Epigenetics and Addiction - American Society for Clinical

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Epigenetics and Addiction JL Cadet1, MT McCoy1 and S Jayanthi1 Addictions are public health menaces. However, despite advances in addiction research, the cellular or molecular mechanisms that cause transition from recreational use to addiction remain to be elucidated. We have recently suggested that addiction may be secondary to long-term epigenetic modifications that determine the clinical course of substance use disorders. A better understanding of epigenetic mechanisms in animal models that mimic human conditions should help to usher in a new area of drug development against addiction. Addiction and abuse of licit and illicit drugs are neuropsychiatric disorders defined by a compulsion to use these substances, loss of control, and continuous abuse despite multiple adverse medicolegal consequences.1 Some patients who are addicted to these substances, including cocaine and methamphetamine, suffer from cognitive decrements that might impact their activities of daily living.2 Despite several decades of studies in animal models, there still exists a great need to develop a theoretical framework that might serve to explain the manner by which certain drugs came to severely modulate the behaviors of certain individuals. It is likely that there are important genetic and environmental influences on the development of addiction.3 Nevertheless, more experiments remain to be conducted in order to clarify the basic substrates of addiction. Several recent articles have indeed provided ample evidence for a role for epigenetic changes in response to acute and repeated exposure to various drugs of abuse including psychostimulants such as cocaine and methamphetamine.3–5 Thus, the purpose of this article is to review recent data on the role of epigenetic mechanisms in addiction. GENERAL EPIGENETIC MECHANISMS IN BIOLOGY

Epigenetics refers to heritable transcriptional changes that are not the result of modified DNA sequences. These are multifactorial and include alterations in the accessibility to DNA because of differential packaging and potential changes in translational processes. This definition may also include meiotically and mitotically inherited transcriptional alterations that are not DNA-encoded.6 Epigenetic alterations include modifications of histones present in chromatin, DNA methylation, and DNA hydroxymethylation.7 In eukaryotic cells, DNA is packaged in chromatin, whose basic unit is the nucleosome. The nucleosome is made of four core histones: histones H2A, H2B, H3, and H4 that form an octamer (two of each histone) surrounded by 146 bp of DNA.8

Histone tails possess lysine residues that can be reversibly acetylated or deacetylated by respective histone acetyltransferases (HATs) or by histone deacetylases (HDACs).9,10 Other histone modifications that can impact gene expression include methylation, phosphorylation, and ubiquitylation (Table 1). These changes promote alterations in gene expression by modifying chromatin conformation and enabling or inhibiting recruitment of regulatory factors onto DNA sequences.11 HISTONE MODIFICATIONS

There are several classes of HATs, HDACs, and lysine methyltransferases (KMTs). Classically, the HATs belong to different classes of proteins that include GNAT (general control nonrepressible 5 [GCN5]-related N-acetyltransferases), cyclic AMPresponsive element binding (CREB)-binding protein (CBP)/ p300, and MYST (a name derived from MOZ [MYST3, monocytic leukemia zinc-finger protein], YBF2, SAS2 [something about silencing 2], and TIP60 [60 kDa trans-acting regulatory protein of HIV-type 1 (tat)-interaction proteins] subclasses.12 The GNAT subfamily of HATs is the largest group and includes histone acetyltransferase 1 (HAT1), GCN5, p300/CBP associated factor (PCAF), elongation protein 3 (ELP3), establishment of cohesion 1 homolog 1 (ESCO1), ESCO2, and others.13 HDACs are important epigenetic enzymes that remove acetyl groups from lysine residues on histone and nonhistone proteins.11 There are 18 HDACs that are further divided into four classes on the basis of sequence similarities.11 These include Class I (HDAC1, HDAC2, HDAC3, and HDAC8), Class IIA (HDAC4, HDAC5, HDAC7, and HDAC9), Class IIB (HDAC6 and HDAC10), Class III (Sirtuins1–7), and Class IV (HDAC11) HDACs.11 Class I, II, and IV HDACs are Zn21-dependent enzymes, whereas Class III HDACs are nicotinamide adenine dinucleotide (NAD)1-dependent enzymes.

1

Molecular Neuropsychiatry Research Branch, NIH/NIDA Intramural Research Program, National Institutes of Health, Baltimore, Maryland, USA. Correspondence: JL Cadet ([email protected]) Received 2 December 2015; accepted 26 January 2016; advance online publication 3 February 2016. doi:10.1002/cpt.345

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REVIEWS Table 1 Posttranslational modifications found on histones Chromatin modifications

Residues

Functions

Acetylation

Lysine

Repair, replication, and transcription

ADP-ribosylation

Glutamate, lysine

Repair, replication, and transcription

Deimination

Arginine

Transcription

Methylation

Lysine

Repair and transcription

Methylation

Arginine

Transcription

Phosphorylation

Serine, threonine

Condensation and transcription

Sumoylation

Lysine

Transcription

Ubiquitylation

Lysine

Repair and transcription

KMTs are a large family of proteins that are involved in mono-, di-, and trimethylation of specific lysine residues on histones.14 These include KMT1A (SUV39H1), KMT1B (SUV39H2), KMT1C (G9a), KMT2A (MLL1), KMT2B (MLL2), KMT3A (SET2), KMT3B (NSD1), among others.8,12 Although histone methylation is associated, for the most part, with decreased gene expression, methylation of histone H3K4 has been shown to lead to increased transcriptional activity.12,15 Moreover, several classes of lysine demethylases (KDMs) exist to counteract the effects of the KMTs.12,16 These include KDM1 (LSD1), KDM2A (JHDM1a), KDM2B (JHM1b), KDM3A (JHDM2a), KDM3B (JHDM2b), KDM4A (JMJD2A), KDM4B (JMJD2B), and others.12 Some of these chromatin-modifying enzymes have been shown to play important roles in biology and medicine17 and may therefore serve as important targets for the pharmacological treatment of human diseases, including addiction. DNA METHYLATION AND DEMETHYLATION

