Nrf2 signaling pathway activation through Keap1-dependent and -independent mechanisms in response to acute...

The presence of benzo[a]pyrene (BaP), a highly lipophilic polycyclic aromatic hydrocarbon (PAH), in aquatic environments poses significant threats to marine organisms. Despite the established role of the Nrf2-Keap1 signaling pathway in mitigating oxidative stress, the specific mechanisms underlying Nrf2 activation in response to BaP exposure in marine bivalves remain unclear. Herein, we explored the dual activation mechanisms of the Nrf2 pathway in Mytilus coruscus through both in vivo and in vitro experiments under 50 μg/L and 16 μM BaP exposure respectively. Then, we examined the variation of key genes associated with Nrf2-Keap1 signaling pathway and antioxidant response element (ARE) after Keap1 knockdown. The results showed that BaP exposure significantly upregulated McNrf2 expression, which was further enhanced by Keap1 knockdown. Additionally, we identified a Keap1-independent mechanism involving Protein Kinase C (PKC) and glycogen synthase kinase-3β (GSK-3β), where PKC knockdown reduced Nrf2 expression and antioxidant defenses, whereas GSK-3β knockdown increased Nrf2 activity. These findings indicate that Nrf2 activation through both Keap1-dependent and -independent pathways plays a critical role in enhancing antioxidant enzyme activities, reducing reactive oxygen species (ROS) levels, and increasing total antioxidant capacity in M. coruscus exposed to BaP. Our study fills an important gap in understanding Nrf2 signaling in marine bivalves, with broader significance for elucidating the adaptive mechanisms employed by aquatic organisms against environmental pollutants.