The genetics of cognitive epigenetics.

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Neuropharmacology xxx (2014) 1e12

Contents lists available at ScienceDirect

Neuropharmacology journal homepage: www.elsevier.com/locate/neuropharm

Invited review

The genetics of cognitive epigenetics Q6

Tjitske Kleefstra a, Annette Schenck a, Jamie Kramer a, Hans van Bokhoven a, b, * a

Radboud University Medical Center, Department of Human Genetics, Nijmegen Center for Molecular Life Sciences (NCMLS), Nijmegen, The Netherlands Radboud University Medical Center, Department of Cognitive Neurosciences, Donders Institute for Brain, Cognition and Behaviour, Nijmegen, The Netherlands b

a r t i c l e i n f o

a b s t r a c t

Article history: Received 16 November 2013 Received in revised form 29 December 2013 Accepted 30 December 2013

Cognitive disorders (CDs) are a heterogeneous group of disorders for which the genetic foundations are rapidly being uncovered. The large number of CD-associated gene mutations presents an opportunity to identify common mechanisms of disease as well as molecular processes that are of key importance to human cognition. Given the disproportionately high number of epigenetic genes associated with CD, epigenetic regulation of gene transcription is emerging as a process of major importance in cognition. The cognate protein products of these genes often co-operate in shared protein complexes or pathways, which is reflected in similarities between the neurodevelopmental phenotypes corresponding to these mutant genes. Here we provide an overview of the genes associated with CDs, and highlight some of the epigenetic regulatory complexes involving multiple CD genes. Such common gene networks may provide a handle for designing therapeutic interventions applicable to a number of cognitive disorders with variable genetic etiology. This article is part of a Special Issue entitled ‘Neuroepigenetic disorders’. Crown Copyright Ó 2014 Published by Elsevier Ltd. All rights reserved.

Keywords: Intellectual disability Cognitive disorder Genetic Epigenetic Mutation Chromatin

1. Introduction Cognitive disorders (CDs) are a large and heterogeneous collection of disorders, which collectively impose a large burden to health care systems. The 2013 update to the authoritative manual of the American Psychiatric Association (APA), the Diagnostic and Statistical Manual of Mental Disorders Fifth Edition (DSM-5; APA, 2013) has 18 diagnostic groups, which each contain a variety of subgroups. This grouping is a subject of much criticism since clinical and genetic research increasingly suggests that mental illnesses are cannot be so clearly defined, and rather are spread along a spectrum with a high degree of overlapping characteristics (Adam, 2013). Disorders that have a major genetic predisposition are found in the diagnostic groups “Neurodevelopmental Disorders”, comprising intellectual disability (ID), autism spectrum disorders (ASD), communication disorders and motor disorders, and the group “schizophrenia (SCZ) and other psychotic disorders”. Regarding the associated direct and indirect costs, ID presents a large burden to our society and in the Dutch health care system it ranks highest in the costs of all disease categories (RIVM, 2011) The large-scale identification of ID and other CD genes is a promising * Corresponding author. Radboud University Medical Center, Department of Human Genetics 855, P.O. Box 9101, 6500 HB Nijmegen, The Netherlands. Tel.: þ31 24 3614017; fax: þ31 24 3668752. E-mail address: [email protected] (H. van Bokhoven).

strategy for identifying networks of interacting proteins and novel regulatory pathways that play a role in CDs. Moreover, it is a prerequisite for understanding the underlying pathogenic mechanisms and for the development of therapies. Severe ID (IQ < 50) affects 0.3e0.4% of the general population. Approximately half of these cases are caused by chromosomal aberrations and single gene mutations, whereas borderline ID (IQ 70e 85) can be expected to be the consequence of multiple genetic variants (Coe et al., 2012). To date, mutations in more than 500 genes have been implicated in ID (van Bokhoven, 2011; Kochinke et al., in preparation). Due to the advent of next generation sequencing (NGS) technology, the number of known ID genes is steeply increasing. Based on current number of well over 100 known ID genes on the X chromosome, an estimated total number of around 2500 genes might be involved in monogenic causes of ID. Likewise, NGS has been performed in large cohorts of patients diagnosed with ASD, SCZ, and epilepsy, with or without associated mental features such as ID (O’Roak et al., 2012; Neale et al., 2012; Carvill et al., 2013). These studies have revealed large numbers of de novo mutations, which might be causative or contributory to the neurodevelopmental phenotype of these patients. Here we will discuss exclusively the genes, collectively referred to as CD genes, for which compelling evidence exist about their contribution to the CD phenotype, based on the identification of recurrent mutations in the respective gene in independent patients or on supporting evidence from functional studies.

0028-3908/$ e see front matter Crown Copyright Ó 2014 Published by Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.neuropharm.2013.12.025

Please cite this article in press as: Kleefstra, T., et al., The genetics of cognitive epigenetics, Neuropharmacology (2014), http://dx.doi.org/10.1016/ j.neuropharm.2013.12.025

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T. Kleefstra et al. / Neuropharmacology xxx (2014) 1e12

1.1. CD genes operate in common molecular networks

1.2. Epigenetic genes and mechanisms

Causative mutations and variants in genes contributing to CDs are rapidly being elucidated by next generation sequencing approaches, which has great benefits for diagnostic and prognostic purposes. The identification of causative and contributory genetic defects provide a stepping stone towards gaining mechanistic insight into pathological pathways involved in CDs. Analysis of the biological functions of large groups of proteins has revealed key cellular processes underlying CD pathogenesis (Chelly et al., 2006). For example, impaired proliferation of neural progenitors is often seen in CDs with microcephaly, and mutations affecting cues involved in neural migration and axonal guidance are characteristic of disorders involving cortical malformations. Disruptions of synaptogenesis and control of synaptic activity are characteristic of ID, ASD, and schizophrenia (Grant, 2012). Indeed, many of the mutated genes associated with these disorders affect genes involved in core synaptic processes (van Bokhoven, 2011). In addition, a disproportionately high number of ID genes encode for proteins involved in chromatin-mediated control of transcription, further alluded to as epigenetic transcription regulation. In the human genome there are 572 genes currently annotated with the GO terms chromatin binding, chromatin remodeling, and chromatin modification, representing about 2.9% of all human genes. Amongst a catalog of 519 ID genes (Kochinke et al., in preparation) there are 40 genes annotated with these GO terms (7.7% of ID genes). Consequently, chromatin-related genes are enriched in ID by a factor of about 2.7 fold (Fig. 1). This is similar to the enrichment seen for synapse-related genes (3 fold), despite the likely bias towards analysis of synaptic function for genes that are implicated in neurological disorders (Hamdan et al., 2009). In combination with this gene ontology analysis we further curated list of 55 ID genes involved in chromatin-related regulation of transcription (Table 1). Mutations in these 55 genes are seen in families with an autosomal dominant, recessive, or X-linked inheritance. Notably, a high proportion of the associated disorders are caused by heterozygous loss of function mutations, suggesting that gene dosage is critically important for many epigenetic genes. The importance of correct dosage is nicely illustrated by the X-linked MECP2 gene, for which loss of function mutations are incompatible with life in males and associated with RETT syndrome in females, whereas gene duplications give rise to syndromic ID in males and females do not usually have any symptoms (Van Esch, 2008).

