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Cotranslational Folding Stimulates Programmed Ribosomal Frameshifting in the Alphavirus Structural Polyprotein

Harrington, Haley R. and Zimmer, Matthew H. and Chamness, Laura M. and Nash, Veronica and Penn, Wesley D. and Miller, Thomas F., III and Mukhopadhyay, Suchetana and Schlebach, Jonathan P. (2020) Cotranslational Folding Stimulates Programmed Ribosomal Frameshifting in the Alphavirus Structural Polyprotein. Journal of Biological Chemistry, 295 (20). pp. 6798-6808. ISSN 0021-9258. PMCID PMC7242702. doi:10.1074/jbc.RA120.012706.

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Viruses maximize their genetic coding capacity through a variety of biochemical mechanisms, including programmed ribosomal frameshifting (PRF), which facilitates the production of multiple proteins from a single mRNA transcript. PRF is typically stimulated by structural elements within the mRNA that generate mechanical tension between the transcript and ribosome. However, in this work, we show that the forces generated by the cotranslational folding of the nascent polypeptide chain can also enhance PRF. Using an array of biochemical, cellular, and computational techniques, we first demonstrate that the Sindbis virus structural polyprotein forms two competing topological isomers during its biosynthesis at the ribosome-translocon complex. We then show that the formation of one of these topological isomers is linked to PRF. Coarse-grained molecular dynamics simulations reveal that the translocon-mediated membrane integration of a transmembrane domain upstream from the ribosomal slip site generates a force on the nascent polypeptide chain that scales with observed frameshifting. Together, our results indicate that cotranslational folding of this viral protein generates a tension that stimulates PRF. To our knowledge, this constitutes the first example in which the conformational state of the nascent polypeptide chain has been linked to PRF. These findings raise the possibility that, in addition to RNA-mediated translational recoding, a variety of cotranslational folding or binding events may also stimulate PRF.

Item Type:Article
Related URLs:
URLURL TypeDescription Information Paper CentralArticle
Chamness, Laura M.0000-0002-3971-2799
Penn, Wesley D.0000-0001-8930-4091
Miller, Thomas F., III0000-0002-1882-5380
Mukhopadhyay, Suchetana0000-0001-9230-4746
Schlebach, Jonathan P.0000-0003-0955-7633
Additional Information:© 2020 Harrington et al. Published under exclusive license by The American Society for Biochemistry and Molecular Biology, Inc. Received for publication, January 17, 2020, and in revised form, March 4, 2020 Published, Papers in Press, March 13, 2020. We thank Renuka Kudva, Gunnar von Heijne, and other members of the Schlebach and Mukhopadhyay laboratories for scientific input. We thank David Giedroc and Jonathan Dinman for editorial input. We thank Christiane Hassel and the Indiana University Bloomington Flow Cytometry Core Facility for experimental support. This work used the Extreme Science and Engineering Discovery Environment (XSEDE) Bridges computer at the Pittsburgh Supercomputing Center through allocation TG-MCB160013 (55). This research was supported in part by NIAID, National Institutes of Health, Grant R21AI142383 (to J. P. S. and S. M.) as well as NIGMS, National Institutes of Health, Grant R01GM125063 (to T. F. M.). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. This article was selected as one of our Editors' Picks. Data availability: The cellular and biochemical data sets described herein will be made freely available by Jonathan Schlebach (Indiana University Department of Chemistry, upon request. All computational data and code relating to the CGMD simulations detailed herein will be made freely available by Thomas Miller III (California Institute of Technology Division of Chemistry and Chemical Engineering, upon request. All remaining data are contained within the article. Viral sequences analyzed herein can be freely accessed through the National Center for Biotechnology Information (NCBI) using the following accession numbers: Sindbis virus (NC_001547), eastern equine encephalitis virus (NC_003899), Middleburg virus (EF536323), sleeping disease virus (NC_003433), southern elephant seal virus (HM147990), Semliki Forest virus (NC_003215), and Venezuelan equine encephalitis virus (NC_001449). Author contributions: H. R. H., M. H. Z., V. N., W. D. P., T. F. M., S. M., and J. P. S. conceptualization; H. R. H., M. H. Z., L. M. C., V. N., W. D. P., T. F. M., S. M., and J. P. S. data curation; H. R. H., M. H. Z., V. N., W. D. P., S. M., and J. P. S. formal analysis; H. R. H., M. H. Z., L. M. C., V. N., W. D. P., and J. P. S. investigation; H. R. H., M. H. Z., and J. P. S. visualization; H. R. H., M. H. Z., L. M. C., V. N., W. D. P., and J. P. S. methodology; H. R. H., M. H. Z., L. M. C., W. D. P., T. F. M., and S. M. writing-review and editing; M. H. Z. and T. F. M. software; M. H. Z. validation; W. D. P., T. F. M., S. M., and J. P. S. supervision; W. D. P., T. F. M., S. M., and J. P. S. project administration; T. F. M. and J. P. S. resources; T. F. M., S. M., and J. P. S. funding acquisition; J. P. S. writing-original draft.
Funding AgencyGrant Number
National Institute of Allergy and Infectious DiseasesUNSPECIFIED
Subject Keywords:protein folding; membrane protein; transmembrane domain; translocation; translation control; ribosome; alphavirus; programmed ribosomal frameshifting; topology; Sindbis virus; translocon; membrane protein folding; cotranslational folding
Issue or Number:20
PubMed Central ID:PMC7242702
Record Number:CaltechAUTHORS:20191003-111416580
Persistent URL:
Official Citation:Haley R. Harrington, Matthew H. Zimmer, Laura M. Chamness, Veronica Nash, Wesley D. Penn, Thomas F. Miller III, Suchetana Mukhopadhyay, and Jonathan P. Schlebach. Cotranslational folding stimulates programmed ribosomal frameshifting in the alphavirus structural polyprotein. J. Biol. Chem. 2020 295: 6798-. doi:10.1074/jbc.RA120.012706
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:99046
Deposited By: Tony Diaz
Deposited On:03 Oct 2019 18:37
Last Modified:15 Feb 2022 21:14

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