Does cellular hydrogen peroxide diffuse or act locally?

Understanding of redox signaling requires data on the spatiotemporal distribution of hydrogen peroxide (H₂O₂) within the cell. The fluorescent reporter HyPer is a powerful instrument for H₂O ₂ imaging. However, rapid diffusion of HyPer throughout the nucleocytoplasmic compartment does not allow visualization of H ₂O₂ gradients on the micrometer scale. Here we dramatically improved the spatial resolution of H₂O₂ imaging by applying subcytoplasmic targeting of HyPer. The membrane-attached reporters identified "microdomains" of elevated H₂O ₂ levels within the cytoplasm of the cells exposed to growth factors. We demonstrate that diffusion of H₂O₂ across the cytoplasm was strongly limited, providing evidence that H₂O ₂ acts locally inside cells.