible modulation of ERMCS, laying the groundwork for developing therapeutics targeting inter-organellar communication in disease. Overall, this research highlights the critical role of organellar heterogeneity and communication in maintaining compartmentalized metabolic regulation.