The genetic information encoded by our genomes is found in highly ordered chromatin structures that facilitate the packaging of DNA into the nucleus and are critical to control basic cellular processes such DNA replication, recombination, transcription, and DNA repair. The core organizational structure is the nucleosome, consisting of 147 basepairs of DNA wrapped around an octamer of histone proteins. Most nuclear DNA is found in this beads-on-astring structure has varying degrees of poorly understood local compaction and long-range contacts between loci. Traditionally, a distinction was made between two types of chromatin, euchromatin and heterochromatin, which were thought to reflect regions of active and inactive transcription. However, this generalization appears to be an oversimplification of the true genomic organization. Recently, combinatorial analysis of different histone modifications and chromatin binding proteins has revealed several novel models for chromatin organization that include anywhere from 5 to 51 different chromatin states, depending on the parameters and organism being analyzed (Bernstein et al., 2012; Zentner and Henikoff, 2013; Kundaje et al., 2012; de Graaf and van Steensel, 2013). The regional chromatin structure, as determined by epigenetic factors such as DNA modification, histone modification, protein complexes, non-coding RNA molecules (ncRNA), and nucleosome positioning, helps to define the gene expression profile of a given cell. DNA methylation is known for a long time as an important regulator of transcription. Methylation of cytosines (mC) at CpG sites, which are often enriched in islands at promoter regions, is generally associated with gene silencing. However, abundant DNA methylation is also seen at non-CpG islands, which may have an important role in the tissue-specific regulation of gene expression. Indeed, wholegenome single base profiling of the DNA methylome has revealed profound changes in the epigenomic profiles during brain development in mice and humans (Lister et al., 2013). Besides cytosine methylation also other cytosine modifications have been identified, such as 5-hydroxymethylcytosine (hmC), 5-formylcytosine (fmC) and 5-carboxylcytosine (caC) (Kriaucionis and Heintz, 2009; reviewed by Song et al. (2012)). Moreover, the recently discovered reversible methylation of N6-methyladenosine in RNA may uncover a totally new field of RNA epigenetics (Zheng et al., 2013). The post-translational modification of amino acid residues in the tails of histone proteins is another major epigenetic mechanism of transcription regulation. A variety of modifications can occur: acetylation, methylation, ubiquitylation, phosphorylation, sumoylation and numerous less well-studied modifications (Tan et al., 2011). The covalent binding of acetyl groups to lysines residues removes the positive charge on the histone tails, thereby decreasing the interaction of the N-termini of histones with the negatively charged phosphate groups of DNA. As a consequence, acetylated chromatin is less condensed, which facilitates transcription by making the DNA more accessible to transcription promoting protein complexes and the basic transcription machinery. In contrast, histone methylation can serve as both an activator and repressor of transcription, depending on the residue being modified and the degree of modification (i.e. mono-, di-, or trimethylation). For several histone modifications a specific activity towards transcription has been established: e.g. histone H3 lysine 9 di- and tri-methylation (H3K9me2/3; repression), histone H3 lysine 4 trimethylation (H3K4me3; activation), histone H3 lysine 27 acetylation (H3K27Ac, active enhancer), histone H3 lysine 27 trimethylation (H3K27me3, repression). However, there is still much to be learned about the effects of specific modifications, let alone about the effects imposed by combinatorial effects of possible DNA and histone modifications.

Fig. 1. Enrichment of Synapse-related and Chromatin-related genes in ID. All human genes associated with the GO terms synapse (GO:0045202), synapse organization (GO:0050808), and regulation of synaptic plasticity (GO:0048167) were combined to form the group “Synapse Related” (N ¼ 636). All genes associated with the terms chromatin binding (GO:0003682), chromatin modification (GO:0016568), and chromatin remodeling (GO:0006338) were combined to form the group “Chromatin Related” (N ¼ 572). Enrichment was based the proportion of genes present amongst all ID genes (N ¼ 519) compared to all human genes (N ¼ 20,687, Pennisi, 2012). Chromatin related, p ¼ 5.9 109; Synapse related, p ¼ 3.4 1012, hypergeometric test.


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Table 1 Genes with epigenetic functions that are involved in cognitive disorders. There are 55 genes in total which are distributed over four categories: 1. writers (n ¼ 17), 2. Erasers (n ¼ 5), 3. Chromatin remodelers of the DEAD/H-ATPase family (n ¼ 13), and 4. Other readers and chromatin remodelers (n ¼ 20). Gene names are provided by the official HGCN symbol (HUGO Gene Nomenclature Committee). In some cases alternative names are provided. The column “OMIM Gene” lists the number by which information of the gene can be assessed in the OMIM database (Online Mendelian inheritance in Man; http://www.ncbi.nlm.nih.gov/omim/). The cognitive disorders associated with mutations in the respective genes are listed and more information about these disorders can be found by following the number provided in the column “OMIM Morbid. The lasts colums list the epigenetic role of the respective protein and the mode of inheritance of the CD caused by mutations. AR, autosomal recessive; AD, autosomal dominant; XL, X-linked. Gene symbol

OMIM gene

1. Writers DNA methylation DNMT1 126375

DNMT3B FTO

602900 610966

Histone modification CREBBP 600140 CUL4B 300304

Phenotype

OMIM Morbid

Function (chromatin)

Inheritance

Hereditary sensory neuropathy type IE (HSN1E) Cerebellar ataxia, deafness, and narcolepsy, ICF syndrome Growth retardation, developmental delay, coarse facies, and early death

