Assessment of cation trapping by cellular acidic compartments

Methods in Enzymology
François MarceauJohanne Bouthillier

Abstract

All nucleated cells, from yeast to animal cells, concentrate cationic chemicals (weak bases with a pKa~8-10) into acidic cell compartments (low retro-diffusion under a protonated form at low pH=ion trapping). The proton pump vacuolar (V)-ATPase is the driving force of this pseudotransport that concerns acidic organelles (mainly late endosomes and lysosomes). The latter rapidly become swollen (osmotic vacuolization) and macroautophagic. Cation concentration in cells is not proved to involve membrane transporters, but is prevented or reversed by inhibitors of V-ATPase, such as bafilomycin A1. Lipophilicity is a major determinant of the apparent affinity of this pseudotransport because simple diffusion of the uncharged form supports it. Quinacrine is a formerly used antiparasitic drug that is intensely fluorescent, lipophilic, and a tertiary amine. The drug, at micromolar concentrations, is proposed as a superior probe for assessing cation trapping by cellular acidic compartments, being readily quantified using fluorometry in cell extracts and analyzed using microscopy and cytofluorometry (fluorescence settings for fluorescein being applicable). Further, cells respond to micromolar levels of quinacrine by autophagic accumulation (...Continue Reading

Citations

Mar 11, 2017·Scientific Reports·Magdalena VaterThomas Kirmeier
Jun 24, 2014·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Agnieszka Pierzyńska-MachJurek W Dobrucki
Sep 30, 2016·Journal of Applied Toxicology : JAT·Dayton M PetiboneDaniel A Casciano
Nov 28, 2020·Cell Death & Disease·Jitka FucikovaLorenzo Galluzzi
Nov 29, 2017·Environmental Science & Technology·Rebekka GuldeKathrin Fenner

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