Mechanisms of Metabolic Adaptation: from Single Molecules to Systems Biology

Project Summary/Abstract Cells have evolved to sense external and internal chemical cues to maintain cellular homeostasis and ensure survival under challenging environmental conditions. In response to environmental stressors, such as the scarcity of nutrients or xenobiotics, cells efficiently adjust their metabolism, a process that is critical for their survival and adaptation. To rewire metabolism, cells have evolved sophisticated mechanisms that sense and regulate pathways in response to stressors. In recent decades, significant advances have been made in identifying metabolic pathways in cells responding to both internal and external chemical cues. Autophagy and cytochrome P450 metabolisms are two pathways that are activated under conditions of nutrient and chemical stress. A considerable challenge persists in understanding how these mechanisms respond during metabolic adaptation. This knowledge gap not only hinders the understanding of how cells maintain response flexibility across diverse conditions but also limits the ability to design targeted and effective treatments that depend on metabolic adaptation. Thus, unraveling these complexities is crucial for developing more precise and impactful strategies against a variety of chronic diseases. The PI has recently developed genomics and imaging approaches that enable the quantitative interrogation of metabolic adaptation in human cell models, and with these tools, proposes to address this knowledge gap. This approach pairs CRISPR-inspired genetic perturbations and genome engineering techniques with state-of-the-art single-molecule microscopy and live-cell imaging. For this proposal, the lab will synergize microscopy and genomics studies with mass spectrometry proteomics, determining the assembly of metabolic complexes and post-translational modifications critical for metabolism rewiring. This unique approach will allow systems biology study of metabolism – from single molecule to cell-wide molecular changes. In this proposal, the lab will determine how autophagy and cytochrome P450 metabolism pathways respond to metabolic adaptation. For autophagy, preliminary data pinpoint WIPI-mediated phospholipid sensing as a key regulatory factor in autophagy progression during nutrient stress. However, the role played by the multiple WIPI isoforms and how they differently concur in autophagy progression remains elusive. The lab will precisely define the molecular interplay between phospholipid signaling and WIPI isoforms in the context of autophagy activation under the condition of nutrient stress. For the cytochrome P450 side of the proposal, the lab will unravel the assembly and function of cytochrome P450 metabolic complexes upon xenobiotic stress in the living cell: this is one of the oldest, yet still unresolved, questions in the field. Not only results from this proposal will provide quantitative measurements on cytochrome P450s molecular assembly but also set a path for discovering novel protein interactors. Together, this proposal will serve as a roadmap for understanding metabolic adaptation in several chronic diseases.