DNA methylation is a relatively stable mark characterized by a covalent modification at the 5-position of cytosine to form 5-methylcytosine. DNA methylation is mediated by the enzymes DNA methyltransferases (DNMT), that include DNMT1, DNMT3A, and DNMT3B.18 Specificity of substrates and different patterns of activation may be important in directing DNMT involvement in various pathological states including medical, neurological, and psychiatric disorders.19 DNA hydroxymethylation consists of the addition of a hydroxyl group to 5-methylcytosine to form hydroxymethylcytosine.7 These reactions are mediated by the three ten-eleven-translocation (TET1, TET2, and TET3) enzymes that are abundant in the brain.7 DNA hydroxymethylation has also been shown to participate in processes involved in cancers, neurodevelopment, and neurological disorders.20 ROLE OF EPIGENETICS IN PSYCHOSTIMULANT EXPOSURE

The abuse of cocaine and amphetamine-like compounds has remained at epidemic proportions despite attempts to curb their use through legal means. In addition, many years of research that have focused on the effects of these drugs on neurotransmitter systems have not helped to develop specific agents that can be used as preventive measures or as treatment of repeated binges. In fact, although those psychostimulants can facilitate transmission at monoaminergic synapses, that knowledge has yet to produce CLINICAL PHARMACOLOGY & THERAPEUTICS | VOLUME 99 NUMBER 5 | MAY 2016

pharmacological agents that have helped patients to remain abstinent. Recently, however, the presence of enduring effects of these drugs, even after long periods of abstinence, have led investigators to refocus attention on the possibility that these drugs may cause persistent molecular events that are engendered by long-lasting epigenetic, transcriptional, and translational effects on brain systems.3 The rest of this article will discuss how epigenetic events may be involved in the manifestation of addiction to these psychostimulants. Cocaine

Cocaine is a drug of abuse whose chronic use is associated with neurological and psychiatric impairments.21 The effects of cocaine addiction are known to involve activation of dopaminergic systems located in various brain regions by blocking the reuptake of dopamine (DA) into nerve terminals and to increase DA levels in synaptic clefts. The increase in DA levels leads to augmented stimulation of various DA receptors located in brain circuits that subsume rewards. These receptors include D1- and D2like proteins.22 Interestingly, a role for DA receptors was supported by the demonstration that DA D1 knockout mice failed to self-administer cocaine,23 whereas D3 knockout mice were reported to show increased cocaine self-administration.24 Because of the potential involvement of DA receptors, several studies have investigated the possible participation of DA genetics in addiction, with some studies reporting diverse roles of DArelated proteins in the manifestation of addictive disorders. These observations have prompted investigations of epigenetic effects on DA receptors. It has been reported that cocaine administration can produce increased histone H3 acetylation on the gene encoding DA D1-interacting protein, increased H4 acetylation at the DRD3 gene, and increased fosB enrichment of both DA D1A and D5 sequences.25 These results suggest that chronic cocaine may have a significant impact on epigenetic markers associated with DA receptors and suggest the possibility that some of the data on cocaine-induced changes in receptor expression and/ or functions26 may be mediated by epigenetic processes. Other studies have investigated the role of HATs in cocaine addiction. HATs are intimately involved in the regulation of gene expression by increasing histone acetylation.27 It was thus of interest to investigate the possibility that cocaine-induced effects on gene expression might be regulated by histone acetylation/ 503

REVIEWS deacetylation processes. Towards that end, Kumar et al.28 investigated the effects of acute and chronic cocaine administration on histone modifications at the promoter of c-fos, fosB, and BDNF in the rat striatum by using chromatin immunoprecipitation assays. They reported that acute, but not chronic, cocaine produced increased H3 phosphoacetylation and H4 acetylation at the c-fos promoter. Acute cocaine caused similar changes in histone acetylation at the fosB promoter. Interestingly, chronic cocaine also caused increased acetylation of H3, but not of H4, at the fosB promoter. Chronic, but not acute, cocaine also resulted in increased H3 acetylation at the BDNF promoter, an effect that was seen even after 1 week of withdrawal from cocaine.28 These long-lasting epigenetic effects of cocaine may play important roles in maintaining a state that is permissive to cue-induced craving in humans29 and animals.30 Levine et al.31 also investigated the role of histone acetylation and of the HAT, CBP, in cocaine-induced fosB mRNA expression and reported that these changes were mediated by increased binding of CBP and of histone H4 on the fosB promoter. The authors also showed that cocaine-induced changes were attenuated in mice that were CBP-haploinsufficient. Interestingly, there were no significant differences in baseline locomotor activity between mutant and control mice, whereas the mutant mice decreased behavioral sensitization to repeated cocaine administration. Malvaez et al.32 also reported that acute and chronic cocaine administration increased the acetylation of histone H3 at lysine 14 (H3K14Ac) in the nucleus accumbens (NAc) of wildtype mice. Consistent with the results of Levine et al.,31 cocaine-induced acetylation was significantly attenuated in mice with CBP deletion.32 Those authors also reported that acute cocaine increased binding of CBP at the c-fos promoter and increased c-fos expression in wildtype mice but attenuated responses in CBP knockout mice.32 Importantly, bilateral deletion of CBP in the NAc also attenuated cocaine-induced conditioned place preference (CPP), a measure of the rewarding effects of the drug. These effects of manipulating CBP may be region-dependent since Madsen et al.33 reported that deletion of CBP in the mouse striatum increased sensitivity to cocaine and amphetamine, in contrast to the observations obtained from manipulation of CBP in the NAc.32 In addition to HATs, HDACs are also involved in the regulation of gene transcription.34 Therefore, it was important to test the possibility that the different HDAC classes might also be involved in animal models of addiction by using pharmacological inhibitors or by virus-induced changes in the expression of HDACs. Schroeder et al.35 showed that pretreatment of mice with the broad-spectrum class I HDAC inhibitor, sodium butyrate, together with a dopamine D1 agonist caused increased cocaine-induced locomotion. The treatment also increased TH and BDNF mRNA expression and associated H3 hypoacetylation at the promoters of these genes. The H3 hypoacetylation is unexpected because histone hypoacetylation is thought to lead to decreased gene expression. Romieu et al.36 also reported that HDAC inhibitors can decrease cocaine self-administration. Related to the role of class I HDAC in addiction, Kennedy et al.37 reported that HDAC1 knockdown caused a reduction of cocaine-induced behavioral sensitization. HDAC2 and HDAC3 504

