Essential role of the histone methyltransferase G9a in cocaine-induced plasticity

Abstract

Cocaine-induced alterations in gene expression cause changes in neuronal morphology and behavior that may underlie cocaine addiction. In mice, we identified an essential role for histone 3 lysine 9 (H3K9) dimethylation and the lysine dimethyltransferase G9a in cocaine-induced structural and behavioral plasticity. Repeated cocaine administration reduced global levels of H3K9 dimethylation in the nucleus accumbens. This reduction in histone methylation was mediated through the repression of G9a in this brain region, which was regulated by the cocaine-induced transcription factor DeltaFosB. Using conditional mutagenesis and viral-mediated gene transfer, we found that G9a down-regulation increased the dendritic spine plasticity of nucleus accumbens neurons and enhanced the preference for cocaine, thereby establishing a crucial role for histone methylation in the long-term actions of cocaine.

Fig. 1

Repeated cocaine represses G9a expression in NAc through a ΔFosB-dependent mechanism. (A) mRNA expression of H3K9/K27 KMTs and KDMs in NAc 24 hr after repeated cocaine. (B) H3K9me2 levels in NAc 24 hr after repeated cocaine. (C) Analysis of gene expression after acute or repeated cocaine. Heatmaps (*) show genes upregulated in NAc 1 hr after a cocaine challenge in naïve animals (Acute), in animals treated repeatedly with cocaine (Repeated + acute) or in animals after 168 hr withdrawal from repeated cocaine (Repeated wd + acute). Associated heatmaps show how genes were affected under the other 2 conditions. Desensitized transcriptional responses following repeated cocaine are indicated (***). (D) H3K9me2 levels in NAc from NSE-tTA × tetOP-ΔFosB mice on (‘Δ FosB off’) or off (‘Δ FosB on’) doxycycline 1 hr after repeated cocaine. (E) G9a mRNA expression in NAc from NSE-tTA × tetOP-ΔFosB mice on (‘Δ FosB off’) and off (‘Δ FosB on’) doxycycline, and from mice infected with AAV-GFP or AAV-ΔFosB. Data are presented as mean ± SEM. For statistical analyses, see full figure legends in “supporting online text.”

Fig. 2

G9a in NAc regulates cocaine-induced behavioral plasticity. (A) Representative image of HSV-mediated transgene expression in NAc. Cartoon of the coronal brain slice was taken from the mouse brain atlas. (B) Conditioned place preference for cocaine and (C) H3K9me2 levels in NAc in animals infected with HSV-GFP, HSV-G9a, or HSV-G9aH1093K. (D) Conditioned place preference for cocaine and (E) H3K9me2 levels in NAc in G9a^fl/fl animals infected with AAV-GFP or AAV-Cre. (F) Conditioned place preference for cocaine and (G) H3K9me2 levels in NAc in animals receiving intra-NAc vehicle or BIX01294. Data are presented as mean ± SEM.

Fig. 3

G9a in NAc regulates cocaine-induced dendritic spine plasticity. (A) Quantitative G9a ChIP in NAc from animals treated acutely or repeatedly with cocaine, at 1 or 24 hr, respectively. APRT was used as a negative control. Data are presented as the relative fold difference from saline controls. (B) Quantitative H3K9me2 ChIP in NAc from repeated cocaine-treated animals at 24 hr, presented as the relative fold difference from saline controls. (C) Dendritic spine analysis of animals infected with HSV-GFP, HSV-G9a, or HSV-ΔJunD following repeated cocaine, and dendritic spines in G9a^fl/fl mice following HSV-Cre infection. (D) Quantitative G9a ChIP in NAc from NSE-tTA × tetOP-ΔFosB mice on (‘ΔFosB off’) and off (‘Δ FosB on’) doxycycline. (E) Dendritic spine analysis in animals infected with AAV-GFP or AAV-ΔFosB following repeated cocaine. Data are presented as mean ± SEM.