614116

DNA methylation

AD

604121 242860 612938

DNA methylation RNA demethylation

AR AR

EHMT1

607001

RubinsteineTaybi syndrome 1 (RSTS1) Mental retardation, X-linked, syndromic 15 (Cabezas type) Kleefstra syndrome

EP300 EZH2

602700 601573

RubinsteineTaybi syndrome 2 (RSTS2) Weaver syndrome

613684 277590

HLCS HUWE1

609018 300697

253270 300706

KAT6B

605880

KMT2A KMT2D KMT2C NSD1

159555 602113 607001 606681

WHSC1

602952

Holocarboxylase synthetase deficiency Mental retardation, X-linked syndromic, Turner type Ohdo syndrome, SBBYS variant Genitopatellar syndrome WiedemanneSteiner syndrome Kabuki syndrome Kleefstra Syndrome spectrum Sotos syndrome BeckwitheWiedemann syndrome WolfeHirschhorn syndrome

603736 606170 605130 147920 610253 117550 130650 194190

UBE2A 2. Erasers HDAC4 HDAC8

312180

X-Linked ID, Nascimento type

300860

605314 300269

AD

180849 300354

Histone acetyltransferase (HAT) Histone ubiquitination (H2AK119Ub1)

AD, de novo XL

610253

Histone methyltransferase (KMT1D; H3K9me1/2) Histone acetyltransferase (HAT) Lysine N-methyltransferase 6 (KMT6A; H3K27me3) Histone biotinylation (H4K16bio) Histone ubiquitination

AD, de novo

Brachydactyly-mental retardation syndrome WilsoneTurner syndrome Cornelia de Lange syndrome 5 KDM5C 314690 X-linked syndromic mental retardation; Claes-Jensen type KDM6A 300128 Kabuki syndrome 2 PHF8 300560 Siderius X-Linked Mental Retardation Syndrome 3. Chromatin remodelers (DEAD/H ATPase family) ACTB 102630 BaraitsereWinter syndrome Dystonia, juvenile onset ARID1A 603024 Mental retardation, Autosomal dominant 14

600430 309585 300882 300534

CoffineSiris syndrome Mental retardation, Autosomal dominant 12

135900 614562 135900 301040 309580 615369

300867 300263

243310 607371 614607

ARID1B

614556

ATRX

300032

CHD2

602119

CoffineSiris syndrome Alpha thalassemia mental retardation syndrome, X-linked (ATRX) Mental retardation-hypotonic facies syndrome Epileptic encephalopathy, childhood-onset

CHD7

608892

CHARGE syndrome

214800

CHD8

610528

Autism, susceptibility to, AUTS18

615032

SMARCA2

600014

NicolaideseBaraitser syndrome

601358

SMARCA4

603254

CoffineSiris syndrome Mental retardation, Autosomal dominant 16

135900 614609

SMARCB1

601607

CoffineSiris Mental retardation, Autosomal dominant 15

135900 614608

603111

CoffineSiris syndrome CoffineSiris syndrome

135900

SMARCE1

Histone acetyltransferase (MYST4) Histone Histone Histone Histone

methylation methylation methylation methylation

(MLL) (MLL2) (MLL3) (KMT3B)

Histone methyltransferase (NSD2) (H3K36me3, H4K20me2) Histone ubiquitination Histone deacetylase Histone deacetylase Histone demethylase (H3K4 tridemethylase) Histone demethylase (H3K27me3/2) Histone demethylase (H4K20me1, H3K9me1/me2) SWI/SNF, INO80 and ISWI complex DEAD/H ATPase helicase family, SWI/SNF subfamily DEAD/H ATPase helicase family, SWI/SNF subfamily DEAD/H ATPase helicase family

DEAD/H ATPase helicase CHD subfamily DEAD/H ATPase helicase CHD subfamily DEAD/H ATPase helicase CHD subfamily DEAD/H ATPase helicase SWI/SNF subfamily

AD, de novo AD, de novo AR XL AD, AD, AD, AD, AD, AD, AD, AD,

de novo de novo de novo de novo de novo mostly de novo de novo deletions

XL AD, de novo XL XL XL XL XL

AD, de novo AD* (single family, caution) AD, de novo AD, de novo AD, de novo AD, de novo XL

family,

XL AD, de novo

family,

AD, de novo

family,

AD, de novo

family,

AD, de novo

DEAD/H ATPase helicase family, SWI/SNF subfamily DEAD/H ATPase helicase family, SWI/SNF subfamily

AD, de novo AD, de novo

AD

(continued on next page)


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Table 1 (continued ) Gene symbol

SRCAP

OMIM gene

611421

Phenotype

Floating-Harbor syndrome

OMIM Morbid

Function (chromatin)

Inheritance

136140

DEAD/H ATPase helicase family, SWI/SNF subfamily DEAD/H ATPase helicase family, INO80/SWR subfamily SWI/SNF complex

AD, de novo

SS18L1 606472 Amyotrophic lateral sclerosis (ALS) 4. Other readers and chromatin remodelers ASXL1 612990 BohringeOpitz syndrome

605039

BCOR

300485

Microphthalmia, syndromic

300166

CHMP1

164010

pontocerbellar hypoplasia 8

614961

CTCF GATAD2B HCFC1

604167 614998 300019

ID, microcephaly and growth retardation Mental retardation, autosomal dominant 18 Mental retardation, X-linked; MRX3/

615074 309541

KANSL1 MBD5

612452 611472

MECP2

300005

PHF6

300414

POGZ

614787

SKI MED12

164780 300188

MED17

603810

MED23 NIPBL RAD21 SALL1 SMC1A SMC3

Methylmalonic acidemia and homocysteinenia (Cobalamin disorder) KooleneDe Vries syndrome Mental retardation, autosomal dominant 1 (2q deletion syndrome) Kleefstra Syndrome spectrum RETT syndrome; RTT RETT syndrome, atypical; Angelman syndrome-like Autism, susceptibility to, X-linked 3; AUTSX3 Encephalopathy, neonatal severe Mental retardation, X-linked, syndromic 13; MRXS13 Duplication MECP2; Lubs X-linked mental retardation syndrome BorjesoneForssmaneLehmann syndrome CoffineSiris-like Autism spectrum disorder

PR-DUB complex, histone H2A deubiquitination Polycomb complexes containing Ring1B Targets polycomb protein BMI to condensed chromatin Chromatin binding factor, insulator NuRD complex Found in repressor and activator complexes

