Engineering a Novel NIR RNA-Specific Probe for Tracking Stress Granule Dynamics in Living Cells

Real-time monitoring of the dynamics of cytosolic RNA-protein condensates, termed stress granules (SGs), is vital for understanding their biological roles in stress response and related disease treatment but is challenging due to the lack of simple and accurate methods. Compared with protein visualization that requires complex transfection procedures, direct RNA labeling offers an ideal alternative for tracking SG dynamics in living cells. Here, we propose a novel molecular design strategy to construct a near-infrared RNA-specific fluorescent probe (HQBT) for tracking SGs in living cells. The positively charged probe HQBT was designed to target the negative groove of RNA, and its binding affinity to RNA was significantly improved by adjusting the position of the nitrogen atom in the molecule. Furthermore, an additional hydroxyl group was introduced to achieve near-infrared emission and enhance the RNA-binding capability. HQBT can rapidly stain RNA within 5 s in living cells and showed performance superior to the commercial SYTO RNA-Select dye in terms of photostability and selectivity. Importantly, the reversible assembly and disassembly dynamics of SGs are successfully visualized in living cells using this simple and direct RNA-selective imaging probe.