@article{e94ef0c385d7498abece844439cf7683,
title = "CRISPR screening reveals a dependency on ribosome recycling for efficient SARS-CoV-2 programmed ribosomal frameshifting and viral replication",
abstract = "During translation of the genomic RNA of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative virus in the COVID-19 pandemic, host ribosomes undergo programmed ribosomal frameshifting (PRF) at a conserved structural element. Although PRF is essential for coronavirus replication, host factors that regulate this process have not yet been identified. Here we perform genome-wide CRISPR-Cas9 knockout screens to identify regulators of SARS-CoV-2 PRF. These screens reveal that loss of ribosome recycling factors markedly decreases frameshifting efficiency and impairs SARS-CoV-2 viral replication. Mutational studies support a model wherein efficient removal of ribosomal subunits at the ORF1a stop codon is required for frameshifting of trailing ribosomes. This dependency upon ribosome recycling is not observed with other non-pathogenic human betacoronaviruses and is likely due to the unique position of the ORF1a stop codon in the SARS clade of coronaviruses. These findings therefore uncover host factors that support efficient SARS-CoV-2 translation and replication.",
keywords = "ABCE1, CP: Microbiology, CP: Molecular biology, DENR, DOHH, EIF5A, SARS-CoV-2, coronavirus, diphthamide, host factors, programmed ribosomal frameshifting, ribosome recycling",
author = "Frederick Rehfeld and Eitson, {Jennifer L.} and Ohlson, {Maikke B.} and Chang, {Tsung Cheng} and Schoggins, {John W.} and Mendell, {Joshua T.}",
note = "Funding Information: We thank John Doench, David Root, Didier Trono, and Feng Zhang for plasmids; Vanessa Schmid and Jo Wagner in the McDermott Center Next Generation Sequencing Core for assistance with high-throughput sequencing; Angie Mobley and the UT Southwestern Flow Cytometry Core facility for assistance with FACS; and Kathryn O'Donnell and members of the Mendell laboratory for helpful feedback on the manuscript. This work was supported by grants from NIH (DP1AI158124 to J.W.S), CPRIT (RP220309 to J.T.M.), the Welch Foundation (I-1961-20210327 to J.T.M.), and the Hamon Center for Regenerative Science and Medicine at UT Southwestern (to J.T.M.). J.W.S. holds an Investigators in the Pathogenesis of Infectious Disease Award from the Burroughs Wellcome Fund. J.T.M. is an Investigator of the Howard Hughes Medical Institute. This article is subject to HHMI's Open Access to Publications policy. HHMI lab heads have previously granted a non-exclusive CC BY 4.0 license to the public and a sublicensable license to HHMI in their research articles. Pursuant to those licenses, the author-accepted manuscript of this article can be made freely available under a CC BY 4.0 license immediately upon publication. F.R. J.W.S. and J.T.M. designed the experiments and interpreted the results. F.R. J.L.E. M.B.O. and T.-C.C. performed experiments. F.R. and J.T.M wrote the manuscript. J.T.M. is a scientific advisor for Ribometrix, Inc. and owns equity in Orbital Therapeutics, Inc. We support inclusive, diverse, and equitable conduct of research. Funding Information: We thank John Doench, David Root, Didier Trono, and Feng Zhang for plasmids; Vanessa Schmid and Jo Wagner in the McDermott Center Next Generation Sequencing Core for assistance with high-throughput sequencing; Angie Mobley and the UT Southwestern Flow Cytometry Core facility for assistance with FACS; and Kathryn O{\textquoteright}Donnell and members of the Mendell laboratory for helpful feedback on the manuscript. This work was supported by grants from NIH ( DP1AI158124 to J.W.S), CPRIT ( RP220309 to J.T.M.), the Welch Foundation ( I-1961-20210327 to J.T.M.), and the Hamon Center for Regenerative Science and Medicine at UT Southwestern (to J.T.M.). J.W.S. holds an Investigators in the Pathogenesis of Infectious Disease Award from the Burroughs Wellcome Fund. J.T.M. is an Investigator of the Howard Hughes Medical Institute. This article is subject to HHMI{\textquoteright}s Open Access to Publications policy. HHMI lab heads have previously granted a non-exclusive CC BY 4.0 license to the public and a sublicensable license to HHMI in their research articles. Pursuant to those licenses, the author-accepted manuscript of this article can be made freely available under a CC BY 4.0 license immediately upon publication. Publisher Copyright: {\textcopyright} 2023 The Author(s)",
year = "2023",
month = feb,
day = "28",
doi = "10.1016/j.celrep.2023.112076",
language = "English (US)",
volume = "42",
journal = "Cell Reports",
issn = "2211-1247",
publisher = "Cell Press",
number = "2",
}