4-Thiouracil

4-Thiouracil (CAS No.: 591-28-6) is a sulfur-substituted nucleobase analog belonging to the class of thionucleobases, widely utilized in RNA biology and photochemical research as a site-specific, photoactivatable probe. It exhibits cytostatic properties and can be metabolically incorporated into RNA through enzymatic conversion to thiolated nucleotides, enabling selective labeling of newly synthesized transcripts. Upon irradiation with ultraviolet A light above 300 nm, 4-thiouracil acts as an efficient photosensitizer, undergoing triplet-state excitation and transferring energy to molecular oxygen to generate reactive oxygen species, particularly singlet oxygen. This photochemical reactivity underlies its utility in probing RNA structure, mapping nucleic acid–nucleic acid interactions, and inducing controlled photocrosslinking or photodamage in vitro and in cellular systems. The resulting oxidative transformations can produce fluorescent derivatives, facilitating detection and analysis. Although precise molecular targets are not defined in the classical receptor-based sense, its mechanism is closely linked to RNA incorporation and photoinduced oxidative processes. In vitro studies typically employ concentrations in the low micromolar range, with experimental conditions optimized according to cell type, irradiation parameters, and assay design. 4-Thiouracil is broadly applied in studies of RNA dynamics, transcriptome labeling, and photodynamic effects, and it continues to serve as a valuable tool in nucleic acid research and the exploration of photoactivated therapeutic strategies.