Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Locus-specific epigenetic remodeling controls addiction- and depression-related behaviors

Abstract

Chronic exposure to drugs of abuse or stress regulates transcription factors, chromatin-modifying enzymes and histone post-translational modifications in discrete brain regions. Given the promiscuity of the enzymes involved, it has not yet been possible to obtain direct causal evidence to implicate the regulation of transcription and consequent behavioral plasticity by chromatin remodeling that occurs at a single gene. We investigated the mechanism linking chromatin dynamics to neurobiological phenomena by applying engineered transcription factors to selectively modify chromatin at a specific mouse gene in vivo. We found that histone methylation or acetylation at the Fosb locus in nucleus accumbens, a brain reward region, was sufficient to control drug- and stress-evoked transcriptional and behavioral responses via interactions with the endogenous transcriptional machinery. This approach allowed us to relate the epigenetic landscape at a given gene directly to regulation of its expression and to its subsequent effects on reward behavior.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: Engineered transcription factors targeting the Fosb promoter bidirectionally regulate gene expression in NAc neurons via epigenetic manipulation.
Figure 2: HSV-Fosb-ZFP35-p65 and -G9a in the NAc specifically regulate FosB/ΔFosB expression.
Figure 3: Cocaine induction of FosB/ΔFosB protein expression and endogenous transcription factor binding is blocked by HSV-Fosb-ZFP35-G9a in the NAc.
Figure 4: Engineered transcription factors bidirectionally modulate cocaine- and stress-evoked behaviors.

References

  1. Renthal, W. et al. Genome-wide analysis of chromatin regulation by cocaine reveals a role for sirtuins. Neuron 62, 335–348 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Kennedy, P.J. et al. Class I HDAC inhibition blocks cocaine-induced plasticity by targeted changes in histone methylation. Nat. Neurosci. 16, 434–440 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Malvaez, M., Mhillaj, E., Matheos, D.P., Palmery, M. & Wood, M.A. CBP in the nucleus accumbens regulates cocaine-induced histone acetylation and is critical for cocaine-associated behaviors. J. Neurosci. 31, 16941–16948 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Robison, A.J. & Nestler, E.J. Transcriptional and epigenetic mechanisms of addiction. Nat. Rev. Neurosci. 12, 623–637 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Feng, J. et al. Chronic cocaine-regulated epigenomic changes in mouse nucleus accumbens. Genome Biol. 15, R65 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  6. Kumar, A. et al. SOM Chromatin remodeling is a key mechanism underlying cocaine-induced plasticity in striatum. Neuron 48, 303–314 (2005).

    Article  CAS  PubMed  Google Scholar 

  7. Kelz, M.B. et al. Expression of the transcription factor ΔFosB in the brain controls sensitivity to cocaine. Nature 401, 272–276 (1999).

    Article  CAS  PubMed  Google Scholar 

  8. Vialou, V. et al. ΔFosB in brain reward circuits mediates resilience to stress and antidepressant responses. Nat. Neurosci. 13, 745–752 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Robison, A.J. et al. Behavioral and structural responses to chronic cocaine require a feedforward loop involving ΔFosB and calcium/calmodulin-dependent protein kinase II in the nucleus accumbens shell. J. Neurosci. 33, 4295–4307 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Maze, I. et al. Cocaine dynamically regulates heterochromatin and repetitive element unsilencing in nucleus accumbens. Proc. Natl. Acad. Sci. USA 108, 3035–3040 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Covington, H.E. et al. A role for repressive histone methylation in cocaine-induced vulnerability to stress. Neuron 71, 656–670 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Maze, I. et al. Essential role of the histone methyltransferase G9a in cocaine-induced plasticity. Science 327, 213–216 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Sun, H. et al. Morphine epigenomically regulates behavior through alterations in histone H3 lysine 9 dimethylation in the nucleus accumbens. J. Neurosci. 32, 17454–17464 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Snowden, A.W., Gregory, P.D., Case, C.C. & Pabo, C.O. Gene-specific targeting of H3K9 methylation is sufficient for initiating repression in vivo. Curr. Biol. 12, 2159–2166 (2002).