AD, de novo AD, de novo XL AR AD, de novo AD, de novo XL XL

610443 156200

NSL1 histone acetyltransferase complex Associated with heterochromatin

AD, de novo AD, de novo

610253 312750 105830

Binds to methylated DNA

AD, de novo XL, mainly females XL

300496

XL

300673 300055

Xl XL

300260

XL

301900 135900

605042 608667 606462 602218

ShprintzeneGoldberg syndrome LujaneFryns syndrome OpitzeKaveggia syndrome OHDO syndrome MaateKieviteBrunner Microcephaly, postnatal progressive, with seizures and brain atrophy Mental retardation, autosomal recessive 18 Cornelia de Lange syndrome 1; CDLS1 Cornelia de Lange syndrome 4; CDLS4 TowneseBrocks syndrome

182212 309520 305450 300895 613668 614249 122470 614701 107480

300040 606062

Cornelia de Lange syndrome 2; CDLS2 Cornelia de Lange syndrome 3; CDLS3

300590 610759

The role of ncRNAs in epigenetic regulation of transcription is suspected to be very large given that most genes overlap with or can bind to multiple ncRNAs. These can affect transcription, translation and RNA breakdown. The important role of ncRNA in epigenetic regulation is most evident during inactivation of the X chromosome in females, where expression of the large ncRNA gene XIST initiates the condensation of the respective X chromosome into a barr body that is decorated with XIST transcripts (Augui et al., 2011). The initiation of X inactivation occurs randomly, leading to a roughly equal silencing of the maternal and paternal X chromosomes. Skewing of the X-inactivation ratio can occur as a consequence of an X-chromosomal aberration or mutation that induces a selective disadvantage for cells in which the mutation-carrying X chromosome is active. Genomic imprinting also appears to be coregulated by ncRNAs. An intriguing example is the silencing of the paternally derived UBE3A gene due to the paternal expression of a UBE3A-antisense transcript (UBE3A-ATS). This transcript is silenced on the maternal chromosome by parent-of-origin-specific DNA methylation in the UBE3A-ATS promoter, which allows for expression of UBE3A from the maternal chromosome (Mabb et al.,

NuRD complex Pogo transposable element with ZNF domain HDAC recruiting complexes Mediator complex Mediator complex Mediator complex Mediator complex Mediator complex Cohesin complex Cohesin complex Member of NuRD histone deacetylase complex Cohesin complex Cohesin complex

XL XL AD, de novo AD XL XL XL AR AR AD AD, de novo AD XL AD, de novo

2011). A maternal mutation in the UBE3A gene can cause Angelman syndrome, a classic example of an imprinting disorder characterized by severe ID, sleep disturbance, seizures, jerky movements, and frequent laughter. In neurons, but not in most other cell lineages, this results in the total absence of UBE3A protein. Mutations affecting ncRNAs have been associated with genetic disorders, including CDs (Berdasco and Esteller, 2013; Willemsen et al., 2011). However, to our knowledge these mutations do not directly impact other modifications of the chromatin structure (DNA and histones). Below we focus on neurodevelopmental disorders with known causal gene disruptions, which result in aberrant modulation of chromatin structure. 1.3. Epigenetic genes underlying cognitive disorders The epigenetic genes that we discuss are presented in Table 1 and have been divided into four major categories: 1. Writers of epigenetic modifications, involved in enzymatic addition of side groups (DNA methylation, RNA methylation and histone modification); 2. Erasers of epigenetic modifications, the enzymes that


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remove these side groups; 3. Chromatin Remodelers of the DEAD/ H ATPase helicase family, which are involved in the regulation of nucleosome positioning; 4. Other readers and chromatin remodelers, containing proteins that recognize and bind to their cognate chromatin marks, as well as proteins that are found in transcription repressor and activator complexes and other complexes that regulate DNA accessibility by the basic transcription machinery. Below we summarize the characteristics of the epigenetic genes and associated disorders. For additional extensive descriptions we refer to other comprehensive reviews (Kramer and van Bokhoven, 2009; van Bokhoven and Kramer, 2010; Berdasco and Esteller, 2013; Millan, 2013; Ronan et al., 2013). 2. Writers 2.1. DNA/RNA methylation In mammals, there are three active DNA methyltransferases, DNMT1, DNMT3A and DNMT3B, and one related protein that lacks catalytic activity (DNMT3L). DNMT3A/B are de novo DNMTs, whereas DNMT1 secures maintenance of DNA methylation. Mutations in two of the three active DNMTs have been linked to CDs. Dominant mutations in DNMT1 are associated with hereditary sensory neuropathy type IE (HSN1E; Klein et al., 2011) and with autosomal dominant cerebellar ataxia, deafness, and narcolepsy (Winkelmann et al., 2012). Recessive mutations in DNMT3B cause immunodeficiency-centromeric instability-facial anomalies syndrome 1 (ICF1; Xu et al., 1999). DNA methylation is known to be important for brain development, as well as memory consolidation (Miller et al., 2010). Hence, it is not surprising that disruption of the methylation machinery can cause a cognitive disorder. Analogous to DNA modifications, numerous different kinds of RNA modifications can occur. Although these modifications are not strictly epigenetic (i.e. “on top of the genetic code”), such modifications regulate gene activity and hence are popularly referred to as RNA-epigenetics (Zheng et al., 2013). Cellular RNAs contain more than a hundred structurally distinct post-transcriptional modifications at thousands of sites. Some RNA modifications are dynamic and may have critical regulatory roles analogous to those of protein and DNA modifications. The internal N6-methyladenosine (m6A) modification in messenger RNA is one of the most abundant RNA modifications in eukaryotes. Demethylation of m6A is mediated by FTO (fat mass and obesity-associated protein). Polymorphisms in the FTO gene are associated with variability in body mass index, but the underlying mechanism has not been studied (Yang et al., 2012). In addition, a homozygous missense mutation in the FTO gene, resulting in an Arg316Gln substitution co-segregated with an ID phenotype in a consanguineous family with growth retardation, developmental delay, coarse facies, and early death (Boissel et al., 2009). 2.2. Histone modifications

