Acc Chem Res. 2025 Jul 21.
ConspectusEngineered photosensitizer proteins, such as miniSOG, KillerRed, and SuperNova, have long been used for light-triggered protein inhibition and cell ablation. Compared to synthetic organic dyes, these genetically encoded tags provide superior spatial precision for subcellular targeting. More recently, the photochemistry of miniSOG has been repurposed for subcellular omics studies. Upon light activation, miniSOG generates reactive oxygen species (ROS) that oxidize nearby nucleic acids or proteins. These oxidized biomolecules can then react with exogenously supplied nucleophilic probes, which introduce bio-orthogonal handles for downstream enrichment and analysis.This labeling strategy, known as photocatalytic proximity labeling (PPL), has emerged as a powerful approach for profiling the molecular architecture of subcellular compartments and identifying RNA or protein interactors of specific targets. The use of light provides exceptional temporal control, enabling labeling windows as short as 1 s. Moreover, PPL readily supports pulse-chase experiments through simple light on/off switching, an advantage not easily achievable with conventional platforms such as APEX or TurboID.In this account, we highlight our recent developments and applications of genetically encoded PPL tools. These include CAP-seq for RNA/DNA labeling, RinID for protein labeling, and LAP-seq/MS/CELL for bioluminescence-activated multi-omic profiling. Together, these tools enable detailed mapping of the cellular biomolecular landscape. For example, CAP-seq revealed enrichment of transcripts encoding secretory and mitochondrial proteins near the endoplasmic reticulum membrane and outer mitochondrial membrane, supporting models of localized translation. Additionally, pulse-chase labeling using RinID in the ER lumen uncovered distinct decay kinetics of secretory proteins.Looking forward, future efforts may focus on developing low-toxicity and low-background chemical probes, engineering red-shifted photosensitizers for deep-tissue and in vivo applications, and integrating multiple proximity labeling (PL) platforms to study organelle contact sites and interorganelle molecular trafficking.