Poster: Cell Cycle & Cytokinesis

Abs # 1089: Cell cycle arrest and redox changes induced by fluorescence microscopy

Presenter:	Dixit, Ram V., rvd10@psu.edu
Authors	Dixit, Ram V. (A)   Cyr, Richard J. (A)
Affiliations:	(A): Penn State University

The green fluorescent protein (GFP) has gained popularity as a reporter because it allows visualization of cellular constituents in living specimens. The use of GFP makes it possible to view dynamic processes in living cells, but extended viewing of cells, using fluorescence microscopy, can result in significant light energy exposure, even with confocal technology. It is therefore possible that cells experience light-induced damage that alters the physiological state of the cell and confounds observations. In order to understand the impact that extended viewing has on cells, we wanted to obtain quantitative information about the effect of light energy dosage, and observation conditions, on tobacco BY-2 cell physiology. Our results, using transgenic BY-2 cells, indicate a non-linear relationship between excitation light energy dosage and cell cycle arrest. The detrimental effects of the excitation light are dependent on its wavelength and on the presence of a GFP form that absorbs the excitation light. Moreover, fluorescence microscopy induces oxidation of the cellular redox environment as assayed using wild-type BY2 cells loaded with oxidation-sensitive dyes. Oxidation of the cellular redox environment, by light, exhibits sigmoidal kinetics and is reversible if the cells are exposed to low irradiation levels. The data highlight the importance of redox regulation for normal progression of the cell cycle, and provide a reference for judiciously choosing conditions for extended fluorescence microscopy that neither damage cells, nor lead to changes in redox physiology.