Q1

The importance of post-translation histone modifications in gene regulation has been established in many studies. Last year, comprehensive genome-wide histone modification profiles related to gene expression and chromatin structure were established by the Encyclopedia of DNA Elements (ENCODE) project, o expression (Bernstein et al., 2012). These studies suggested that up to 80% of the genome outside the coding regions may have a function and not reflect “junk DNA”. Fourteen epigenetic genes associated with ID phenotypes encode proteins engaged in the post-translational modification of histone tails (Table 1). Histone acetyltransferases (HATs) such as CBP (encoded by CREBBP) and p300 (EP300) are the least specific of these histone-modifying enzymes as they can

5

acetylate a variety of lysine residues of histone proteins (H2A, H2B, H3 and H4) and non-histone proteins. They bind specifically to phosphorylated CREB (cAMP response element-binding protein) and enhance its transcriptional activity toward cAMP-responsive genes. In the nervous system this cAMP-mediated response is involved in the formation of long-term memories (Dash et al., 1990; Bourtchuladze et al., 1994). Interestingly, de novo mutations in these closely related HATs give rise to RubinsteineTaybi syndrome, characterized by ID, postnatal growth deficiency, microcephaly, broad thumbs and halluces, and characteristic facial appearance (Petrij et al., 1995; Roelfsema et al., 2005). The notion that CBP and p300 mutations give rise to similar phenotypes suggests that these paralogous proteins have shared functions, which is supported by their high sequence similarity and similar domain structure, including a bromodomain, cysteineehistidine-rich regions, and a histone acetyltransferase domain. The Lysine acetyltransferase 6B (KAT6B, also known as MYST4) underlies two distinct CDs: the SayeBarbereBieseckereYounge Simpson variant of Ohdo syndrome, which is usually associated with severe ID, delayed motor milestones, and significantly impaired speech (Clayton-Smith et al., 2011), and Genitopatellar syndrome, with severe psychomotor retardation and microcephaly. The different phenotypic features of these two conditions and the location of the underlying mutations suggest that distinct molecular mechanisms account for the respective phenotypes (Campeau et al., 2012). Of interest, the MaateKieviteBrunner variant of Ohdo syndrome is caused by a mutation in the MED12 subunit of the mediator complex, which bridges chromatin modifications to the transcription machinery (see below). In contrast to acteyltransferases, histone methyltransferases (HMTs) have a higher degree of specificity for catalyzing the modification of specific lysine residues. This is reflected by the large number of histone methyltransferases (at least 27 according to the HUGO Gene Nomenclature Committee; http://www.genenames. org/genefamilies/KDM-KAT-KMT; Allis et al., 2007) that are encoded in the mammalian genome. Mutations in seven HMTs are currently linked to a CD. Mutations are almost exclusively de novo and give rise to loss of one functional copy of the gene. Three of these genes are implicated in so-called overgrowth syndromes. Mutations in the NSD1 (Nuclear receptor binding SET domain) gene cause Sotos syndrome, which is characterized by pre- and postnatal overgrowth, macrocephaly, advanced bone age, typical facial features, and variable degrees of ID (Kurotaki et al., 2002). Interestingly, NSD1 mutations were also observed in patients with another overgrowth disorder, BeckwitheWiedeman syndrome (BWS). BWS is an imprinting disorder involving a cluster of genes on chromosome 11p15, including H19, CDKN1C and KCNQ1OT1. Therefore, it has been suggested that NSD1 is involved in imprinting of the chromosome 11p15 region (Baujat et al., 2004). Also of interest is that there is considerable phenotypic overlap between Sotos and Weaver syndrome, and NSD1 mutations have been identified in three patients initially diagnosed with Weaver syndrome (Douglas et al., 2003; Tatton-Brown and Rahman, 2013). Mutations in the related gene NSD2, which is not listed among the 27 HMTs in the HUGO gene nomenclature overview, is located in the minimal critical region defined by hemizygous deletions of chromosome 4p16.3 that are associated with WolfeHischhorn syndrome (Stec et al., 1998). WolfeHirschhorn syndrome is a congenital malformation syndrome characterized by pre- and postnatal growth deficiency, intellectual disability (usually severe), characteristic craniofacial features, cardiac features, seizures, and midline defects. NSD2, also known as WHSC1 is a histone H3 lysine 36 trimethyltransferase which is associated with active transcription. Interactions between NSD2 and EZH2, which is mutated in an overgrowth syndrome (see below), have been reported in


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oncogenesis (Asangani et al., 2013). In addition, NSD2 interacts with NKx2-5 in heart development and Sall1, a cell-type specific transcriptional co-regulator of the NuRD complex, which is mutated in syndromic ID (TowneseBrock syndrome, see below and Table 1) (Nimura et al., 2009). Weaver syndrome is an overgrowth syndrome for which de novo mutations were recently identified in the EZH2 gene (Enhancer of Zeste Homolog 2) (Gibson et al., 2012; Tatton Brown et al., 2011). The observed phenotypic overlaps seen for NSD1 and EZH2 mutations suggest that the corresponding proteins act in similar epigenetic regulatory pathways. NSD1 preferentially methylates lysine residue 36 of histone 3 (H3K36) and lysine 20 of histone H4 (H4K20), which are primarily associated with active transcription. In contrast, EZH2 shows specificity for trimethylation of histone H3 on lysine 27 (H3K27me3), which is associated with transcriptional repression. These apparent opposite activities might be differently affected by the mutations identified in the respective genes as NSD1 mutations in Sotos syndrome typically exhibit loss of function effects whereas EZH2 mutations in Weaver syndrome are expected to give rise to the synthesis of mutant EZH2 protein, with possible gain of function effects. The four other HMT proteins affected by mutations in CDs include three members of the mixed-lineage leukemia protein (MLL) protein family and the euchromatic histone-lysine N-methyltransferase 1 (EHMT1). EHMT1 preferentially mono- and dimethylates histone H3 at lysine 9 (H3K9me1/me2). This is thought to contribute to epigenetic transcriptional repression by recruiting HP1 proteins (Ogawa et al., 2002). MLL proteins are class 2 lysine-specific methyltransferases (KMT2 proteins, their official name), which preferentially mono- and tri-methylate lysine 4 of histone H3 (H3K4me1/me3). H3K4me1 and H3K4me3, are thought to be specific tags for enhancers and for epigenetic transcriptional activation, respectively (Del Rizzo and Trievel, 2011; Goldsworthy et al., 2013; Herz et al., 2012). Chimeric protein fusions involving KMT2 proteins as well as somatic mutations are frequently found leukemia, lymphomas and other tumors, such as medulloblastoma. Strikingly, germline mutations in these genes do not give rise to cancer-prone phenotypes, but rather to three syndromes with multiple congenital anomalies that show little phenotypic overlap beside their neurodevelopmental defects. De novo mutations in KMT2A (MLL) cause WiedemanneSteiner syndrome, characterized by hairy elbows, short stature, facial dysmorphism, ID, and speech delay (Jones et al., 2012). De novo mutations in KMT2D (MLL2) are seen in Kabuki syndrome, a congenital syndrome with ID and additional features, such as postnatal dwarfism and a typical facial appearance, reminiscent of the make-up of traditional Japanese Kabuki actors (Ng et al., 2010). Finally, de novo mutations in KMT2C (MLL3) have been identified in a patient with a phenotype resembling Kleefstra syndrome (Kleefstra et al., 2012) and in a patient with autism spectrum disorder and a nonverbal IQ of 82 (O’Roak et al., 2012). Mutations in the EHMT1 gene give rise to the archetypal Kleefstra syndrome, suggesting a functional relationship between EHMT1 and KMT2C. Indeed these two proteins and several others are part of an epigenetic underlying the Kleefstra syndrome phenotypic spectrum (Kleefstra et al., 2012), as further discussed below. The final three epigenetic writers implicated in CD mediate histone modifications with poorly defined effects on transcription, ubiquitination and biotinylation. Two proteins with a role in histone ubiquitination are involved in unrelated X-linked ID syndromes. Cullin 4B (CUL4B) is the main component of the Cullin4BRing E3 ligase complex, which physically associates with polycomb-repressive complex 2. It was shown that CUL4B represses gene expression through its intrinsic monoubiuitination activity towards histone H2A (H2AK19ub1) (Hu et al., 2012). UBE2A

