Transcription factor activator protein‐1 expressed by kainate treatment can bind to the non‐coding region of mitochondrial genome in murine hippocampus

Abstract

We have demonstrated previously that the transcription factor activator protein‐1 (AP‐1) complex is translocated into mitochondria into the nucleus in murine hippocampus after systemic kainate injection (Ogita et al. [2002] J. Neurosci. 22:2561–2570). The present study investigates whether the mitochondrial AP‐1 complex translocated in response to kainate treatment binds to AP‐1‐like sites located at the non‐coding region of the mitochondrial genome in mouse hippocampus. There are 10 sites with sequences similar to the nuclear AP‐1 site in the non‐coding region. Of 10 pieces (MT‐1–MT‐10) of synthesized double‐stranded oligonucleotides, each containing a 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 probes demonstrated marked enhancement with binding of these 2 probes in hippocampal mitochondrial extracts prepared 2–6 hr after kainate treatment. Unlabeled AP‐1 probe was more potent than unlabeled MT‐9 probe in inhibiting the mitochondrial MT‐9 binding. Supershift analysis revealed participation of particular Fos/Jun family proteins, such as c‐Fos, Fos‐B, c‐Jun, Jun‐B, and Jun‐D, in MT‐9 binding in hippocampal mitochondrial extracts prepared 4 hr after kainate treatment. Immunoprecipitation analysis using anti‐c‐Fos antibody demonstrated that c‐Fos associated with the mitochondrial genome in hippocampal mitochondria prepared from kainate‐treated animals. These results suggest that the AP‐1 complex expressed by in vivo kainate treatment would bind to AP‐1‐like sites in the non‐coding region of the mitochondrial genome after translocation into mitochondria from murine hippocampus.