Targeting Keap1/Nrf2/ARE Signaling Pathway by Esculin in Zebrafish Larvae

Nuclear factor E2-related factor 2 (Nrf2) is a basic leucine zipper (bZIP) transcription factor that plays a crucial role in cellular defense mechanisms by regulating the expression of phase II detoxification enzymes and antioxidant proteins. Nrf2 exerts its protective effects by binding to antioxidant response elements (AREs), which are cis-acting regulatory sequences located upstream of the transcription start site (TSS) of cytoprotective genes. This study aims to identify and characterize AREs in the whole genome of zebrafish larvae exposed to esculin, a bioactive coumarin compound with antioxidant properties. Therefore, a chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-Seq) was employed using an Nrf2-specific polyclonal antibody to map genome-wide Nrf2 binding sites. Our findings reveal that Nrf2 interacts with ARE sequences in promoter-proximal and distal regions, highlighting the intricate and dynamic nature of Nrf2-mediated transcriptional regulation. These results provide novel insights into the genome-wide regulatory landscape of Nrf2 in response to esculin exposure, further elucidating its role in modulating oxidative stress-related gene expression. Overall design: Control and Esculin exposed Zebrafish larvae were immunoprecipitated with Nrf2 polyclonal antibody and precipitated with magnetic G beads, followed by downstream sequencing.