Detecting RNA–RNA Interactions Using Psoralen Derivatives

Abstract

Psoralens are tricyclic compounds that intercalate into double-stranded DNA or RNA and, on irradiation with long-wavelength (365-nm) UV light, covalently link pyrimidines on adjacent strands. More rarely, psoralen cross-links can be observed at the ends of helices (i.e., double-stranded–single-stranded boundaries). Although psoralens can, in some instances, cross-link protein to RNA, their primary application is to detect RNA–RNA base-pairing interactions. The most useful psoralen derivative is 4′-aminomethyl trioxsalen (AMT), which is soluble in H2O. This protocol describes the use of AMT to detect RNA–RNA interactions in tissue culture cells or in extracts. Cross-linked RNAs are detectable by their reduced mobility in polyacrylamide gels. Cross-links can be reversed by exposure to short-wavelength (254 nm) UV light.