knockdown was without any behavioral consequences. In contrast, Host et al.38 reported that cocaine self-administration increased HDAC2 expression. It is, however, not clear to what extent increased HDAC2 was involved in regulating the behavioral effects of cocaine, since this was not specifically determined in that study. It is also not clear how this increase might be involved in regulating cocaine-associated changes in gene expression because HDAC2 can bind to genes that show either increased or decreased expression.39 A later study reported that cocaine-induced HDAC2 expression can be attenuated by increased expression of the cyclic GMP-dependent protein kinase.40 This was followed by another article that documented a specific role for HDAC2 in cocaine self-administration.41 The participation of another class I HDAC, HDAC3, has also been documented in models of cocaine addiction.42,43 Using a selective HDAC3 inhibitor called RGFP966, Malvaez et al.42 demonstrated that the drug promoted extinction of cocaine CPP. RGFP966 also prevented reinstatement of cocaine-seeking behavior. The drug also increased acetylation of histone H4 at lysine 8 (H4K8Ac) in the infralimbic cortex but not in the NAc and hippocampus and increased H3K14Ac in all three brain regions.42 Moreover, there was decreased HDAC3 binding at the c-fos promoter.42 The results of that study are supported by the report that focal deletion of HDAC3 in the mouse NAc also facilitated cocaine CPP.43 In addition to its effects on class I HDACs, cocaine has also been shown to regulate the expression of class II HDACs. For example, Wang et al.44 reported that HDAC4 overexpression was able to attenuate the motivation to self-administer cocaine. Chronic cocaine injection also produced increased HDAC5 phosphorylation at 30 minutes, but not at 24 hours, after the last injection.45 This was accompanied by export of HDAC5 from the cell nucleus to the cytoplasm. Moreover, increased HDAC5 expression in the NAc via a viral vector led to attenuated cocaine CPP, indicating a decrease in the rewarding effect of the drug. The authors also showed that HDAC5 knockout mice, pre-exposed to chronic cocaine, exhibited increased responses to the rewarding effects of cocaine.45 Interestingly, cocaine self-administration also caused decreased nuclear localization of HDAC5.38 Moreover, injections of cocaine increased phosphorylation of HDAC5, followed by shuttling of phosphorylated HDAC5 into the cytoplasm.46 The results of these three studies, however, are not consistent with the report by Taniguchi et al.,47 who observed cocaine-induced HDAC5 dephosphorylation and increased HDAC5 nuclear accumulation in the mouse striatum. The reasons for these dichotomous results are not clear at present and will await more experiments to provide a detailed explanation for the role of HDAC5 phosphorylation and of its regional and subcellular distribution in modulating cocaine addiction. Interestingly, the role of multiple zinc-dependent HDACs in cocaine-induced behavioral effects was further supported by the demonstration that cocaine self-administration increased the expression of the class IV HDAC, HDAC11,38 which is highly expressed in the brain48 and shows great similarities to class I HDACs.49 Class III HDACs, the sirtuins, also participate in the effects of cocaine in the brain. Renthal et al.25 used chromatin VOLUME 99 NUMBER 5 | MAY 2016 | www.wileyonlinelibrary/cpt

REVIEWS Table 2 Summary of epigenetic alterations caused by cocaine use Chronic: IP (20 mg/kg; 1 injection/ day) for 7 days

Striatum: Histone H3 hyperacetylation at the Bdnf promoter

Self-administration (15 days; 4-h session)

Striatum: Histone H3 hyperacetylation at the Bdnf promoter

Self-administration (3-4 weeks; 4-h session)

NAc shell: Histone H3 hyperacetylation at the Bdnf promoter

(44)

CamkIIa


NAc shell: Histone H3 hyperacetylation at the CamkIIa gene associated with drug motivated behavior

(44)

Cart

Chronic: IP (20 mg/kg; 1 injection/ day) for 7 days

NAc: Histone H3 hyperacetylation at the Cart promoter

(25)

CBP


NAc shell: Histone H3 hyperacetylation at the CBP promoter

(44)

Cdk5


Striatum: Histone H3 hyperacetylation at the Cdk5 promoter

(28)


Striatum: Histone H3 hyperacetylation at the Cdk5 promoter


NAc shell: Histone H3 hyperacetylation at the Cdk5 promoter

(44)

Acute: IP (20 mg/kg)

Striatum: Histone H4 hyperacetylation at the c-fos promoter

(28)


NAc: Histone H3 hyperacetylation at the c-fos promoter

(25)

Acute IP (10mg/kg) Chronic IP (10mg/kg; 1 injection/day) for 5 days

NAc: Histone H3 hyperacetylation and increased binding of CBP to c-fos promoter leads to increased c-fos expression

(32)


NAc: Increased Dnmt3a and Dnmt3b expression

(55)


PFCtx: Increased Dnmt3a and Dnmt3b expression

(53)

Bdnf

c-fos

Dnmt3a Dnmt3b

(28)

CPu: Transient Increase in Dnmt3a and Dnmt3b expression Egr1


NAc shell: Histone H4 hyperacetylation at the Egr1 promoter

(44)

FosB


Striatum: Histone H4 hyperacetylation and increased binding of CBP to FosB promoter leads to FosB expression

(31)


Striatum: Histone H4 hyperacetylation at the FosB promoter

(28)





Acute: IP (15 mg/kg) Chronic: IP (15 mg/kg; 1 injection/day) for 7 days

NAc: Hypomethylation at the FosB promoter decreased MeCP2 binding and increased FosB expression

(55)


NAc shell: Histone H3 hyperacetylation at the FosB promoter

(44)


NAc shell: Histone H3 hyperacetylation at the GluA2 promoter

(44)

GluA2

Table 2 Continued on next page

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GluN2a


NAc shell: Histone H3 hyperacetylation at the GluN2a promoter

(44)

