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T cell calcium dynamics visualized in a ratiometric tdTomato-GCaMP6f transgenic reporter mouse

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Cite this article as: eLife 2017;6:e32417 doi: 10.7554/eLife.32417

Abstract

Calcium is an essential cellular messenger that regulates numerous functions in living organisms. Here we describe development and characterization of 'Salsa6f', a fusion of GCaMP6f and tdTomato optimized for cell tracking while monitoring cytosolic Ca2+, and a transgenic Ca2+ reporter mouse with Salsa6f targeted to the Rosa26 locus for Cre-dependent expression in specific cell types. The development and function of T cells was unaffected in Cd4-Salsa6f mice. We describe Ca2+ signals reported by Salsa6f during T cell receptor activation in naïve T cells, helper Th17 T cells and regulatory T cells, and Ca2+ signals mediated in T cells by an activator of mechanosensitive Piezo1channels. Transgenic expression of Salsa6f enables ratiometric imaging of Ca2+ signals in complex tissue environments found in vivo. Two-photon imaging of migrating T cells in the steady-state lymph node revealed both cell-wide and localized sub-cellular Ca2+ transients ('sparkles') as cells migrate.

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Author details

  1. Tobias X Dong

    Department of Physiology and Biophysics, University of California, Irvine, Irvine, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5500-7099
  2. Shivashankar Othy

    Department of Physiology and Biophysics, University of California, Irvine, Irvine, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6832-5547
  3. Amit Jairaman

    Department of Physiology and Biophysics, University of California, Irvine, Irvine, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Jonathan Skupsky

    Department of Physiology and Biophysics, University of California, Irvine, Irvine, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Angel Zavala

    Department of Physiology and Biophysics, University of California, Irvine, Irvine, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Ian Parker

    Department of Physiology and Biophysics, University of California, Irvine, Irvine, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Joseph L Dynes

    Department of Physiology and Biophysics, University of California, Irvine, Irvine, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Michael D Cahalan

    Department of Physiology and Biophysics, University of California, Irvine, Irvine, United States
    For correspondence
    mcahalan@uci.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4987-2526

Funding

National Institutes of Health (AI117555)

  • Joseph L Dynes
  • Michael D Cahalan

National Institutes of Health (NS14609)

  • Michael D Cahalan

National Institutes of Health (AI121945)

  • Michael D Cahalan

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: Use of blood samples from healthy human subjects has been approved by the University of California, Irvine Institutional Review Board (UCI IRB HS #1995-459). All animal procedures were approved by the UCI Institutional Animal Care and Use committee (IACUC) (protocol #1998-1366-11).

Human subjects: Human blood was prepared using support from the National Center for Research Resources and the National Center for Advancing Translational Sciences (NIH Grant UL1 TR000153).

Reviewing Editor

  1. Michael L Dustin, University of Oxford, United Kingdom

Publication history

  1. Received: October 2, 2017
  2. Accepted: December 11, 2017
  3. Accepted Manuscript published: December 14, 2017 (version 1)
  4. Version of Record published: December 29, 2017 (version 2)

Copyright

© 2017, Dong et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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