associates with the E3 enzyme BRE1 and catalyzes the monoubiquitination of histone H2B (H2BK120ub1), a specific tag for epigenetic activation of transcription (Kim et al., 2005). Interestingly, the ASXL1 gene, involved in Boring Opitz syndrome, is a core component of the PR-DUB protein complex involved in histone deubiquitination, as discussed below. Finally, the Holocarboxylase synthetase (HLCS) catalyzes the covalent binding of biotin to lysines in histones H3 and H4, thereby creating rare gene repression marks such as K16-biotinylated histone H4 (H4K16bio) that promote nucleosome condensation (Singh et al., 2013). Furthermore, HLCS interacts physically with both DNMT1 and MeCP2 in the transcriptional repression of long-terminal repeats (Xue et al., 2013). 3. Erasers Five erasers of epigenetic marks have been linked to CD, including two histone deacetylases (HDAC) and three histone demethylases (Table 1). HDAC4 is a class II HDAC expressed in the cranial neural crest, osteoblasts, heart, skeletal muscle, and brain. It acts to inhibit a variety of transcription factors, including MEF2C and RUNX2, both of which are essential for proper skeletal development. HDAC4 expression and activities are in line with the phenotype observed in mouse models and in human individuals carrying de novo deletions and mutations affecting the HDAC4 gene that have brachydactyly-mental retardation syndrome (Williams et al., 2010). HDAC8 is a class I HDAC that is expressed in the nervous and alimentary system and controls patterning of the skull by repressing transcription factors in the neural crest (Haberland et al., 2009). Mutations in the X-linked HDAC8 gene have been identified in males with WilsoneTurner syndrome, a syndromic form of ID, characterized by severe intellectual disability, dysmorphic facial features, gynecomastia, hypogonadism, short stature, and truncal obesity in males, whereas females have a milder phenotype (Haraklova et al., 2012). These features resemble those of Borjesone ForssmaneLehmann syndrome, another X-linked disorder caused by mutation of the PHF6 gene which is a component of the nucleosome remodeling and deacetylase complex (NuRD complex, see below). Interestingly, HDAC8 mutations were recently shown to be responsible for a small subgroup of patients with Cornelia de Lange syndrome. The majority of cases with this syndrome have mutation in components of the cohesin complex (see Table 1, subgroup 4): NIPBL, SMC1A, SMC3 and RAD21. Interestingly, it was demonstrated that HDAC8 also acts as a deacetylase of SMC3, thus linking HDAC8 to the cohesin acetylation cycle (Deardorff et al., 2007, 2012a, 2012b). The histone demethylases KDM5C (JARID1C), KDM6A (UTX), and PHF8 are involved in the removal of methyl groups from specific lysines. KDM5C and KDM6A are members of the Jumonji C family of proteins. Mutations in the X-linked KDM5C gene cause syndromic ID with variable severity of cognitive and dysmorphic phenotypes in males and females. There is some correlation between phenotypic severity and the predicted disruptive effect of the KDM5C mutation (Rujirabanjerd et al., 2010). KDM5C specifically acts as a demethylase for di- and trimethylated H3K4me3, thus converting an activating histone mark into a repressive one by recruiting histone deacetylases and RE1-silencing transcription factor (REST) to neuron-restrictive silencer elements. This activity is antagonistic to the H3K4 trimethylase activity of the KMT2 proteins. KDM6A occupies the promoters of HOX gene clusters and regulates their transcription by modulating the recruitment of the Polycomb Repressive Complex PRC1 and the monoubiquitination of histone H2A. KDM6a demethylates H3K27me3, which through association with KMT2C/D complexes, is concomitant with methylation of H3K4 (Lee et al., 2007). The phenotype associated with KDM6A mutations resembles classical Kabuki syndrome,


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which is usually caused by mutations in KMT2D/MLL2 (Lederer et al., 2012). The PHD finger protein 8 (PHF8) is a demethylase that acts on repressive marks such as H3K9me/me2, H3K27me2, and H4K20me1, thus promoting transcription. Mutations in this gene cause ID with cleft lip/palate (Laumonnier et al., 2005).

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functions, found to be an important constituent of DEAD/H ATPase complexes, including the nBAF, INO80 and ISWI complexes (Hargreaves and Crabtree, 2011).

