Our laboratory investigates the glycosylation, assembly, structure, trafficking and function of ion channel complexes. We rely on traditional electrophysiological, biochemical, and imaging modalities, but we also design, develop, and utilize novel chemical tools to interrogate a wide variety of ion channels and membrane transport proteins responsible for cardiac and neuronal function. Thus, we have synthetic organic chemists, glycobiologists, membrane protein biochemists, and electrophysiologists working together to elucidate the molecular underpinnings of these membrane transport proteins in both healthy and diseased tissues.
Given the laboratory's enthusiasm for studying ion channels, our lab has been developing a new approach to visualize ions exiting and entering cells. Our first publication in Cell Chemical Biology enabled the visualization of proton accumulation and depletion on the extracellular side of the membrane. Proton fluxes were visualized from voltage-gated ion channels, transporters, and mutant channels harboring mutations associated with human disease. The video (left) shows protons rushing into a cell after the channels were opened with hyperpolarizing pulse (-120 mV). The initial fluorescent signal is due to protonated fluorescent sensors covalently attached to the cell's glycocalyx. Proton channel activation at -120 mV results in proton depletion and loss of the fluorescent signal, which slowly returns after the channels are closed (30 mV).
Fluorescent Visualization of Cellular Proton Fluxes.
Cell Chem Biol. 2016 Nov 14;:
Authors: Zhang L, Bellve K, Fogarty K, Kobertz WR
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.
PMID: 27916567 [PubMed - as supplied by publisher]
Lazare Research Building 804
Campus Map (pdf)
William R. Kobertz, Ph.D.
Department of Biochemistry and Molecular Pharmacology
University of Massachusetts Medical School
364 Plantation Street LRB804, Worcester, MA 01605-4321
We are always interested in applications from qualified candidates at the postdoctoral and research associate levels. UMMS GSBS graduate students interested in rotating in the Kobertz Lab should email Dr. Kobertz to set up an appointment.
Undergraduates interested in pursuing a PhD at UMass Medical School should apply directly to the Graduate School of Biomedical Sciences Program.