Activation of Nrf2 at Critical Windows of Development Alters Tissue-Specific Protein S-Glutathionylation in the Zebrafish (Danio rerio) Embryo

Abstract

Activation of Nrf2—the master regulator of antioxidative response—at different stages of embryonic development has been shown to result in changes in gene expression, but the tissue-specific and downstream effects of Nrf2 activation during development remain unclear. This work seeks to elucidate the tissue-specific Nrf2 cellular localization and the downstream changes in protein S-glutathionylation during critical windows of zebrafish (Danio rerio) development. Wild-type and mutant zebrafish embryos with a loss-of-function mutation in Nrf2a were treated with two canonical activators, sulforaphane (SFN; 40 µM) or tert-butylhydroquinone (tBHQ; 1 µM), for 6 h at either pharyngula, hatching, or the protruding-mouth stage. Nrf2a protein and S-glutathionylation were visualized in situ using immunohistochemistry. At the hatching stage, Nrf2a protein levels were decreased with SFN, but not tBHQ, exposure. Exposure to both activators, however, decreased downstream S-glutathionylation. Stage- and tissue-specific differences in Nrf2a protein and S-glutathionylation were identified in the pancreatic islet and liver. Protein S-glutathionylation in Nrf2a mutant fish was increased in the liver by both activators, but not the islets, indicating a tissue-specific and Nrf2a-dependent dysregulation. This work demonstrates that critical windows of exposure and Nrf2a activity may influence redox homeostasis and highlights the importance of considering tissue-specific outcomes and sensitivity in developmental redox biology.