    Article  CAS  PubMed  Google Scholar 

  15. Sanjana, N.E. et al. A transcription activator-like effector toolbox for genome engineering. Nat. Protoc. 7, 171–192 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Konermann, S. et al. Optical control of mammalian endogenous transcription and epigenetic states. Nature 500, 472–476 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Mendenhall, E.M. et al. Locus-specific editing of histone modifications at endogenous enhancers. Nat. Biotechnol. 31, 1133–1136 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Laganiere, J. et al. An engineered zinc finger protein activator of the endogenous glial cell line-derived neurotrophic factor gene provides functional neuroprotection in a rat model of Parkinson's disease. J. Neurosci. 30, 16469–16474 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Beerli, R.R. & Barbas, C.F. Engineering polydactyl zinc-finger transcription factors. Nat. Biotechnol. 20, 135–141 (2002).

    Article  CAS  PubMed  Google Scholar 

  20. Gerritsen, M.E. et al. CREB-binding protein/p300 are transcriptional coactivators of p65. Proc. Natl. Acad. Sci. USA 94, 2927–2932 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Covington, H.E. et al. Antidepressant actions of histone deacetylase inhibitors. J. Neurosci. 29, 11451–11460 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Malvaez, M., Mhillaj, E., Matheos, D.P., Palmery, M. & Wood, M.A. CBP in the nucleus accumbens regulates cocaine-induced histone acetylation and is critical for cocaine-associated behaviors. J. Neurosci. 31, 16941–16948 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Kennedy, P.J. et al. Class I HDAC inhibition blocks cocaine-induced plasticity by targeted changes in histone methylation. Nat. Neurosci. 16, 434–440 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Kumar, A. et al. Chromatin remodeling is a key mechanism underlying cocaine-induced plasticity in striatum. Neuron 48, 303–314 (2005).

    Article  CAS  PubMed  Google Scholar 

  25. Peixoto, L. & Abel, T. The role of histone acetylation in memory formation and cognitive impairments. Neuropsychopharmacology 38, 62–76 (2013).

    Article  CAS  PubMed  Google Scholar 

  26. Rogge, G.A., Singh, H., Dang, R. & Wood, M.A. HDAC3 is a negative regulator of cocaine-context-associated memory formation. J. Neurosci. 33, 6623–6632 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Rea, S. et al. Regulation of chromatin structure by site-specific histone H3 methyltransferases. Nature 406, 593–599 (2000).

    Article  CAS  PubMed  Google Scholar 

  28. Garriga-Canut, M. et al. Synthetic zinc finger repressors reduce mutant huntingtin expression in the brain of R6/2 mice. Proc. Natl. Acad. Sci. USA 109, E3136–E3145 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Smith, A.E., Hurd, P.J., Bannister, A.J., Kouzarides, T. & Ford, K.G. Heritable gene repression through the action of a directed DNA methyltransferase at a chromosomal locus. J. Biol. Chem. 283, 9878–9885 (2008).

    Article  CAS  PubMed  Google Scholar 

  30. Fritsch, L. et al. A subset of the histone H3 lysine 9 methyltransferases Suv39h1, G9a, GLP and SETDB1 participate in a multimeric complex. Mol. Cell 37, 46–56 (2010).

    Article  CAS  PubMed  Google Scholar 

  31. Filion, G. J. & van Steensel, B. Reassessing the abundance of H3K9me2 chromatin domains in embryonic stem cells. Nat. Genet. 42, 4 (2010).

    Article  CAS  PubMed  Google Scholar 

  32. Vialou, V. et al. Serum response factor and cAMP response element binding protein are both required for cocaine induction of FosB. J. Neurosci. 32, 7577–7584 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Zhang, F. et al. Efficient construction of sequence-specific TAL effectors for modulating mammalian transcription. Nat. Biotechnol. 29, 149–153 (2011).

    Article  PubMed  PubMed Central  Google Scholar 

  34. Maniatis, T., Goodbourn, S. & Fischer, J.A. Regulation of inducible and tissue-specific gene expression. Science 236, 1237–1245 (1987).

    Article  CAS  PubMed  Google Scholar 

  35. Ernst, J. & Kellis, M. Interplay between chromatin state, regulator binding, and regulatory motifs in six human cell types. Genome Res. 23, 1142–1154 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Chaudhury, D. et al. Rapid regulation of depression-related behaviors by control of midbrain dopamine neurons. Nature 493, 532–536 (2013).

    Article  CAS  PubMed  Google Scholar 

  37. Smith, D.J. & Konarska, M.M. A critical assessment of the utility of protein-free splicing systems. RNA 15, 1–3 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Grimmer, M.R. et al. Analysis of an artificial zinc finger epigenetic modulator: widespread binding but limited regulation. Nucleic Acids Res. 42, 10856–10868 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Kouzarides, T. Chromatin modifications and their function. Cell 128, 693–705 (2007).