5. Other readers and chromatin remodelers 4. Readers of the DEAD/H ATPase helicase family Proteins involved in reading of the epigenetic code and in remodeling of the chromatin structure constitute the largest and most variable category of CD-associated proteins. We have subdivided these in essentially two classes. Group 3 contains proteins of the DEAD/H ATPase helicase family, which are key to positioning of nucleosomes. The removal and exchange of nucleosomes requires energy, which is provided by the intrinsic ATPase activity of proteins of the DEAD/H family of helicase. This family contains four subfamilies, SWI/SNF, INO80/SWR1, ISWI, and CHD ATPases (Hargreaves and Crabtree, 2011). Several members of the SWI/SNF family have been implicated in CoffineSiris syndrome, which is characterized by variable phenotypic manifestations including ID, coarse facial features, hypertrichosis, and hypoplastic or absent fifth fingernails or toenails. CoffineSiris syndrome is most often caused by mutation of the ARID1B gene, but de novo mutations have also been identified in other genes: ARID1A, SMARCA2, SMARCA4, SMARCB1 and SMARCE1 encoding subunits of the neuronal chromatin remodeling complex nBAF (Tsurusaki et al., 2012). The disruption of individual constituents of the same protein complex underlying a group of similar phenotypes is in line with the general notion that similar phenotypes share a common molecular and cellular etiology (Oti and Brunner, 2007). Interestingly, mutations affecting the chromatin remodeler PHF6 were originally identified in BorjessoneForssmaneLehmann syndrome, but more recently also in CoffineSiris syndrome (Wieczorek et al., 2013). PHF6 appears to be part of the NuRD complex, that also includes CHD4 and HDAC1 (Todd and Picketts, 2012), but no functional interactions have been reported with members of the nBAF complex that could explain the similarities to the Coffin-Siris phenotype. It should be noted that the Coffin-Siris phenotype is highly variable and that mutations in the associated genes can lead to other phenotypes, such as NicolaideseBaraitser syndrome (SMARCA2), Kleefstra syndrome spectrum (SMARCB1) or unspecified dominant ID (several genes). Moreover, de novo mutations were recently identified in SS18L1 (also known as CREST), which encodes another subunit of the nBAF complex (Chesi et al., 2013). The respective patients had no similarity with CoffineSiris syndrome, but had developed amyotrophic lateral sclerosis, a devastating neurodegenerative disorder. Three members of the chromodomain (CHD) subfamily of ATPase helicases have been associated with various neurodevelopmental disorders. Mutation of CHD7 is the most common cause of CHARGE syndrome, an acronym for the most prominent clinical features: Coloboma, Heart anomaly, choanal Atresia, Retardation, Genital and Ear anomalies (Vissers et al., 2004). More recently, de novo mutations in CHD2 and CHD8 were identified in several patients with epileptic encephalopathy (Carvill et al., 2013) and ASD (Neale et al., 2012), respectively. The latter study and several other large studies involving exome sequencing in ASD trios have also revealed single de novo mutations in other CHD genes, but these remain to be confirmed in further patients. The ATRX gene, involved in alpha-thalassemia-mental retardation (Gibbons et al., 1995) and SRCAP (SNF2-related CBP activator protein), mutated in floating harbor syndrome (Hood et al., 2012), are also members of the DEAD/H ATPase family. Finally, BaraitsereWinter syndrome is caused by mutations affecting the ubiquitous b-actin protein (Rivière et al., 2012), which is, among many other cellular

This category includes genes that encode integral members of protein complexes that recognize and bind to chromatin and help to reshape the chromatin structure to regulate accessibility to the transcription machinery (Table 1). The MECP2 protein, which is involved in a variety of neurological disorders including Rett syndrome, is an archetypal reader of the epigenetic code as it mediates binding of protein complexes to methylated CpG sites. Because of its interaction with histone deacetylases and co-repressor complexes, and because CpG methylation at promoters is associated with transcriptional repression, it was initially thought that MECP2 acts strictly in transcriptional repression. However, it is now well established that MECP2 has a much broader regulatory role. It binds to methylated CpG sites globally and is also engaged in transcription activation (Charour et al., 2008). The methyl-CpG binding domain protein 5 (MBD5) is another potential reader of DNA methylation, although its direct binding to DNA has not been demonstrated. MBD5 is associated with heterochromatin (Laget et al., 2010). Deletions of chromosome 2q23.1 encompassing the MBD5 gene are relative common and haploinsufficiency of MBD5 appears to be causative for ID in these patients as MBD5-specific mutations give rise to similar phenotypes (Wagenstaller et al., 2007; Kleefstra et al., 2012; Hodge et al., 2013). Several of the genes defined in this last category do not have a clear enzymatic function, but are consistently found as core components in various different transcriptional repressor (ASXL1, BCOR, SKI), or activator (HCFC1, KANSL1) complexes. Loss of function mutations in ASXL1 (Additional sex combs-like) are found in BohringeOpitz syndrome, a severe developmental and malformation disorder characterized by intrauterine growth retardation, poor feeding, profound ID, and various other congenital anomalies (Hoischen et al., 2011). ASXL1 is a core component of the Polycomb repressive deubiquitinase (PR-DUB) complex, a dimeric protein complex that removes monoubiquitin from histone H2A. The chromatin-modifying protein, charged multivesicular body protein (CHMP1) is also involved in polycomb-mediated gene silencing by targeting the Polycomb group (PcG) protein BMI1/PCGF4 to regions of condensed chromatin (Stauffer et al., 2001). Recessive mutations in CHMP1 cause pontocerbellar hypoplasia 8, characterized by severe psychomotor retardation, abnormal movements, hypotonia, spasticity, and variable visual defects (Mochida et al., 2012). Mutations in the X-linked BCL6 corepressor gene, BCOR, are found in Lenz microphthalmia and Oculofaciocardiodental syndrome (OFCD), which typically features eye anomalies (microphthalmia or anophthalmia), microcephaly and structural brain anomalies, but usually no cognitive impairments. BCOR acts as a corepressor by recruiting BCL6, MLLT3 and several HDACs (Huynh et al., 2000). Also the SKI protein is a transcriptional corepressor, in particular for repressing TGFb target genes through interaction with PRDM16. This promotes recruitment of the SMAD3-HDAC1 complex to the promoter of these target genes (Harada et al., 2003). GATAD2B is a core component of the NuRD complex that seems to regulate the localization of this complex to specific subnuclear foci via interactions with MBD2 (Brackertz et al., 2006). PHF6 was shown to interact with members of the NuRD complex in a proteomic analysis, however its function in this complex is not known (Todd and Picketts, 2012). Mutations in these NuRD complex components are associated with differential types of syndromic ID


