Unraveling Dynamic Molecular Networks through the Evolution of Proximity Labeling Technologies

Proximity labeling (PL) has become a powerful strategy for capturing biomolecular interactions in their native cellular environments. Over the past decade, this approach has evolved from a single enzyme based tool into a diverse technological platform, yielding specialized systems optimized for distinct biological applications. Here, we provide a systematic overview of PL technologies, tracing their development from early enzyme-based systems to new-generation tools including split-enzyme systems, activatable probes, and orthogonal dual enzyme strategies. We highlight how recent innovations have expanded applications from single target interactome mapping to dynamic, network level analyses of protein trafficking, intercellular communication, and multi-compartment signaling. We further explore the growing translational impact of PL in disease modeling, biomarker discovery, and therapeutic development. By synthesizing key technological milestones and emerging trends, this review provides a perspective on how next-generation PL platforms will reshape the study of molecular networks and advance precision medicine.