Complexity of subcellular metabolism: strategies for compartment-specific profiling

Subcellular compartmentalization provides cells with tremendous advantages for the operation of cellular metabolism. Spatial separation of metabolism generates microenvironments with distinct concentrations of metabolites and cofactors allowing the cell to execute otherwise thermodynamically exclusive reactions simultaneously. Moreover, compartmentalization is also involved in the fine-tuned regulation of gene expression. Thus, the elucidation of compartment-specific metabolic processes has become essential for a thorough understanding of cellular metabolism. The limited availability of reliable methods to inspect metabolic activities in subcellular compartments has been a major impediment in this field. During the last decade, progress has been greatly extended through methodological advances in subcellular metabolic profiling. This review summarizes recent strategies applied for the direct and indirect assessment of compartment-specific metabolism. We review contemporary techniques, recent advances, limitations, and future trends in the field of subcellular metabolomics. •Assessment of subcellular metabolism is moving beyond traditional fractionation approaches with new immunocapture techniques.•Compartmentalized metabolism can be effectively probed through targeted genetic models and selective tracers.•Establishment of a compartmentalized flux model with integrated thermodynamic analysis is already feasible.