In situ imaging of bacterial outer membrane projections and associated protein complexes using electron cryo-tomography

Mohammed Kaplan, Georges Chreifi, Lauren Ann Metskas, Janine Liedtke, Cecily R. Wood, Catherine M. Oikonomou, William J. Nicolas, Poorna Subramanian, Lori A. Zacharoff, Yuhang Wang, Yi Wei Chang, Morgan Beeby, Megan J. Dobro, Yongtao Zhu, Mark J. McBride, Ariane Briegel, Carrie L. Shaffer, Grant J. Jensen

Research output: Contribution to journalArticlepeer-review

5 Scopus citations


The ability to produce outer membrane projections in the form of tubular membrane extensions (MEs) and membrane vesicles (MVs) is a widespread phenomenon among diderm bacteria. Despite this, our knowledge of the ultrastructure of these extensions and their associated protein complexes remains limited. Here, we surveyed the ultrastructure and formation of MEs and MVs, and their associated protein complexes, in tens of thousands of electron cryo-tomograms of ~90 bacterial species that we have collected for various projects over the past 15 years (Jensen lab database), in addition to data generated in the Briegel lab. We identified outer MEs and MVs in 13 diderm bacterial species and classified several major ultrastructures: (1) tubes with a uniform diameter (with or without an internal scaffold), (2) tubes with irregular diameter, (3) tubes with a vesicular dilation at their tip, (4) pearling tubes, (5) connected chains of vesicles (with or without neck-like connectors), (6) budding vesicles and nanopods. We also identified several protein complexes associated with these MEs and MVs which were distributed either randomly or exclusively at the tip. These complexes include a secretin-like structure and a novel crown-shaped structure observed primarily in vesicles from lysed cells. In total, this work helps to characterize the diversity of bacterial membrane projections and lays the groundwork for future research in this field.

Original languageEnglish
Article numbere73099
StatePublished - Sep 2021

Bibliographical note

Funding Information:
This project was funded by the NIH (grant R35 GM122588 to GJJ, and P20 GM130456 to CLS) and a Baxter postdoctoral fellowship from Caltech to MK. Cryo-ET work was done in the Beckman Institute Resource Center for Transmission Electron Microscopy at the California Institute of Technology. We are grateful to Prof. Martin Pilhofer for collecting the P. luteoviolacea data and for critically reading the manuscript. We thank Prof. Elitza I Tocheva for collecting the D. acidovorans data. We thank Prof. Mohamed El-Naggar for insights into preparing S. oneidensis samples and Dr. Yuxi Liu for discussions. Briegel lab data was collected at the Netherlands Center for Electron Nanoscopy with support from Dr Wen Yang. This data was collected with support from the National Roadmap for Large-Scale Research Infrastructure 2017–2018 with project number 184.034.014, which is financed in part by the Dutch Research Council (NWO). This work was also supported by the NWO OCENW. GROOT.2019.063 grant.

Publisher Copyright:
© Kaplan et al.

ASJC Scopus subject areas

  • Neuroscience (all)
  • Biochemistry, Genetics and Molecular Biology (all)
  • Immunology and Microbiology (all)


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