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A live cell reporter of exosome secretion and uptake reveals pathfinding behavior of migrating cells

Author

Listed:
  • Bong Hwan Sung

    (Vanderbilt University School of Medicine)

  • Ariana Lersner

    (Vanderbilt University School of Medicine)

  • Jorge Guerrero

    (University of Wisconsin-Madison)

  • Evan S. Krystofiak

    (Vanderbilt University Cell Imaging Shared Resource)

  • David Inman

    (University of Wisconsin-Madison)

  • Roxanne Pelletier

    (Vanderbilt University School of Medicine)

  • Andries Zijlstra

    (Vanderbilt University School of Medicine
    Vanderbilt University Medical Center)

  • Suzanne M. Ponik

    (University of Wisconsin-Madison)

  • Alissa M. Weaver

    (Vanderbilt University School of Medicine
    Vanderbilt University School of Medicine
    Vanderbilt University Medical Center)

Abstract

Small extracellular vesicles called exosomes affect multiple autocrine and paracrine cellular phenotypes. Understanding the function of exosomes requires a variety of tools, including live imaging. Our previous live-cell reporter, pHluorin-CD63, allows dynamic subcellular monitoring of exosome secretion in migrating and spreading cells. However, dim fluorescence and the inability to make stably-expressing cell lines limit its use. We incorporated a stabilizing mutation in the pHluorin moiety, M153R, which now exhibits higher, stable expression in cells and superior monitoring of exosome secretion. Using this improved construct, we visualize secreted exosomes in 3D culture and in vivo and identify a role for exosomes in promoting leader–follower behavior in 2D and 3D migration. Incorporating an additional non-pH-sensitive red fluorescent tag allows visualization of the exosome lifecycle, including multivesicular body (MVB) trafficking, MVB fusion, exosome uptake and endosome acidification. This reporter will be a useful tool for understanding both autocrine and paracrine roles of exosomes.

Suggested Citation

  • Bong Hwan Sung & Ariana Lersner & Jorge Guerrero & Evan S. Krystofiak & David Inman & Roxanne Pelletier & Andries Zijlstra & Suzanne M. Ponik & Alissa M. Weaver, 2020. "A live cell reporter of exosome secretion and uptake reveals pathfinding behavior of migrating cells," Nature Communications, Nature, vol. 11(1), pages 1-15, December.
    • Handle: RePEc:nat:natcom:v:11:y:2020:i:1:d:10.1038_s41467-020-15747-2
    DOI: 10.1038/s41467-020-15747-2
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