IMAT R61 Abstract
In addition to its well-recognized roles in neuronal computation and cardiac contractions, the voltage across the
plasma membrane has been implicated as a fundamental regulator of differentiation and proliferation. The
concurrent regulation of proliferation and stem cell states by membrane voltage contributes to normal tissue
morphogenesis, development, and diseases such as cancer. Membrane hyperpolarization tends to promote
increased differentiation and reduced proliferation while depolarization is associated with immature and
proliferative stem-like cell states. Of clinical relevance, cancer cells and cancer stem-like cells exhibit depolarized
membrane voltage values compared with the normal differentiated cells from which they arise. Despite the
strong association between voltage and cancer cell phenotypes, voltage modulation has not been effectively
exploited as a drug target against cancer, largely reflecting the technological challenges with membrane voltage
measurements in physiologically relevant contexts. Current techniques to measure membrane voltage, including
electrode arrays and voltage sensitive dyes, have limitations in the number of experimental observations
possible, compatibility with complementary phenotypic readouts, the duration of sampling, and the inability to
perform longitudinal cell analyses. These technologic challenges motivate our hypothesis that integration of
genetically encoded voltage and cell cycle sensors will provide a robust and novel high-throughput platform to
identify modulators of membrane voltage and cell proliferation. We propose to develop, optimize, and validate
a cell-based technology platform integrating JEDI, a novel voltage indicator, and FUCCI cell-cycle sensors to
enable real time high-throughput analysis of concurrent changes in voltage and cell proliferation in living cells.
To build the platform, we propose to demonstrate high-throughput imaging of voltage dynamics (Aim 1) and cell
cycle states (Aim 2) in glioblastoma stem cells. We next propose to deploy these technologies for high-throughput
quantification of the impact of ion channel-modulating drugs on the membrane voltage and cell-cycle changes
(Aim 3). This novel platform is expected, for the first time, to provide a tool for real-time live cell readouts of
concurrent changes in membrane voltage and cell proliferation. Besides its implications for the treatment of
cancer, we anticipate this platform to have far reaching basic science and clinical translational applications in
neuroscience, stem cell and iPSC biology, cell-based therapies, and cardiovascular diseases.