GluN2b


NAc shell: Histone H3 hyperacetylation at the GluN2b promoter

HDAC2 HDAC5 HDAC11


NAc: Increased phospho HDAC5 protein

(45)

Self-administration (4 days; 2h session)

CgCtx, NAc, CPu: Increased HDAC2 mRNA and protein

(38)

Decreased HDAC5 and increased HDAC11 protein Acute: IP (20 mg/kg)

Striatum: Increased phospho-HDAC5

(46)


Striatum: Transient decreases in phosphorylation of HDAC5

(47)


Striatum: Increased MeCP2 protein

(52)


CgCtx, NAc, CPu: Increased MeCP2 mRNA and protein

(38)


NAc: Increased phospho-MeCP2 protein level

(54)


CgCtx, NAc, CPu: Increased MEF2c mRNA expression

(38)


Striatum: Increased MEF2c through activation of SIK1 and phosphorylation of HDAC5

(46)

PP1Cb


PFCtx, CPu: Hypermethylation and increased MeCP2 binding at PP1Cb promoter lead to decreased mRNA and protein levels

(53)

Psd95


NAc shell: Histone H3 and H4 hyperacetylation at the Psd95 promoter

(44)

SIRT1 SIRT2


NAc: Histone H3 hyperacetylation at Sirt1 and Sirt2 gene promoters

(25)


NAc: Increased Sirt1 and Sirt2 mRNA expression

(50)

MeCP2

MEF2c

Abbreviations: Cpu, caudate putamen; CgCtx, cingulate cortex; IP, intraperitoneal injection; Nac, nucleus accumbens; PFCtx, prefrontal cortex.

immunoprecipitation (ChIP) to investigate the effects of repeated cocaine injections on genome-wide distribution of acetylated histones the NAc of mice. Pan-acetylated histones H3 and H4 were used in their experiments. They found that chronic cocaine increased binding of acetylated H3 and H4 at the promoters of a large number of genes. Only 15% of the genes showed increased H3 and H4 acetylation. Genes of interest were Sirt1 and Sirt2.25 The authors further tested the potential participation of the Sirts in cocaine-induced behavioral effects by using the Sirt agonist, resveratrol, which enhanced cocaine CPP, whereas the Sirt inhibitor, sirtinol, suppressed CPP. Increasing Sirt1 expression in the mouse NAc increased cocaine CPP and BDNF and CBP expression, whereas Sirt1 deletion caused decreased cocaine CPP.50 Increasing Sirt2 expression also increased cocaine CPP.50 In addition to causing changes in the expression of HDACs, HDAC phosphorylation, and histone acetylation, cocaine can also 506

increase methyl-CpG binding protein 2 (MeCP2) expression.51–53 Specifically, cocaine self-administration for 7 days increased MeCP2 protein expression in the dorsal striatum.52 In addition, MeCP2 knockdown decreased cocaine self-administration in rats by regulating BDNF expression. Recently, cocaine self-administration was also reported to increase MeCP2 phosphorylation in the NAc and that MeCP2 phosphorylation was involved in limiting cocaine intake.54 Cocaine has also been shown to cause changes in DNA methylation in the brain. For example, Anier et al.55 reported that acute, but not chronic, cocaine increased DNMT3A and DNMT3B mRNA expression in the NAc. The authors also reported that both acute and chronic cocaine caused DNA hypermethylation and increased MeCP2 binding at the promoter of protein phosphatase-1 catalytic subunit (PP1c), changes that were associated with decreased PP1C mRNA expression.55 Pol Bodetto et al.53 also reported that repeated cocaine increased DNA methylation at the PP1Cbeta gene, which was associated with decreased PP1Cbeta mRNA and VOLUME 99 NUMBER 5 | MAY 2016 | www.wileyonlinelibrary/cpt

REVIEWS Table 3 Summary of epigenetic alterations caused by amphetamine/ methamphetamine use Bdnf

Self-administration: METH (8 days; 15-h session)

Striatum: Increased binding of phospho CREB at the Bdnf promoter

(5)

CamkIIa

Chronic: SC METH (1 mg/kg) once every 2 days for 6 days

Limbic forebrain: Histone H3 hyperacetylation at the CamkIIa promoter

(60)

Cdk5


Limbic forebrain: Histone H3 and H4 hyperacetylation at the Cdk5 promoter regions

(60)

c-fos

Acute: IP METH (5 mg/kg)

Striatum: Increased Histone H4K5Ac binding at the c-fos promoter

(62)


Striatum: Increased binding of phospho CREB at the c-fos promoter

(5)

CoREST

Chronic: IP METH (Escalating dose- 0.5–3 mg/kg) for 2 weeks

Striatum: Increased CoREST protein

(4)

Creb


Limbic forebrain: Histone H3 hyperacetylation at the Creb promoter

(60)

Dlg4


Limbic forebrain: Histone H3 hyperacetylation at the Dlg4 promoter

(60)

Egr2


Striatum: Increased Histone H4K5Ac binding at the Egr2 promoter

(62)

FosB


Striatum: Increased binding of phospho CREB at the fosB promoter

(5)

DNMT1


Striatum: Increased DNMT1 protein

(4)


NCau, Cerebellum: Increased DNMT1 expression

(72)

Chronic: IP METH (4 mg/kg) once daily for 21 days

NAc: Increased DNMT1 expression


Limbic forebrain: Histone H3 hyperacetylation at the GluA1 promoter

(60)


Striatum: Increased binding of repressor elements CoREST, MeCP2 and HDAC2 on GluA1 and GluA2 promoters leads to decreased GluA1 and GluA2 expression.

