Always on the move: Mapping spatiotemporal interactome with proximity labeling in mammalian cells

Compartmentalization is central to coordinate complex biochemical processes in eukaryotic cells. Proteins governing organelle function and proteome complexity are dynamically trafficked intracellularly and intercellularly to maintain cellular homeostasis and communication. Proximity labeling (PL) enzymes such as TurboID and APEX2 coupled with mass spectrometry emerged as powerful tools to capture snapshots of subcellular proteomes. However, as protein trafficking is a dynamic process, this presents a challenge to current technologies due to limitations in temporal resolution. In this review, we highlight the recent developments in PL-based methodologies, including orthogonal PL enzymes (TransitID and OrthoID) and protein synchronization strategies coupled with PL (POTATOMap). Together, these new approaches enable the study of dynamic intracellular protein trafficking between organelles and intercellular communications.