A DEAD-box RNA helicase promotes thermodynamic equilibration of kinetically trapped RNA structures in vivo.
RNAs must assemble into specific structures in order to carry out their biological functions, but in vitro RNA folding reactions produce multiple misfolded structures that fail to exchange with functional structures on biological time scales. We used carefully designed self-cleaving mRNAs that assemble through well-defined folding pathways to identify factors that differentiate intracellular and in vitro folding reactions. Our previous work showed that simple base-paired RNA helices form and dissociate with the same rate and equilibrium constants in vivo and in vitro. However, exchange between adjacent secondary structures occurs much faster in vivo, enabling RNAs to quickly adopt structures with the lowest free energy. We have now used this approach to probe the effects of an extensively characterized DEAD-box RNA helicase, Mss116p, on a series of well-defined RNA folding steps in yeast. Mss116p overexpression had no detectable effect on helix formation or dissociation kinetics or on the stability of interdomain tertiary interactions, consistent with previous evidence that intracellular factors do not affect these folding parameters. However, Mss116p overexpression did accelerate exchange between adjacent helices. The nonprocessive nature of RNA duplex unwinding by DEAD-box RNA helicases is consistent with a branch migration mechanism in which Mss116p lowers barriers to exchange between otherwise stable helices by the melting and annealing of one or two base pairs at interhelical junctions. These results suggest that the helicase activity of DEAD-box proteins like Mss116p distinguish intracellular RNA folding pathways from nonproductive RNA folding reactions in vitro and allow RNA structures to overcome kinetic barriers to thermodynamic equilibration in vivo.
['Ruminski|Dana J|DJ|', 'Watson|Peter Y|PY|', 'Mahen|Elisabeth M|EM|', 'Fedor|Martha J|MJ|']
D001483:Base Sequence D053487:DEAD-box RNA Helicases D007700:Kinetics D009690:Nucleic Acid Conformation D012331:RNA, Fungal D012441:Saccharomyces cerevisiae D029701:Saccharomyces cerevisiae Proteins D013816:Thermodynamics
RNA RNA structure RNA folding Mss116p DEAD-box protein RNA chaperone RNA-protein interaction catalytic RNA ribozyme