(4)

Chronic: IP METH (Escalating dose-0.5–3 mg/kg) for 2 weeks

Striatum: Increased binding of repressor element REST and HDAC1 on GluN1 promoter leads to decreased GluN1 expression

(4)


Limbic forebrain: Histone H3 hyperacetylation at the GluN2a and GluN2b promoter regions

(60)


NAc: Transient decrease in HDAC1 and increase in HDAC2 protein levels

(61)

Acute: SC (5 mg/kg)

PFCtx: Decreased HDAC1 and HDAC2 expression

(66)

Chronic: IP METH (Escalating dose0.5–3 mg/kg) for 2 weeks

Striatum: Increased HDAC1 and HDAC2 protein levels

(4)

Acute IP AMPH (3 mg/kg)

NAc: Transient increase in phospho MeCP2

(51)


Striatum: Increased MeCP2 protein

(4)

Chronic: SC METH (0.5–2 mg/kg) once every 2 days for 6 days

Limbic forebrain: Histone H3 hyperacetylation at the Nrxn promoter

(60)

GluA1 GluA2

GluN1 GluN2a GluN2b

HDAC1 HDAC2

MeCP2

Nrxn

Table 3 Continued on next page


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REST


Striatum: Increased REST protein

(4)

SIRT1 SIRT2


Striatum: Increased SIRT1 and SIRT2 protein levels

(4)

Syp


Limbic forebrain: Histone H3 hyperacetylation at the Syp promoter

(60)


Striatum: Increased binding of phospho CREB at the Syp promoter

(5)

Abbreviations: IP, intraperitoneal injection; Nac, nucleus accumbens; Ncau, nucleus caudatus; PFCtx, prefrontal cortex; SC, subcutaneous.

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Epigenetic effects of repeated METH exposure GluA1 mRNA GluA2 mRNA Histone H4 hypoacetylation SirT2 CoREST

The abuse of amphetamine-like psychostimulants has been a major problem in the world for many years, with its prevalence outpacing that of cocaine. There are, nevertheless, only a few studies investigating the epigenetic effects of these drugs when compared to a number of studies investigating the effects of cocaine. There is, however, a wealth of information on the neurodegenerative effects of the drug.59 Nevertheless, experiments have recently been conducted in attempts to elucidate potential epigenetic effects of these groups of drugs.3 The fact that the behavioral effects of psychostimulants can last for long periods of time suggests that epigenetic alterations may be involved3 (Table 3). This suggestion was supported initially by the report that histone acetylation was involved in the behavioral effects of methamphetamine (METH).60 The authors of that article reported that METH CPP was accompanied by increased histone H3 acetylation. Martin et al.61 also reported that METH increased the abundance of acetylated H4K5 but decreased the abundance of acetylated H3K9 and H3K18 in the NAc. METH administration also produced decreased HDAC1 expression but increased HDAC2 levels in the rats NAc. Moreover, METH upregulated Egr2 and c-fos expression via increased H4K5Ac binding at the promoters of these genes.62 In a subsequent article, Harkness et al.63 also reported that acute METH increased H4 acetylation in the striatum. It is thus possible that METH-induced increased H4 acetylation may be secondary to increased CBP expression because increased CBP expression is associated with increased histone acetylation.64 CBP was also reported to mediate cocaineinduced histone acetylation.31 The role of HDAC1 and HDAC2 in the acute effects of amphetamine-like drugs is also supported by a behavioral study that an HDAC1/2 inhibitor, Cp60, was able to inhibit amphetamine (AMPH)-induced locomotion.65 In a recent report, Li et al.66 also provided evidence

METH METH METH METH

HDAC2

Amphetamine and methamphetamine

that a single METH injection decreased HDAC1 and HDAC2 in the prefrontal cortex. The epigenetic effects of repeated METH exposure have also been investigated in a model of increasing METH doses over weeks4 (see Figure 1). The authors reported that chronic METH produced decreased mRNA and protein expression of glutamate AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid) receptors in the dorsal striatum. They also found that the METH injections increased HDAC1 and HDAC2 protein expression in the dorsal striatum, with associated hypoacetylation of histone H4 acetylation at lysines 4, 8, 12, and 16.4 The repeated METH injections also increased the expression of repressor element-1 silencing transcription factor,67 corepressor 1 that usually exists in complexes with HDACs.68 ChIP / polymerase chain reaction (PCR) also showed that METH decreased the enrichment of acetylated histone H4 on AMPA receptors and of histone deacetylase 2 enrichment onto GluA1 and GluA2 gene sequences. Of note is that valproic acid, a histone deacetylase inhibitor, which is used in the treatment of various neuropsychiatric disorders,69 was able to block METHinduced decreased expression of AMPA receptor subunits. In contrast, chronic METH decreased HDAC2 expression in the

CoREST

protein expression. These results are consistent with a recent article that has documented substantial changes in DNA methylation in the rat NAc at 1 and 30 days of withdrawal from cocaine selfadministration.56 Importantly, methyl supplementation was able to attenuate cocaine-seeking behaviors in a DNA methylationdependent fashion.57 Taken together, these results indicate that cocaine can produce genome-wide changes in DNA methylation in the rodent brain. More generally, cocaine-induced epigenetic changes (Table 2) may serve as substrates for the large-scale changes in gene expression observed in various models of cocaine administration.25,58

M M

M

M M

M

Increasing doses of METH

Figure 1 Schematic model showing METH-induced epigenetic effects in the rat dorsal striatum. METH injections increased the expression of MeCP2, HDAC2, CoREST, and SirT2 in the striatum. The scheme suggests that these proteins participate in a repressor complex that resulted in decreased enrichment of acetylated histone H4 on GluA1 and GluA2 DNA sequences, with consequent decreased mRNA transcription. This model also supports the idea that the use of HDAC inhibitors in humans might prevent the chronic effects of psychostimulants in the brains of addicted patients. VOLUME 99 NUMBER 5 | MAY 2016 | www.wileyonlinelibrary/cpt

