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Axonal/Dendritic Growth and Circuit Formation

開催日 2014/9/11
時間 11:00 - 12:00
会場 Poster / Exhibition(Event Hall B)

Dynamics of CREB in neuronal activity-dependent transcription studied by single-molecule imaging in cortical neurons

  • P1-097
  • 北川 宏信 / Hironobu Kitagawa:1 菅生 紀之 / Noriyuki Sugo:1 大國 紋 / Aya Ohkuni:1 山本 亘彦 / Nobuhiko Yamamoto:1 :1
  • 1:大阪大院生命機能細胞分子神経生物学 / Graduate School of Frontier Biosciences, Osaka Univ, Osaka, Japan 

Dynamics of CREB in neuronal activity-dependent transcription studied by single-molecule imaging in cortical neurons

Hironobu Kitagawa, Noriyuki Sugo, Aya Ohkuni, Nobuhiko Yamamoto
Graduate School of Frontier Biosciences, Osaka University, Osaka, Japan

Activity-dependent transcription is essential for neuronal circuit formation and synaptic plasticity. An intriguing question is how neuronal activity affects the dynamic interactions between sequence-specific transcription factors and their target sites. Here we investigated activity dependence of DNA binding and dissociation events of cAMP-response element binding protein (CREB) in living cortical neurons. For visualization of CREB at single molecule level, the fluorescent-tagged CREB in living cell nuclei was observed by highly inclined laminated optical sheet microscopy. CREB resided at restricted positions for a few seconds (dissociate rate constant: 0.54 s-1), but mutant CREB which lacks binding to its target sequence cAMP-response element (CRE) scarcely stayed at fixed locations for such a long period. To test the possibility that the residence time depends on neuronal activity, an optogenetic method with light-gated channel Channelrhodopsin-2 was applied to cultured cortical neurons. Although the photostimulation efficiently induced nuclear translocation of CREB-regulated transcriptional coactivator 1 (CRTC1), the residence time distribution of CREB was not changed before and after the photostimulation. Moreover, it was affected by neither high KCl nor tetrodotoxin treatment, which alters spontaneous firing activity. Taken together, these results suggest that basal binding of CREB to CRE in the time range of several seconds and its binding to the cofactor are crucial for activity-dependent gene expression.

Research funds: MEXT KAKENHI (Nos. 23115102, 23111516) and JSPS KAKENHI (No. 25640035).

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