Unwinding Quadruplex Structures with RNA Helicases

The interaction of RNA helicases and non-canonical G-rich quadruplex (G4) structures in all aspects of RNA metabolism is a rapidly expanding field of study. Both cellular and viral RNA helicases dynamically maintain and disrupt G4 RNA structures in multiple compartments of the cell for efficient trafficking, transcription, splicing, translation, and, in the case of viral RNA genomes, replication. The conserved domains shared by diverse members of the helicase superfamilies for RNA binding and ATP-mediated duplex unwinding suggest that quadruplex disruption is a critical activity for many helicases. To probe the conditions and required energetics for quadruplex interaction and unwinding, robust in vitro assays are needed. To overcome the challenges of studying G4 unwinding under optimized conditions, we have adapted standard helicase unwinding assays to closely examine quadruplex dynamics. This chapter provides detailed protocols for describing the capacity for RNA helicases to remodel G4 structures.