REVIEWS prefrontal cortex,66 suggesting that neuroscientists and pharmacologists need to be cognizant of the brain regions being investigated when studying models of addiction or drug effects because of the diversity of responses that drugs can cause. Chronic METH also reduced the expression of the class II HDAC, HDAC4, without any effect on HDAC5.66 Both HDAC4 and HDAC5 expression was decreased when measured at 7 days of withdrawal from METH treatment. Chronic METH also increased expression of two class III HDACs, SIRT1 and SIRT2, in the dorsal striatum.4 Methylation of histone H3 at lysine 4 (H3K4 me2/3) also appears to be important in methamphetamine-induced CPP because increased methylation is associated with increased CPP, whereas decreased methylation is related to decreased CPP.70 METH-induced behavioral sensitization is also accompanied by increased H3K4me3.71 Krasnova et al.5 also reported that METH self-administration was associated with an increased abundance of trimethylated H3K4 (H3K4me3) in the rat dorsal striatum. However, the extent to which these changes are involved in mediating the effects of METH self-administration remains to be clarified. In any case, when taken together, these results suggest that amphetamine-like drugs can impact brain functions by causing multiple posttranslational histone modifications. METH administration can also cause changes in DNA methylation since the drug increases DNMT1 expression in the brain.4,72 Withdrawal from chronic AMPH treatment is associated with increased DNA methylation in the NAc, prefrontal cortex, and orbitofrontal cortex.73 The significant increases in DNA methylation are, in part, consistent with our recent report of global decreases in gene expression measured by microarray analysis at 1 month of withdrawal from METH self-administration.74 Mice exposed to METH in utero show differentially DNA methylation in their hippocampi as adults.75 The results on DNA methylation are consistent with observations that both AMPH and METH can impact MeCP2 expression in the brain.4,51 Interestingly, MeCP2 overexpression in the NAc was shown to attenuate the rewarding effects of AMPH.51 Mice that express a nonphosphorylated MeCP2 protein exhibited increased AMPH-induced behavioral sensitization.54 These results suggest that modifications of histones and nonhistone proteins can impact amphetamineinduced behavioral changes in models of drug addiction. Nevertheless, much more research needs to be done to understand the relative meaning of METH-induced changes in DNA methylation and how these alterations might influence gene expression and brain function during the transition from recreational drug use to full-blown addiction. CONCLUDING REMARKS AND THERAPEUTIC IMPLICATIONS

Reductionist approaches have led to a number of discoveries related to the molecular and cellular substrates of drug-induced plasticity in the brain. In the case of addiction to psychostimulant, the theoretical reliance on the demonstrated biochemical, molecular, and epigenetic effects of contingent and noncontingent administration of these substances promises to help us to develop more rational therapeutic approaches. The findings that CLINICAL PHARMACOLOGY & THERAPEUTICS | VOLUME 99 NUMBER 5 | MAY 2016

these drugs can significantly influence the expression of several HATs and HDACs in the brain suggest that more effort should be spent trying to develop brain-specific agents that would avert the peripheral effects of drugs presently being used as anticancer therapeutics. Because both cocaine and the amphetamine-like compounds also appear to influence DNA methylation processes in the brain, the possibility also exists that methylation mechanisms need to be more thoroughly investigated in models of addiction. It is hoped that an expanded portfolio for antiaddiction epigenetic agents will help to bolster our pharmacological arsenals against drug addiction. ACKNOWLEDGMENT This work was supported by funds of the Intramural Research Program of the DHHS/NIH/NIDA. CONFLICT OF INTEREST The authors declare no conflicts of interest. Published 2016. This article is a US Government work and is in the public domain in the USA.

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Cadet, J.L., Bisagno, V. & Milroy, C.M. Neuropathology of substance use disorders. Acta Neuropathol. 127, 91–107 (2014). Cadet, J.L. & Bisagno, V. The primacy of cognition in the manifestations of substance use disorders. Front. Neurol. 4, 189 (2013). Cadet, J.L. Epigenetics of stress, addiction, and resilience: therapeutic implications. Mol. Neurobiol. 53, 545–563 (2016). Jayanthi, S. et al. Methamphetamine downregulates striatal glutamate receptors via diverse epigenetic mechanisms. Biol. Psychiatry 76, 47–56 (2014). Krasnova, I.N. et al. CREB phosphorylation regulates striatal transcriptional responses in the self-administration model of methamphetamine addiction in the rat. Neurobiol. Dis. 58, 132–143 (2013). Kota, S.K. & Feil, R. Epigenetic transitions in germ cell development and meiosis. Dev. Cell 19, 675–686 (2010). Hill, P.W., Amouroux, R. & Hajkova, P. DNA demethylation, Tet proteins and 5-hydroxymethylcytosine in epigenetic reprogramming: an emerging complex story. Genomics 104, 324–333 (2014). Kouzarides, T. Chromatin modifications and their function. Cell 128, 693–705 (2007). Schneider, A. et al. Acetyltransferases (HATs) as targets for neurological therapeutics. Neurotherapeutics 10, 568–588 (2013). Seto, E. & Yoshida, M. Erasers of histone acetylation: the histone deacetylase enzymes. Cold Spring Harb. Perspect. Biol. 6, a018713 (2014). Sterner, D.E. & Berger, S.L. Acetylation of histones and transcriptionrelated factors. Microbiol. Mol. Biol. Rev. 64, 435–459 (2000). Allis, C.D. et al. New nomenclature for chromatin-modifying enzymes. Cell 131, 633–636 (2007). Marmorstein, R. & Roth, S.Y. Histone acetyltransferases: function, structure, and catalysis. Curr. Opin. Genet. Dev. 11, 155–161 (2001). Black, J.C., Van Rechem, C. & Whetstine, J.R. Histone lysine methylation dynamics: establishment, regulation, and biological impact. Mol. Cell 48, 491–507 (2012). Shilatifard, A. Molecular implementation and physiological roles for histone H3 lysine 4 (H3K4) methylation. Curr. Opin. Cell. Biol. 20, 341–348 (2008). Natoli, G., Testa, G. & De Santa, F. The future therapeutic potential of histone demethylases: a critical analysis. Curr. Opin. Drug Discov. Dev. 12, 607–615 (2009). Falkenberg, K.J. & Johnstone, R.W. Histone deacetylases and their inhibitors in cancer, neurological diseases and immune disorders. Nat. Rev. Drug Discov. 13, 673–691 (2014). 509

