Dissecting stimulus-specific Ca²⁺signals in amyloplasts and chloroplasts ofArabidopsis thalianacell suspension cultures

Abstract

Highlight Arabidopsis suspension-cultured cells stably expressing aequorin in the plastid stroma were used as a suitable tool to dissect differential Ca2+ responses of non-green plastids versus chloroplasts to several environmental cues.

Chloroplast-Specific in Vivo Ca2+ Imaging Using Yellow Cameleon Fluorescent Protein Sensors Reveals Organelle-Autonomous Ca2+ Signatures in the Stroma

“…2, B, C, and E), which are likely to reflect the differential Ca 2+ binding affinities of both sensors. Hence, our observations empirically identify YC3.6 as the sensor with the preferable characteristics to monitor physiological Ca 2+ dynamics in the plastid stroma, consistent with previous aequorin-based estimations of the free Ca 2+ concentrations in the stroma in the high nanomolar to low micromolar range (Johnson et al, 1995;Sello et al, 2016).…”

“…2, B and F). This mirrors recent aequorin-based observations where different stimuli (NaCl, mannitol, cold shock, and oligogalacturonides) applied to cultured Arabidopsis cells induced stromal Ca 2+ elevations that were lower than those in the cytosol (Sello et al, 2016). Both stromal Cameleon probes indicated the maximum ratio change at about 140 s after the eATP stimulus (150 6 13.5 s and 136 6 19.9 s for 2Bam-YC3.6 and 2Bam4-YC4.6 lines respectively; Fig.…”

“…We demonstrate the functionality of the sensing approach in vivo in plastids of roots and leaves and show that at tissue level the measured stromal Ca 2+ signatures resemble those reported using aequorin (Sello et al, 2016 …”

“…In vivo monitoring of stromal free Ca 2+ concentration ([Ca 2+ ] str ) using the aequorin probe subsequently enabled the observation of diurnal fluctuations in [Ca 2+ ] str between the high nanomolar and the low micromolar range (Johnson et al, 1995;Sai and Johnson, 2002;Nomura et al, 2012). The transition from light to darkness, in particular, was shown to induce stromal Ca 2+ accumulation (Johnson et al, 1995;Sai and Johnson, 2002;Sello et al, 2016). Consequently it has been hypothesized that chloroplasts sequester Ca 2+ into the thylakoids during the day and release it into the stroma at onset of darkness (Sai and Johnson, 2002).…”

“…Despite quantitative estimation and measurement of relative changes in free Ca 2+ concentrations in different chloroplast subcompartments using aequorin Sello et al, 2016), the extent and direction of intraplastidial Ca 2+ fluxes or Ca 2+ transport into and from the cytosol are currently not understood and none of the transporters involved has been identified (Rocha and Vothknecht, 2012;Finazzi et al, 2015). Accordingly, it remains unknown how stromal Ca 2+ dynamics and signaling are orchestrated.…”