Painting Cell–Cell Interactions by Horseradish Peroxidase and Endogenously Generated Hydrogen Peroxide

Cell-cell interactions are fundamental in biology for maintaining physiological conditions with direct contact being the most straightforward mode of interaction. Recent advancements have led to the development of various chemical tools for detecting or identifying these interactions. However, the use of exogenous cues, such as toxic reagents, bulky probes, and light irradiation, can disrupt normal cell physiology. For example, the toxicity of hydrogen peroxide (H₂O₂) limits the applications of peroxidases in the proximity labeling field. In this study, we aimed to address this limitation by demonstrating that membrane-localized horseradish peroxidase (HRP-TM) efficiently utilizes endogenously generated extracellular H₂O₂. By harnessing endogenous H₂O₂, we observed that HRP-TM-expressing cells can effectively label contacting cells without the need for exogenous H₂O₂ treatment. Furthermore, we confirmed that HRP-TM labels proximal cells in an interaction-dependent manner. These findings offer a novel approach for studying cell-cell interactions under more physiological conditions without the confounding effects of exogenous stimuli. Our study contributes to elucidating cell-cell interaction networks in various model organisms, providing valuable insights into the dynamic interplay between cells in their native network.