REVIEWS 18. Kinney, S.R. & Pradhan, S. Regulation of expression and activity of DNA (cytosine-5) methyltransferases in mammalian cells. Prog. Mol. Biol. Transl. Sci. 101, 311–333 (2011). 19. Baets, J. et al. Defects of mutant DNMT1 are linked to a spectrum of neurological disorders. Brain 138, 845–861 (2015). 20. Sherwani, S.I. & Khan, H.A. Role of 5-hydroxymethylcytosine in neurodegeneration. Gene 570, 17–24 (2015). 21. Nnadi, C.U., Mimiko, O.A., McCurtis, H.L. & Cadet, J.L. Neuropsychiatric effects of cocaine use disorders. J. Natl. Med. Assoc. 97, 1504–1515 (2005). 22. Yan, Y., Newman, A.H. & Xu, M. Dopamine D1 and D3 receptors mediate reconsolidation of cocaine memories in mouse models of drug self-administration. Neuroscience 278, 154–164 (2014). 23. Caine, S.B. et al. Lack of self-administration of cocaine in dopamine D1 receptor knock-out mice. J. Neurosci. 27, 13140–13150 (2007). 24. Song, R., Zhang, H.Y., Li, X., Bi, G.H., Gardner, E.L. & Xi, Z.X. Increased vulnerability to cocaine in mice lacking dopamine D3 receptors. Proc. Natl. Acad. Sci. U. S. A. 109, 17675–17680 (2012). 25. Renthal, W. et al. Genome-wide analysis of chromatin regulation by cocaine reveals a role for sirtuins. Neuron 62, 335–348 (2009). 26. Ramoa, C.P., Doyle, S.E., Lycas, M.D., Chernau, A.K. & Lynch, W.J. Diminished role of dopamine D1-receptor signaling with the development of an addicted phenotype in rats. Biol. Psychiatry 76, 8–14 (2014). 27. Choudhary, C., Weinert, B.T., Nishida, Y., Verdin, E. & Mann, M. The growing landscape of lysine acetylation links metabolism and cell signalling. Nat. Rev. Mol. Cell. Biol. 15, 536–550 (2014). 28. Kumar, A. et al. Chromatin remodeling is a key mechanism underlying cocaine-induced plasticity in striatum. Neuron 48, 303–314 (2005). 29. Vorspan, F. et al. Self-reported cue-induced physical symptoms of craving as an indicator of cocaine dependence. Am. J. Addict. 24, 740–743 (2015). 30. Pickens, C.L., Airavaara, M., Theberge, F., Fanous, S., Hope, B.T. & Shaham, Y. Neurobiology of the incubation of drug craving. Trends Neurosci. 34, 411–420 (2011). 31. Levine, A.A., Guan, Z., Barco, A., Xu, S., Kandel, E.R. & Schwartz, J.H. CREB-binding protein controls response to cocaine by acetylating histones at the fosB promoter in the mouse striatum. Proc. Natl. Acad. Sci. U. S. A. 102, 19186–19191 (2005). 32. 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). 33. Madsen, H.B. et al. CREB1 and CREB-binding protein in striatal medium spiny neurons regulate behavioural responses to psychostimulants. Psychopharmacology 219, 699–713 (2012). 34. Chen, H.P., Zhao, Y.T. & Zhao, T.C. Histone deacetylases and mechanisms of regulation of gene expression. Crit. Rev. Oncog. 20, 35–47 (2015). 35. Schroeder, F.A. et al. Drug-induced activation of dopamine D(1) receptor signaling and inhibition of class I/II histone deacetylase induce chromatin remodeling in reward circuitry and modulate cocaine-related behaviors. Neuropsychopharmacology 33, 2981– 2992 (2008). 36. Romieu, P., Host, L., Gobaille, S., Sandner, G., Aunis, D. & Zwiller, J. Histone deacetylase inhibitors decrease cocaine but not sucrose self-administration in rats. J. Neurosci. 28, 9342–9348 (2008). 37. 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). 38. Host, L., Dietrich, J.B., Carouge, D., Aunis, D. & Zwiller, J. Cocaine self-administration alters the expression of chromatin-remodelling proteins; modulation by histone deacetylase inhibition. J. Psychopharmacol. 25, 222–229 (2011). 39. Wang, Z. et al. Genome-wide mapping of HATs and HDACs reveals distinct functions in active and inactive genes. Cell 138, 1019–1031 (2009). 40. Deschatrettes, E., Jouvert, P. & Zwiller, J. Overexpression of cyclic GMP-dependent protein kinase reduces MeCP2 and HDAC2 expression. Brain Behav. 2, 732–740 (2012). 41. Deschatrettes, E., Romieu, P. & Zwiller, J. Cocaine selfadministration by rats is inhibited by cyclic GMP-elevating agents: involvement of epigenetic markers. Int. J. Neuropsychopharmacol. 16, 1587–1597 (2013). 510

