# The Ancestral KEAP1-NRF Pathway in Amphioxus Branchiostoma japonicum: Implications for the Evolution of Antioxidant Defense System

## TL;DR

This study explores the KEAP1-NRF pathway in amphioxus, revealing its role in antioxidant defense and evolutionary significance.

## Contribution

The discovery of BjKEAP1 and BjNRF as ancestral homologs in amphioxus provides new insights into the evolutionary origins of the antioxidant defense system.

## Key findings

- BjNRF combines features of vertebrate NRF1 and NRF2, suggesting an evolutionary intermediate.
- BjKEAP1 regulates BjNRF's cytoplasmic localization, and sulforaphane induces its nuclear translocation.
- Exposure to Benzo[a]pyrene activates the stress response system in amphioxus.

## Abstract

The Kelch-like ECH-associated protein 1 (KEAP1)/Nuclear factor E2-related factor 2 (NRF2) pathway is a key mechanism that responds to oxidative stress and xenobiotic stimuli in vertebrates. However, knowledge of its evolutionary origins remains limited. In this study, we identify the ancestral homologues of KEAP1 and NRF (BjKEAP1 and BjNRF) in cephalochordate amphioxus (Branchiostoma japonicum). BjNRF uniquely combines the feature domains of vertebrates NRF1 and NRF2, marking it as an evolutionary intermediate. High expression levels of Bjkeap1 and Bjnrf in the gill, hepatic cecum, and intestine highlight their roles in environmental defense at key interface tissues. Functional studies reveal that BjKEAP1 regulates the cytoplasmic localization of BjNRF. Typical NRF2 activator sulforaphane (SFN) induces its nuclear translocation and significantly elevates the transcriptional expression of BjNRF and phase II detoxification enzymes. Moreover, exposure to the environmental toxin Benzo[a]pyrene (BaP) activates this stress response system. These findings bridge critical gaps in our understanding of this pathway in basal chordates and offer new insights into the evolutionary trajectory of the KEAP1-NRF system. Furthermore, this study highlights crucial implications for the conservation of amphioxus in deteriorating marine environments.