Revealing Subcellular Redox Dynamics with Multiplex Imaging of Compartment‐Specific Redox Probes

Abstract

Reactive oxygen species (ROS), redox enzymes, and redox buffers are central to the propagation of redox signals and oxidative stress between cells in the tissue environment and between organelles in the cellular environment. For example, hydrogen peroxide generated in mitochondria can alter the thiol oxidation states on proteins to alter their function, and subsequently redox active biomolecules such as glutathione can act as reducing equivalents in the face of oxidative stress. In some cases, it has become apparent that ROS mediate physiologically important signaling across compartments, such as in mitonuclear retrograde signaling. However, it has remained technically challenging to quantitatively correlate compartment‐specific redox dynamics with subcellular resolution within live single cells. In order to address this challenge, we are developing genetically‐encoded biosensors that enable the visualization of redox dynamics simultaneously within multiple compartments within the same cell. Our approach utilizes Förster‐type resonance energy transfer in a spectral relay strategy to extend the fluorescence emission of the redox‐sensitive green fluorescent protein (roGFP) into orange and red emission wavelengths. Furthermore, we are developing multiplexed, multiphoton imaging methods to study redox coupling between organelles and compartment‐specific dynamics in thick neural tissues.Support or Funding InformationWe gratefully acknowledge support from the Ralph W. and Grace M. Showalter Trust, the Purdue Research Foundation, and from National Institutes of Health grants R21 NS092010 and R21 EY026425.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.