42. Malvaez, M. et al. HDAC3-selective inhibitor enhances extinction of cocaine-seeking behavior in a persistent manner. Proc. Natl. Acad. Sci. U. S. A. 110, 2647–2652 (2013). 43. 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). 44. Wang, L. et al. Chronic cocaine-induced H3 acetylation and transcriptional activation of CaMKIIalpha in the nucleus accumbens is critical for motivation for drug reinforcement. Neuropsychopharmacology 35, 913–928 (2010). 45. Renthal, W. et al. Histone deacetylase 5 epigenetically controls behavioral adaptations to chronic emotional stimuli. Neuron 56, 517–529 (2007). 46. Dietrich, J.B., Takemori, H., Grosch-Dirrig, S., Bertorello, A. & Zwiller, J. Cocaine induces the expression of MEF2C transcription factor in rat striatum through activation of SIK1 and phosphorylation of the histone deacetylase HDAC5. Synapse 66, 61–70 (2012). 47. Taniguchi, M., Carreira, M.B., Smith, L.N., Zirlin, B.C., Neve, R.L. & Cowan, C.W. Histone deacetylase 5 limits cocaine reward through cAMP-induced nuclear import. Neuron 73, 108–120 (2012). 48. Takase, K., Oda, S., Kuroda, M. & Funato, H. Monoaminergic and neuropeptidergic neurons have distinct expression profiles of histone deacetylases. PLoS One 8, e58473 (2013). 49. Gao, L., Cueto, M.A., Asselbergs, F. & Atadja, P. Cloning and functional characterization of HDAC11, a novel member of the human histone deacetylase family. J. Biol. Chem. 277, 25748–25755 (2002). 50. Ferguson, D. et al. Essential role of SIRT1 signaling in the nucleus accumbens in cocaine and morphine action. J. Neurosci. 33, 16088– 16098 (2013). 51. Deng, J.V., Rodriguiz, R.M., Hutchinson, A.N., Kim, I.H., Wetsel, W.C. & West, A.E. MeCP2 in the nucleus accumbens contributes to neural and behavioral responses to psychostimulants. Nat. Neurosci. 13, 1128–1136 (2010). 52. Im, H.I., Hollander, J.A., Bali, P. & Kenny, P.J. MeCP2 controls BDNF expression and cocaine intake through homeostatic interactions with microRNA-212. Nat. Neurosci. 13, 1120–1127 (2010). 53. Pol Bodetto, S., Carouge, D., Fonteneau, M., Dietrich, J.B., Zwiller, J. & Anglard, P. Cocaine represses protein phosphatase-1Cbeta through DNA methylation and methyl-CpG binding protein-2 recruitment in adult rat brain. Neuropharmacology 73, 31–40 (2013). 54. Deng, J.V. et al. MeCP2 phosphorylation limits psychostimulantinduced behavioral and neuronal plasticity. J. Neurosci. 34, 4519– 4527 (2014). 55. Anier, K., Malinovskaja, K., Aonurm-Helm, A., Zharkovsky, A. & Kalda, A. DNA methylation regulates cocaine-induced behavioral sensitization in mice. Neuropsychopharmacology 35, 2450–2461 (2010). 56. Massart, R. et al. Role of DNA methylation in the nucleus accumbens in incubation of cocaine craving. J. Neurosci. 35, 8042–8058 (2015). 57. Wright, K.N. et al. Methyl supplementation attenuates cocaineseeking behaviors and cocaine-induced c-Fos activation in a DNA methylation-dependent manner. J. Neurosci. 35, 8948–8958 (2015). 58. Krasnova, I.N. et al. Transcriptional responses to reinforcing effects of cocaine in the rat hippocampus and cortex. Genes Brain Behav. 7, 193–202 (2008). 59. Cadet, J.L. & Krasnova, I.N. Molecular bases of methamphetamineinduced neurodegeneration. Int. Rev. Neurobiol. 88, 101–119 (2009). 60. Shibasaki, M., Mizuno, K., Kurokawa, K. & Ohkuma, S. L-type voltagedependent calcium channels facilitate acetylation of histone H3 through PKCgamma phosphorylation in mice with methamphetamineinduced place preference. J. Neurochem. 118, 1056–1066 (2011). 61. Martin, T.A. et al. Methamphetamine causes differential alterations in gene expression and patterns of histone acetylation/hypoacetylation in the rat nucleus accumbens. PLoS One 7, e34236 (2012). 62. Cadet, J.L. et al. Genome-wide profiling identifies a subset of methamphetamine (METH)-induced genes associated with METH-induced increased H4K5Ac binding in the rat striatum. BMC Genomics 14, 545 (2013). 63. Harkness, J.H., Hitzemann, R.J., Edmunds, S. & Phillips, T.J. Effects of sodium butyrate on methamphetamine-sensitized locomotor activity. Behav. Brain Res. 239, 139–147 (2013). VOLUME 99 NUMBER 5 | MAY 2016 | www.wileyonlinelibrary/cpt

REVIEWS 64. Henry, R.A., Kuo, Y.M. & Andrews, A.J. Differences in specificity and selectivity between CBP and p300 acetylation of histone H3 and H3/ H4. Biochemistry 52, 5746–5759 (2013). 65. Schroeder, F.A. et al. A selective HDAC 1/2 inhibitor modulates chromatin and gene expression in brain and alters mouse behavior in two mood-related tests. PLoS One 8, e71323 (2013). 66. Li, H., Li, F., Wu, N., Su, R.B. & Li, J. Methamphetamine induces dynamic changes of histone deacetylases in different phases of behavioral sensitization. CNS Neurosci. Ther. 20, 874–876 (2014). 67. Robinson, T.E., Becker, J.B. & Presty, S.K. Long-term facilitation of amphetamine-induced rotational behavior and striatal dopamine release produced by a single exposure to amphetamine: sex differences. Brain Res. 253, 231–241 (1982). 68. Griffith, E.C., Cowan, C.W. & Greenberg, M.E. REST acts through multiple deacetylase complexes. Neuron 31, 339–340 (2001). 69. Cipriani, A., Reid, K., Young, A.H., Macritchie, K. & Geddes, J. Valproic acid, valproate and divalproex in the maintenance treatment of bipolar disorder. Cochrane Database Syst. Rev. 10, CD003196 (2013).


70. Aguilar-Valles, A. et al. Methamphetamine-associated memory is regulated by a writer and an eraser of permissive histone methylation. Biol. Psychiatry 76, 57–65 (2014). 71. Ikegami, D. et al. Epigenetic modulation at the CCR2 gene correlates with the maintenance of behavioral sensitization to methamphetamine. Addict. Biol. 15, 358–361 (2010). 72. Numachi, Y. et al. Methamphetamine alters expression of DNA methyltransferase 1 mRNA in rat brain. Neurosci. Lett. 414, 213–217 (2007). 73. Mychasiuk, R., Harker, A., Ilnytskyy, S. & Gibb, R. Paternal stress prior to conception alters DNA methylation and behaviour of developing rat offspring. Neuroscience 241, 100–105 (2013). 74. Cadet, J., Brannock, C., Jayanthi, S. & Krasnova, I. Transcriptional and epigenetic substrates of methamphetamine addiction and withdrawal: evidence from a long-access self-administration model in the rat. Mol. Neurobiol. 1–22 (2014). 75. Itzhak, Y., Ergui, I. & Young, J.I. Long-term parental methamphetamine exposure of mice influences behavior and hippocampal DNA methylation of the offspring. Mol. Psychiatry 20, 232–239 (2014).

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