Heterogeneity in the population response of a human cell line to hydrogen peroxide as measured by a genetically encoded sensor

Abstract

Genetically encoded ratiometric sensors can provide valuable mechanistic understanding of biological systems. Characterization of cellular response of these sensors is the first step in validating their use. Here, we characterize the response of a genetically encoded H₂O₂ sensor, HyPer, expressed in HeLa cells. Using quantitative fluorescence microscopy, we found significant heterogeneity in HyPer response among the cell population. Further analysis showed that the variation in HyPer response was dependent on expression of HyPer protein as well as on cell cycle phase. Cells with higher levels of expressed HyPer protein showed a stronger HyPer response to H₂O₂. Cells synchronized in S-phase showed a weaker HyPer response than unsynchronized cells. It was determined that this weaker response could be a function of higher antioxidant capacity in S-phase cells. The dependence of HyPer response on these factors needs to be accounted for to avoid experimental artifacts.