    Article  CAS  PubMed  Google Scholar 

  40. Lachner, M., O'Carroll, D., Rea, S., Mechtler, K. & Jenuwein, T. Methylation of histone H3 lysine 9 creates a binding site for HP1 proteins. Nature 410, 116–120 (2001).

    Article  CAS  PubMed  Google Scholar 

  41. Hathaway, N.A. et al. Dynamics and memory of heterochromatin in living cells. Cell 149, 1447–1460 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Cedar, H. & Bergman, Y. Linking DNA methylation and histone modification: patterns and paradigms. Nat. Rev. Genet. 10, 295–304 (2009).

    Article  CAS  PubMed  Google Scholar 

  43. Paxinos, G. & Franklin, K.B.J. The Mouse Brain in Stereotaxic Coordinates (Academic Press, 2012).

    Google Scholar 

  44. Langmead, B., Trapnell, C., Pop, M. & Salzberg, S.L. Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol. 10, R25 (2009).

    Article  PubMed  PubMed Central  Google Scholar 

  45. Shen, L. et al. diffReps: detecting differential chromatin modification sites from ChIP-seq data with biological replicates. PLoS ONE 8, e65598 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Lobo, M.K. et al. ΔFosB induction in striatal medium spiny neuron subtypes in response to chronic pharmacological, emotional and optogenetic stimuli. J. Neurosci. 33, 18381–18395 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Becker, A. et al. Glutaminyl cyclase-mediated toxicity of pyroglutamate-beta amyloid induces striatal neurodegeneration. BMC Neurosci. 14, 108 (2013).

    Article  PubMed  PubMed Central  Google Scholar 

  48. US National Research Council Committee on Guidelines for the Use of Animals in Neuroscience and Behavioral Research. Guidelines for the Care and Use of Mammals in Neuroscience and Behavioral Research (National Academies Press (US), 2003).

  49. Maze, I. et al. Cocaine dynamically regulates heterochromatin and repetitive element unsilencing in nucleus accumbens. Proc. Natl. Acad. Sci. USA 108, 3035–3040 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Allan, R.S. et al. An epigenetic silencing pathway controlling T helper 2 cell lineage commitment. Nature 487, 249–253 (2012).

    Article  CAS  PubMed  Google Scholar 

  51. Jennings, J.H. et al. Distinct extended amygdala circuits for divergent motivational states. Nature 496, 224–228 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Golden, S.A. et al. Epigenetic regulation of RAC1 induces synaptic remodeling in stress disorders and depression. Nat. Med. 19, 337–344 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

The authors wish to thank G. Stuber and R. Ung for their help in generating social interaction heat maps. This work was supported by grants from the National Institute on Drug Abuse (E.A.H. and E.J.N.), the National Institute of Mental Health and the Hope for Depression Research Foundation (E.J.N.).

Author information

Authors and Affiliations

Authors

Contributions

E.A.H. designed and executed the biochemical, molecular and behavioral experiments (viral packaging constructs, qRT-PCR expression analysis, chromatin immunoprecipitation (mouse and human), immunohistochemical preparation and behavioral assays). H.M.C. and H.S. generated viral packaging constructs, conducted expression analysis and prepared chromatin. C.J.P. and S.A.G. analyzed immunohistochemical data. N.S. and L.S. conducted genome-wide sequence analysis. J.F. and E.A.H. performed DNA methylation sequencing. S.A.G. and S.J.R. generated human chromatin. E.A.H., H.M.C., H.S., J.J.W., M.M.-R., D.F. and M.-H.H. performed stereotaxic mouse surgery. J.P.H. generated mouse locomotor heat maps. C.S.T. prepared human postmortem brain tissue. R.L.N. generated HSV viral vectors. H.S.Z. generated the G9a catalytic domain construct. S.K., M.A.G. and C.N. generated the ZFP constructs. F.Z. generated the TALE constructs. C.J.P., E.A.H. and H.M.C. analyzed the data conducted statistical analyses. E.A.H. and E.J.N. discussed the data and wrote the manuscript.

Corresponding author

Correspondence to Eric J Nestler.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 A suite of Fosb-ZFPs bidirectionally regulate Fosb expression in vitro.

