Gdt1p, a new player involved in calcium homeostasis in Saccharomyces cerevisiae? Development of organelles-localized calcium and pH-sensitive probes.

Deschamps, Antoine;Sluysmans, Sophie;Colinet, Anne-Sophie;Poncelet, Guillaume;Morsomme, Pierre
(2015) Gordon Research Conference - Organellar Channels and Transporters — Location: Bentley University in Waltham MA United States (14.June.2015)

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Authors
  • Deschamps, AntoineUCLouvain
    Author
  • Sluysmans, SophieUCLouvain
    Author
  • Colinet, Anne-SophieUCLouvain
    Author
  • Poncelet, GuillaumeUCLouvain
    Author
  • Morsomme, Pierreorcid-logoUCLouvain
    Author
Abstract
The Ca2+ ion is a ubiquitous intracellular messenger involved in various signaling processes. The basal concentration of cytosolic calcium is maintained at a very low level. Upon stimulation, a transient increase of cytosolic Ca2+ originated from internal and/or external stores is observed. Therefore, a complex toolkit of pumps, exchangers and channels localized in the different compartments of the cell are essential to tightly control this mechanism. Although a significant number of those are already well known, there are still some lacking actors in the Saccharomyces cerevisiae Ca2+-signaling toolkit. Recently, we have identified a novel family of putative Ca2+/H+ antiporters (UPF0016), whose Gdt1p is the yeast member. Gdt1p is a six-transmembrane spans protein located to the Golgi apparatus. It possesses a characteristic conserved hydrophilic motif in two transmembrane spans. Phenotypical assays and cytosolic Ca2+ concentration measurement suggested that Gdt1p could play an important role in Ca2+ homeostasis. However, the precise function of Gdt1p remains unclear. In order to elucidate the physiological function of Gdt1p related to calcium and pH homeostasis within the secretory pathway, we try to get direct evidences of Ca2+/H+ transport. For this purpose, we are addressing genetically-encoded Ca2+ sensors and H+ probes to the lumen of different organelles (endoplasmic reticulum, Golgi apparatus, vacuole). Aequorin, GAP (GFP-Aequorin Probe) and pHluorin are bioluminescent/fluorescent probes useful for Ca2+ or pH measurement, respectively. So far, we correctly localized these sensors to the endoplasmic reticulum and the Golgi apparatus lumens. However, the low expression level does not allow us to distinguish specific signal from noise. The next challenge is then to increase the expression level of these probes while preserving the right localization.
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Deschamps, A., Sluysmans, S., Colinet, A.-S., Poncelet, G., & Morsomme, P. (2015). Gdt1p, a new player involved in calcium homeostasis in Saccharomyces cerevisiae? Development of organelles-localized calcium and pH-sensitive probes. Gordon Research Conference - Organellar Channels and Transporters, Bentley University in Waltham MA United States. https://hdl.handle.net/2078.5/177545