Fluorescent Visualization of Cellular Proton Fluxes

Lejie Zhang; Karl Bellve; Kevin Fogarty; William R Kobertz

doi:10.1016/j.chembiol.2016.10.013

Fluorescent Visualization of Cellular Proton Fluxes

Cell Chem Biol. 2016 Dec 22;23(12):1449-1457. doi: 10.1016/j.chembiol.2016.10.013. Epub 2016 Dec 1.

Authors

Lejie Zhang¹, Karl Bellve², Kevin Fogarty², William R Kobertz³

Affiliations

¹ Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605-2324, USA; Programs in Neuroscience and Chemical Biology, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605-2324, USA.
² Biomedical Imaging Group, Molecular Medicine, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605-2324, USA.
³ Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605-2324, USA; Programs in Neuroscience and Chemical Biology, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605-2324, USA. Electronic address: william.kobertz@umassmed.edu.

Abstract

Cells use plasma membrane proton fluxes to maintain cytoplasmic and extracellular pH and to mediate the co-transport of metabolites and ions. Because proton-coupled transport often involves movement of multiple substrates, traditional electrical measurements provide limited information about proton transport at the cell surface. Here we visualize voltage-dependent proton fluxes over the entire landscape of a cell by covalently attaching small-molecule fluorescent pH sensors to the cell's glycocalyx. We found that the extracellularly facing sensors enable real-time detection of proton accumulation and depletion at the plasma membrane, providing an indirect readout of channel and transporter activity that correlated with whole-cell proton current. Moreover, the proton wavefront emanating from one cell was readily visible as it crossed over nearby cells. Given that any small-molecule fluorescent sensor can be covalently attached to a cell's glycocalyx, our approach is readily adaptable to visualize most electrogenic and non-electrogenic transport events at the plasma membrane.

Keywords: glycocalyx; membrane transport; pH; proton-coupled transport; voltage-gatedion channels.

Grants and funding

R01 GM070650/GM/NIGMS NIH HHS/United States