Intravital quantification reveals dynamic calcium concentration changes across B cell differentiation stages

Elife. 2021 Mar 22:10:e56020. doi: 10.7554/eLife.56020.

Abstract

Calcium is a universal second messenger present in all eukaryotic cells. The mobilization and storage of Ca2+ ions drives a number of signaling-related processes, stress-responses, or metabolic changes, all of which are relevant for the development of immune cells and their adaption to pathogens. Here, we introduce the Förster resonance energy transfer (FRET)-reporter mouse YellowCaB expressing the genetically encoded calcium indicator TN-XXL in B lymphocytes. Calcium-induced conformation change of TN-XXL results in FRET-donor quenching measurable by two-photon fluorescence lifetime imaging. For the first time, using our novel numerical analysis, we extract absolute cytoplasmic calcium concentrations in activated B cells during affinity maturation in vivo. We show that calcium in activated B cells is highly dynamic and that activation introduces a persistent calcium heterogeneity to the lineage. A characterization of absolute calcium concentrations present at any time within the cytosol is therefore of great value for the understanding of long-lived beneficial immune responses and detrimental autoimmunity.

Keywords: B cells; calcium; fluorescence lifetime imaging; germinal center; immunology; inflammation; mouse.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • B-Lymphocytes / metabolism*
  • Female
  • Fluorescence Resonance Energy Transfer / methods*
  • Lymphocyte Activation*
  • Male
  • Mice

Associated data

  • Dryad/10.5061/dryad.cc2fqz63d

Grants and funding

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