Video3.AVI Audrey Kelner Nuno Leitão Mireille Chabaud Myriam Charpentier Fernanda de Carvalho-Niebel 10.3389/fpls.2018.00245.s004 https://frontiersin.figshare.com/articles/media/Video3_AVI/5928574 <p>Spatiotemporal changes in cellular calcium (Ca<sup>2+</sup>) concentrations are essential for signal transduction in a wide range of plant cellular processes. In legumes, nuclear and perinuclear-localized Ca<sup>2+</sup> oscillations have emerged as key signatures preceding downstream symbiotic signaling responses. Förster resonance energy transfer (FRET) yellow-based Ca<sup>2+</sup> cameleon probes have been successfully exploited to measure the spatiotemporal dynamics of symbiotic Ca<sup>2+</sup> signaling in legumes. Although providing cellular resolution, these sensors were restricted to measuring Ca<sup>2+</sup> changes in single subcellular compartments. In this study, we have explored the potential of single fluorescent protein-based Ca<sup>2+</sup> sensors, the GECOs, for multicolor and simultaneous imaging of the spatiotemporal dynamics of cytoplasmic and nuclear Ca<sup>2+</sup> signaling in root cells. Single and dual fluorescence nuclear and cytoplasmic-localized GECOs expressed in transgenic Medicago truncatula roots and Arabidopsis thaliana were used to successfully monitor Ca<sup>2+</sup> responses to microbial biotic and abiotic elicitors. In M. truncatula, we demonstrate that GECOs detect symbiosis-related Ca<sup>2+</sup> spiking variations with higher sensitivity than the yellow FRET-based sensors previously used. Additionally, in both M. truncatula and A. thaliana, the dual sensor is now able to resolve in a single root cell the coordinated spatiotemporal dynamics of nuclear and cytoplasmic Ca<sup>2+</sup> signaling in vivo. The GECO-based sensors presented here therefore represent powerful tools to monitor Ca<sup>2+</sup> signaling dynamics in vivo in response to different stimuli in multi-subcellular compartments of plant cells.</p> 2018-02-27 12:00:24 Medicago truncatula Arabidopsis thaliana root symbiosis root hairs root elongation zone calcium GECO sensors biotic and abiotic stimuli