Structural and molecular basis for Cardiovirus 2A as a viral gene switch

Published: 1 November 2021| Version 3 | DOI: 10.17632/gkpwngy65h.3
Contributors:
Chris Hill,
,
,
,
,
,
Neva Caliskan,

Description

Programmed ribosome frameshifting in cardioviruses is activated by the 2A protein: a multi-functional virulence factor that also inhibits cap-dependent translational initiation. Here, we show that 2A binds directly to the frameshift-stimulatory element in the viral RNA with nanomolar affinity and equimolar stoichiometry, and we define the minimal RNA element required for binding. Through site-directed mutagenesis and the use of single-molecule optical tweezers, we study the dynamics of EMCV RNA (wild type- CCC as well CUC mutant), both alone and in the presence of 2A. By observing short-lived intermediate states in real-time, we demonstrate that the EMCV stimulatory element exists in at least two conformations and binding of 2A stabilizes one of these, an RNA pseudoknot, increasing the force required to unwind it.

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Institutions

Helmholtz-Zentrum fur Infektionsforschung GmbH, University of Cambridge, Helmholtz-Institut fur RNA-basierte Infektionsforschung

Categories

Electrophoretic Mobility-Shift Assay, Thermophoresis, Optical Tweezer

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