The developing brain is a primary target site for lead (Pb) induced toxicity. Pb-exposure during critical periods of pre- and postnatal development may impair maturation of glia and neurons and alter neural network. The activator protein (AP)-1 plays an important function in regulating the transcription of developmentally-important genes, such as glial fibrillary acidic protein, whose promoter region contains a AP-1 DNA binding site.
The aims of this project are: 1. To understand the effect of Pb on the induction of AP-1 in glial-rich primary cultures from embryonic mouse brain. 2. To study the effect of Pb on AP-1 DNA binding activity. 3. To study the direct effect of Pb on Ref-1, a protein involved in the redox regulation of AP-1 DNA binding activity. Electrophoretic mobility shift assays performed in our laboratory showed a 3 fold increase of AP-1 DNA binding activity after a 1 h exposure to 10 microM Pb in cortical astrocyte cultures. Whereas, Ref-1 protein in nuclear extracts measured by Western blotting was already increased after a 15 minutes exposure to 10 microM Pb, reaching a 2 fold increase after 30 minutes of exposure. In contrast, no difference in Ref-1 protein level was observed from total cell extracts after Pb exposure, suggesting that Pb may facilitate nuclear translocation of Ref-1 but not promote the induction of Ref-1. The present data suggest that the increase of AP-1 DNA binding activity by Pb may be enhanced or elicited by the nuclear translocation of Ref-1. Our present working hypothesis suggests that Pb may interact either with Zn-binding sites or with the nuclear targeting domain of Ref-1, thereby, facilitating the transport of Ref-1 from the cytoplasm into the nucleus.
