Finally, we asked whether the mechanism of transcriptional repression by Nurr1 in astrocytes is similar to that in microglia. Treatment of primary mouse astrocytes with IL1β induced the interaction of Nurr1 with p65 (Fig. 6A) and induced recruitment of both Nurr1 and p65 to the iNOS-promoter (Fig. 6B). Recruitment of Nurr1 to the iNOS promoter was blocked by inhibition of GSK3β by SB21 (Fig. 6C), consistent with TLR4 and IL1β receptors sharing the MyD88 signaling pathway (Verstrepen et al., 2008). Nurr1 also interacted with CoREST in astrocytes in a manner that was stimulated by IL1β (Fig. 6D), and both molecules were recruited to the iNOS-promoter, as was observed in microglia (Fig. 6E). Knockdown of the components of CoREST repressor complex such as LSD1, G9a and HDAC1 also up-regulated iNOS, CSF1 and Ncf1 genes, suggesting that the CoREST-complex is required for Nurr1-mediated transcriptional repression in astrocytes (Fig. 6F–H). Finally, knockdown of Nurr1 in astrocytes resulted in prolonged occupancy of p65 on the iNOS promoter (Fig. 6I), similar to results obtained in microglia (Fig. 4H).
