Methylmercury-induced oxidative stress alters the dynamic association of Nrf2 and Fyn in primary astrocytes

Abstract

Methylmercury (MeHg) is an environmentally persistent neurodevelopmental toxicant. Humans are primarily exposed via contaminated fish consumption, which is of particular concern to populations that subsist on seafood. Extensive regulation has virtually eliminated high-dose exposure incidents. However, there continue to be significant risks associated with low-dose chronic exposure, especially in utero. Despite these concerns, research thus far has not fully elucidated the mechanism[s] of toxicity. It is, however, well-established MeHg-induces an oxidative stress response in the brain. Furthermore, this response was found to activate nuclear factor erythroid 2-related factor 2 (Nrf2) in cortical astrocytes. Nrf2 is a transcription factor, which promotes the expression of detoxifying enzymes. MeHg was demonstrated to increase detoxifying enzyme transcription. Further, Nrf2 knockdown diminished this response and exacerbated toxicity in astrocytes. In spite of these effects, it is not entirely evident whether Nrf2 directly mediates MeHg-induced neurotoxicity. Therefore, we sought to investigate other factors involved in the Nrf2-regulated detoxifying enzyme response consequent to exposure. Particular focus was dedicated to Fyn, a non-receptor tyrosine kinase, recognized to negatively regulate Nrf2. The present results indicate a toxic MeHg exposure decreases Fyn nuclear abundance concomitant with maximal Nrf2 activation in astrocytes. Moreover, at this same MeHg concentration and time-point GSK-beta3 and Akt were phosphorylated at inhibitory and activating sites, respectively. Thus, these data suggest MeHg stimulates Akt activation and GSK-beta3 inhibition to decrease Fyn nuclear localization, and sustain the Nrf2 detoxifying enzyme response in cortical astrocytes. The results further demonstrated MeHg represses Fyn expression at a sub-toxic exposure. Interestingly, this effect was concurrent with increased specificity protein 1 (Sp1), the transcription factor that promotes Fyn transcription, nuclear abundance. Sp1 was found to interact with Nrf2. Further, this interaction repressed an Sp1 specific target gene. Therefore, the present data indicate MeHg promotes Sp1 interaction with Nrf2, which subsequently represses Fyn expression in cortical astrocytes. These results provide greater insight into the MeHg and Nrf2 connection. Although by no means exhaustive of this relationship, it is evident Nrf2 has a potential role in MeHg-induced neurotoxicity.