title: Fluorescent biosensors illuminate calcium levels within defined beta-cell endosome subpopulations.
abstract: Live cell imaging has revealed that calcium ions (Ca(2+)) pass in and out of many cellular organelles. However, technical hurdles have limited measurements of Ca(2+) in acidic organelles, such as endosomes. Although evidence hints that endosomes play a role in Ca(2+) signaling, direct measurements within endosomal lumina represent one of the final frontiers in organelle imaging. To measure Ca(2+) in a TiVAMP-positive endosome sub-population, the pH-resistant ratiometric Ca(2+) biosensor GEM-GECO1 and the ratiometric pH biosensor mKeima were used. A positive correlation between acidic endosomal pH and higher Ca(2+) was observed within these Rab5a- and Rab7-positive compartments. Ca(2+) concentration in most endosomes was estimated to be below 2μM, lower than Ca(2+) levels in several other intracellular stores, indicating that endosomes may take up Ca(2+) during physiological stimulation. Indeed, endosomes accumulated Ca(2+) during glucose-stimulation, a condition where endosomal pH did not change. Our biosensors permitted the first measurements revealing a role for endosomes in cellular Ca(2+) homeostasis during physiological stimulation.
authors: Department of Cellular and Physiological Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Tobias
authors: T
authors: Albrecht
authors: Department of Chemistry, University of Alberta, Edmonton, AB, Canada.
authors: Yongxin
authors: Y
authors: Zhao
authors: Department of Chemistry, University of Alberta, Edmonton, AB, Canada.
authors: Trang Hai
authors: TH
authors: Nguyen
authors: Department of Chemistry, University of Alberta, Edmonton, AB, Canada.
authors: Robert E
authors: RE
authors: Campbell
authors: Department of Cellular and Physiological Sciences, University of British Columbia, Vancouver, BC, Canada. Electronic address: James.D.Johnson@ubc.ca.
authors: James D
authors: JD
authors: Johnson
mesh_terms: D000818:Animals; D015374:Biosensing Techniques / Q000706:statistics & numerical data*; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D002460:Cell Line; D002648:Child; D011992:Endosomes / Q000378:metabolism* / Q000648:ultrastructure; D005456:Fluorescent Dyes / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D050417:Insulin-Secreting Cells / Q000648:ultrastructure*; D008297:Male; D051379:Mice; D061848:Optical Imaging / Q000379:methods*
chemical_list: D005456:Fluorescent Dyes; D005947:Glucose; D002118:Calcium
keywords: Ca(2+) biosensor; Endosome; Fluorescence imaging; Glucose; Pancreatic beta-cell; pH biosensor