(a) FosB/ΔFosB mRNA was significantly induced by several FosB-ZFP-p65 and -G9a constructs expressed in N2a cells and harvested after 48 hours. FosB-ZFP-NFD constructs activated gene expression to a lesser extent or not at all. Data are normalized to mock transfected cells. Complete statistics are available in Supplementary Table 3. Student's unpaired t-test: *P<0.05, *P<0.10. Data are presented as mean ± s.e.m.

Source data

Supplementary Figure 2 HSV-ZFPs specifically infect neurons in NAc and inhibit basal Fosb expression.

(a) HSV-GFP specifically infects DARPP-32 positive neurons in the NAc. White arrows indicate double labeled cells. (b) NAc injection of HSV-FosB-ZFP35-G9a repressed expression of FosB/ΔFosB in HSV infected (GFP+) cells [t13=3.55, *P=0.033; n=7, 8] compared to HSV-FosB-ZFP35-G9a (72 hours post HSV injection). Data are presented as mean ± s.e.m.

Source data

Supplementary Figure 3 Regulation of Fosb expression and reward behavior by Fosb-TALEs and a suite of additional catalytic domains fused to Fosb-ZFP35.

(a) Locations of FosB-ZFP and -TALE binding relative to the FosB TSS. The location of functional SRF and CREB sites are shown. (b) FosB/ΔFosB mRNA expression in the NAc was significantly induced by HSV-FosB-TALE2-VP64 [FosB: t8=3.03, *P=0.016; ΔFosB: t8=6.40, *P=0.000; n=5], and -FosB-TALE3-VP64 [FosB: t8=2.79, *P=0.023; ΔFosB: t8=4.01, *P=0.004; n=5] compared to control virus. (c) The binding sites of the 6-finger ZFP35 and 17-RVD (repeat variable diresidue) TALE1 recognize the FosB promoter at overlapping sites approximately 250 bp upstream from the FosB TSS. (d) FosB-ZFP35-p65 [FosB: t4=5.91, *P=0.004; ΔFosB: t4=26.11, *P=0.000; n=3], -p65x2 [FosB: t4=2.30, *P=0.000; ΔFosB: t4=3.46, *P=0.026; n=3], -VP16 [FosB: t4=9.04, *P=0.001; ΔFosB: t4=7.45, *P=0.002; n=3], and -VP64 [FosB: t4=19.40, *P=0.001; ΔFosB: t4=7.45, *P=0.001; n=3] and FosB-TALE1-VP64 [FosB: t4=6.80, *P=0.002; ΔFosB: t4=15.17, *P=0.000; n=3] activate FosB/ΔFosB mRNA levels when expressed in Neuro2a cells and harvested after 48 hours. Data are normalized to mock transfected cells. (e) HSV-FosB-TALE1-VP64 in NAc sensitizes cocaine-induced hyperactivity over time. There is a significant interaction between day, cocaine treatment, and virus [F(3,32)=3.42, *P=0.029]. TALE1-VP64 sensitizes the effect of cocaine on locomotor activity [main effect of day among TALE1-VP64 [F(3,35)=9.92, *P=0.000, n=10], but not GFP control [F(3,36)=2.05, P=0.126, n=9]. HSV-GFP data are the same as in Fig. 4d-e. Heat maps show representative locomotor data within the chamber for mice over the course of repeated cocaine exposure. Data are presented as mean ± s.e.m.

Source data

Supplementary Figure 4 Differential regulation by Fosb-ZFPs in vitro and in vivo.

(a) Expression of FosB-ZFP35-G9a differs between N2a cells and NAc [t6=4.50, *P=0.004; n=3,5]. (b) Titration of the amount of FosB-ZFP35-G9a expression in N2a cells leads to a reduction in the amount of induced mRNA expression of ΔFosB but not FosB. Using a 2-tailed Pearson correlation, we found a significant correlation between μg of transfected DNA and mRNA fold change of ΔFosB [R12=0.83, *P=0.001] but not FosB [R12=.66, P=0.665]. Data are presented as mean ± s.e.m.

Source data

Supplementary Figure 5 Full blots of experiment shown in Figure 1e.

Supplementary information

Supplementary Figures and Tables

Supplementary Figures 1–5 and Supplementary Tables 1–3 (PDF 18467 kb)

Supplementary Methods Checklist

Reporting Checklist for Nature Neuroscience (PDF 122 kb)

Source data

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Heller, E., Cates, H., Peña, C. et al. Locus-specific epigenetic remodeling controls addiction- and depression-related behaviors. Nat Neurosci 17, 1720–1727 (2014). https://doi.org/10.1038/nn.3871

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nn.3871

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing