Novel mechanisms for regulation by glutamate signals in expression of mitochondrial gene

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A systemic administration of kainate dramatically enhanced AP-1 DNA binding in both mitochondrial and nuclear extracts of mouse cerebral cortex and hippocampus 1 h to 3 days later. Unlabeled AP-1 probe selectively competed for AP-1 DNA binding in mitochondrial extracts of cortex and hippocampus obtained from mice injected with kainate. Supershift and immunoblotting analyses revealed participation of c-Fos, Fos-B and Jun-B proteins in potentiation by kainate of mitochondrial AP-1 DNA binding in cortex and hippocampus. An immunohistochemical study demonstrated marked expression by kainate of c-Fos protein in the pyramidal and dentate granular layers, while an immunoelectron microscopic analysis showed localization of c-Fos protein within mitochondria, as well as nuclei, of the CA1 pyramidal and dentate granular cells in hippocampus obtained 2 h after the administration of kainate. There are 10 sites with sequences similar to the nuclear AP-1 site in the non-coding region. Of 10 pieces(termed as MT-1 to MT-10) of synthesized double-stranded oligonucleotides containing each mitochondrial AP-1-like site, MT-3,MT-4, and MT-9 were effective in inhibiting mitochondrial AP-1 DNA binding enhanced by kainate. Electrophoresis mobility shift analysis using radiolabeled MT-3 and MT-9 as probes demonstrated that marked enhancement was seen with binding of these 2 probes in hippocampal mitochondrial extracts prepared 2 to 6 h after kainate treatment. Unlabeled AP-1 probe was more potent than unlabeled MT-9 in inhibiting the mitochondrial MT-9 binding. Immunoprecipitation analysis using anti-c-Fos antibody demonstrated that c-Fos associated with mitochondrial genome in the hippocampal mitochondria prepared from kainate-treated animals. These results suggest that AP-1 complex expressed by in vivo treatment with kainate would bind to AP-1-like sites in the non-coding region of mitochondrial genome following translocation into the mitochondria in murine hippocampus.