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Published June 13, 2022 | Supplemental Material + Published
Journal Article Open

Ribosome profiling reveals multiple roles of SecA in cotranslational protein export


SecA, an ATPase known to posttranslationally translocate secretory proteins across the bacterial plasma membrane, also binds ribosomes, but the role of SecA's ribosome interaction has been unclear. Here, we used a combination of ribosome profiling methods to investigate the cotranslational actions of SecA. Our data reveal the widespread accumulation of large periplasmic loops of inner membrane proteins in the cytoplasm during their cotranslational translocation, which are specifically recognized and resolved by SecA in coordination with the proton motive force (PMF). Furthermore, SecA associates with 25% of secretory proteins with highly hydrophobic signal sequences at an early stage of translation and mediates their cotranslational transport. In contrast, the chaperone trigger factor (TF) delays SecA engagement on secretory proteins with weakly hydrophobic signal sequences, thus enforcing a posttranslational mode of their translocation. Our results elucidate the principles of SecA-driven cotranslational protein translocation and reveal a hierarchical network of protein export pathways in bacteria.

Additional Information

© The Author(s) 2022. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. Received 13 January 2022. Accepted 26 May 2022. Published 13 June 2022. We thank A. Buskirk, J. Chartron and J. Chen for advice on the ribosome profiling protocol, P. He for advice on data analysis, and members of the Shan lab for discussions and advice. Sequencing was performed at the Millard and Muriel Jacobs Genetics and Genomics Laboratory at California Institute of Technology. This work was supported by NIH grant R35 GM136321 to S.-o.S. These authors contributed equally: Zikun Zhu, Shuai Wang. Contributions. Conceptualization, Z.Z., S.W., and S.-o.S.; Methodology, Z.Z. and S.W.; Investigation, Z.Z. and S.W.; Formal Analysis, Z.Z.; Data curation, Z.Z.; Visualization, Z.Z.; Software, Z.Z. and S.W.; Resources, Z.Z. and S.W.; Writing – Original Draft, Z.Z. and S.-o.S.; Writing – Review & Editing, Z.Z., S.W., and S.-o.S.; Supervision, S.-o.S.; Funding Acquisition, S.-o.S. Reporting summary. Further information on research design is available in the Nature Research Reporting Summary linked to this article. Data availability. The data supporting the findings of this study are available from the corresponding authors upon reasonable request. The accession number for the data reported in this paper is GSE185572. The protein structures used to calculate absolute contact order were downloaded from AlphaFold Protein Structure Database (https://alphafold.ebi.ac.uk/). Source data for the figures and supplementary figures are provided as a Source Data file. Source data are provided with this paper. The authors declare no competing interests. Peer review information. Nature Communications thanks the anonymous reviewers for their contribution to the peer review of this work.

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August 22, 2023
October 24, 2023