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In situ imaging of the bacterial flagellar motor disassembly and assembly processes

Kaplan, Mohammed and Subramanian, Poorna and Ghosal, Debnath and Oikonomou, Catherine M. and Pirbadian, Sahand and Starwalt‐Lee, Ruth and Mageswaran, Shrawan Kumar and Ortega, Davi R. and Gralnick, Jeffrey A. and El‐Naggar, Mohamed Y. and Jensen, Grant J. (2019) In situ imaging of the bacterial flagellar motor disassembly and assembly processes. EMBO Journal, 38 (10). Art. No. e100957. ISSN 0261-4189. PMCID PMC6627242. https://resolver.caltech.edu/CaltechAUTHORS:20190520-104026571

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Abstract

The self‐assembly of cellular macromolecular machines such as the bacterial flagellar motor requires the spatio‐temporal synchronization of gene expression with proper protein localization and association of dozens of protein components. In Salmonella and Escherichia coli, a sequential, outward assembly mechanism has been proposed for the flagellar motor starting from the inner membrane, with the addition of each new component stabilizing the previous one. However, very little is known about flagellar disassembly. Here, using electron cryo‐tomography and sub‐tomogram averaging of intact Legionella pneumophila, Pseudomonas aeruginosa, and Shewanella oneidensis cells, we study flagellar motor disassembly and assembly in situ. We first show that motor disassembly results in stable outer membrane‐embedded sub‐complexes. These sub‐complexes consist of the periplasmic embellished P‐ and L‐rings, and bend the membrane inward while it remains apparently sealed. Additionally, we also observe various intermediates of the assembly process including an inner‐membrane sub‐complex consisting of the C‐ring, MS‐ring, and export apparatus. Finally, we show that the L‐ring is responsible for reshaping the outer membrane, a crucial step in the flagellar assembly process.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.15252/embj.2018100957DOIArticle
http://www.ncbi.nlm.nih.gov/pmc/articles/pmc6627242/PubMed CentralArticle
ORCID:
AuthorORCID
Kaplan, Mohammed0000-0002-0759-0459
Ghosal, Debnath0000-0002-2227-0330
Oikonomou, Catherine M.0000-0003-2312-4746
Ortega, Davi R.0000-0002-8344-2335
Gralnick, Jeffrey A.0000-0001-9250-7770
Jensen, Grant J.0000-0003-1556-4864
Additional Information:© 2019 The Authors. Received 18 October 2018; Revised 11 April 2019; Accepted 18 April 2019. This work was supported by the National Institutes of Health (NIH, grant R01 AI127401 to G.J.J.). M.K. is supported by a postdoctoral Rubicon fellowship from De Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO). S.P. and M.Y.E.‐N. are supported by the Air Force Office of Scientific Research Presidential Early Career Award for Scientists and Engineers (FA955014‐1‐0294, to M.Y.E.‐N.). R.S.L. and J.A.G acknowledge support from the Office of Naval Research (N00014‐18‐1‐2632 to J.A.G.). We would like to thank Dr. Spiros Garbis, Dr. Annie Moradian, Dr. Michael Sweredoski, and Dr. Brett Lomenick from the Caltech Proteome Exploration Laboratory for their help in mass spectrometry experiments and data analysis. All P. aeruginosa mutants were kindly supplied by Dianne Newman's laboratory at Caltech. ECT was performed in the Beckman Institute Resource Center for Cryo‐EM. Funding: Air Force Office of Scientific Research Presidential Early Career Award for Scientists and Engineers FA955014‐1‐0294; National Institutes of Health R01 AI127401; De Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO).
Funders:
Funding AgencyGrant Number
NIHR01 AI127401
Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO)UNSPECIFIED
Air Force Office of Scientific Research (AFOSR)FA955014-1-0294
Office of Naval Research (ONR)N00014-18-1-2632
Subject Keywords:assembly; bacterial flagellar motor; disassembly; electron cryo-tomography; in situ imaging
Issue or Number:10
PubMed Central ID:PMC6627242
Record Number:CaltechAUTHORS:20190520-104026571
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20190520-104026571
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:95599
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:20 May 2019 18:22
Last Modified:10 Apr 2020 22:51

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