Biophysical Society Thematic Meeting - June 28-July 1, 2015

New Biological Frontiers Illuminated by Molecular Sensors and Actuators

Poster Abstracts

11-POS Board 11 Revealing Proteostasis Capacity in Cells by a Fluorescent Sensor Yuning Hong . University of Melbourne, Melbourne, Australia.

Maintaining proteostasis is an essential housekeeping function for cell survival. Proteostasis is in principle affected in any disease that involves misfolded or mutant proteins that do not fold with normal efficiencies; and hence overdraw on the finite proteostasis resources of the cell. Tracking the proteostasis capacity of cells has the generic potential to track neurodegenerative diseases of diverse specific molecular origins. Building new approaches to identify the efficiency of proteostasis is thus highly desired in order to track the risk of cells succumbing to damage from protein misfolding and aggregation. Here we describe our approach to revealing the proteostasis states in Huntington’s disease cell culture models by a cysteine-reactive aggregation-induced emission (AIE) fluorogen. Stress conditions, such as protein misfolding and environmental stimuli, will lead to an elevated level of unfolded proteins. The fluorogen, TPE-MI, can react with the unfolded proteins with free cysteine residue exposed and turns on its fluorescence. Because of the unique AIE characteristic, reacting with small biothiols, such as glutathione, would not change the fluorescence of TPE-MI. The signal from TPE-MI can thus be correlated to the amount of unfolded proteins and reflect the proteostasis capacity in cells. We then apply this method to investigate the proteostasis capacity in the subpathogenic and pathogenic Httexon 1 cells by using flow cytometry. Our results show distinct fluorescence signals between the pathogenic and subpathogenic cells. By incorporating pulse shape analysis, we are able to differentiate the signals from the non-inclusion and inclusion cell population within the same sample. Our results suggest that the collapse of proteostasis is one of the features for Huntington’s diseases and such effect emerges in cells prior to the formation of visible aggregates.

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