mRNA stem-loops can pause the ribosome by hindering A-site tRNA binding

• Main Authors: Chen Bao, Sarah Loerch, Clarence Ling, Andrei A Korostelev, Nikolaus Grigorieff, Dmitri N Ermolenko
• Format: Article
• Language: English
• Published: eLife Sciences Publications Ltd 2020-05-01
• Series: eLife
• Subjects: ribosome; translation; mRNA; smFRET; cryo-EM
• Online Access: https://elifesciences.org/articles/55799

Although the elongating ribosome is an efficient helicase, certain mRNA stem-loop structures are known to impede ribosome movement along mRNA and stimulate programmed ribosome frameshifting via mechanisms that are not well understood. Using biochemical and single-molecule Förster resonance energy transfer (smFRET) experiments, we studied how frameshift-inducing stem-loops from E. coli dnaX mRNA and the gag-pol transcript of Human Immunodeficiency Virus (HIV) perturb translation elongation. We find that upon encountering the ribosome, the stem-loops strongly inhibit A-site tRNA binding and ribosome intersubunit rotation that accompanies translation elongation. Electron cryo-microscopy (cryo-EM) reveals that the HIV stem-loop docks into the A site of the ribosome. Our results suggest that mRNA stem-loops can transiently escape the ribosome helicase by binding to the A site. Thus, the stem-loops can modulate gene expression by sterically hindering tRNA binding and inhibiting translation elongation.