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(Lower et al., 2002; de Ligt et al., 2012; Wieczorek et al., 2013; Willemsen et al., 2013). Mutations in HCFC1 are associated with ID and specific metabolic effects (cobalamin disorder; Yu et al., 2013). HCFC1 is engaged in a large number of proteineprotein interactions both in transcription repressor (SIN3A/HDAC) and activator complexes such as the highly conserved COMPASS (Shilatifard, 2012) and NSL complexes (Wysocka et al., 2003). The COMPASS complexes activate transcription via H3K4 methylation, while the NSL complex activates transcription through acetylation of H4K16. Another core component of the NSL complex, KANSL1, is the culprit in Koolen-de Vries syndrome, a recognizable form of syndromic ID formerly known as chromosome 17q21.31 microdeletion syndrome (Koolen et al., 2012). Other CD-associated genes encode proteins with various roles in chromatin remodeling during transcription and other cellular processes. For example, POGZ, which has recurrent de novo mutations in ASD patients (Iossifov et al., 2012), is essential for normal mitotic progression. The CCTC-binding factor CTCF is involved in several transcriptional mechanism: repression, activation, and enhancer blocking (Weth and Renkawitz, 2011) respectively. Moreover, CTCF regulates the three-dimensional chromatin structure by binding to insulator elements. The binding to insulators prevents interactions between promoter and nearby enhancers and silencer elements, which makes CTCF a key regulator of transcription through control of long-range chromatin-mediated interactions (Phillips-Cremins and Corces, 2013). Mutations in the CTCF gene are associated with ID, microcephaly, and growth retardation (Gregor et al., 2013). Besides the epigenetic co-regulator proteins, category 4 contains proteins that are found in large complexes bridging chromatin remodeling complexes to basic transcription factors. One such complex is the cohesion complex, for which mutations of several of its components (NIPBL, SMC1A, SMC3, RAD21, HDAC8) give rise to a clinically similar phenotype known as Cornelia de Lange syndrome (Deardorff et al., 2007, 2012a, 2012b). Cohesin is a conserved multi-protein complex that plays a crucial role in keeping sister chromatids together from S-phase until mitosis or meiosis. Cohesion facilitates DNA damage repair that occurs during replication, and enforces faithful segregation of chromosomes during cell division. In addition, cohesin regulates gene expression by stabilizing the chromatin loops formed by CTCF (Mehta et al., 2013). Another “bridging complex” that is involved in ID is the mediator complex, which links chromatin modifying complexes to the basal transcriptional machinery. Three mediator components, MED12, MED17 and MED23, are implicated in various unrelated forms of ID. The Mediator complex consists of 25e30 subunits, and plays a central role in the regulation of RNA Pol II mediated transcription (Ansari et al., 2012). Different subunits of Mediator are clustered in modules (head, middle, tail and CDK domain) that mediate contacts to other proteins and complexes involved in various activities including transcriptional co-regulators, general transcription factors, subunits of RNA Pol II, and chromatin. In contrast to mutations in various members of the cohesion complex, the phenotypes associated with mutations in individual mediator genes is variable aside from the occurrence of ID. This may be a reflection of the wide variety of different interactions that each of the mediator protein has. Our compilation of epigenetic regulators in CD does include transcription factors that are known to interact with remodeling complexes discussed above, because by definition, transcription factors (TF) are not epigenetic regulators. In general, by binding to specific target DNA sequences these TFs provide specificity in the range of controlled target genes by repressor and activator complexes. Mutations in a variety of TFs give rise to phenotypes that

include CD, for example MEF2C, YY1, Sall1 and members of the KRAB-domain Zinc finger family (ZNFs) to name a few. 5.1. Similar CD phenotypes reflect functional relationships between epigenetic CD genes The phenotypic similarity that is seen for a number of genetic disorders despite the involvement of mutations in distinct genes is a reflection of common molecular pathways and cellular processes that are affected by such mutations. There are many examples for which a common clinical phenotype is caused by mutations in different genes that have a known functional relationship, for example proteineprotein interactions, complementary enzymatic activities, or co-regulation of target genes. This is also the case for CD genes with an epigenetic function. Construction of a protein interaction network based on currently annotated proteineprotein interactions reveals a striking degree connectivity amongst the epigenetic ID genes (Fig. 2). Several examples of such interactions have already been discussed above: the cohesion mutations in Cornelia de Lange syndrome, nBAF mutations in CoffineSiris syndrome, and mutations affecting the interacting proteins KMT2D and KDM6A in Kabuki syndrome. We have also discussed some cases where the same phenotype involves different proteins for which functional or even molecular relationships are still to be elucidated, e.g. KAT6B and MED12, which underlie different variants of Ohdo syndrome. It is possible that the respective genes are involved in parallel pathways that eventually converge onto similar cellular processes. However, it is also likely that in such cases our knowledge of the activities is currently insufficient to recognize their shared activities or protein networks. The identification of mutations in distinct genes underlying a similar disorder can be an incentive to look for interactions between these genes. For Kleefstra syndrome spectrum phenotypes, genetic interaction studies in Drosophila combined with knowledge about proteineprotein interactions has uncovered a chromatin modification module encompassing EHMT1 and four other genes that were found to carry de novo mutations associated with this recognizable phenotype (Fig. 2; Kleefstra et al., 2012). Undoubtedly, this module will be extended, as a significant number of patients with the Kleefstra syndrome phenotypic spectrum do not have mutations in any of these five genes. Moreover, the proteins contained in the module, e.g. SMARCB1 and KMT2C are engaged in a wide variety of other proteineprotein interactions, including proteins involved in CD that are presented in Table 1 (e.g. MED12 and KDM5C via EHMT2/ G9a. and with other SWI/SNF complex members via SMARCB1). Thus, our network of annotated proteineprotein interactions between chromatin remodeling CD genes, will likely expand in size and medical importance with the emergence of additional functional connections (Fig. 2). 5.2. Variable phenotypes can be due to mutation of the same epigenetic gene The opposite of the “shared pathway-shared phenotype paradigm” can also be seen when mutations in the same gene give rise to different clinical phenotypes. For example, mutations in SMARCB1 have been identified both in the Kleefstra syndrome phenotypic spectrum and in CoffineSiris syndrome. The reason for the differential phenotype could be due to the differential consequences of the underlying mutations. This seems to be the case for MECP2 mutations, where highly disruptive mutations give rise to femalerestricted Rett syndrome and mutations that predict residual protein function give rise to ID in males and no features in female mutation carriers. In addition, gene duplications predicting enhanced MECP2 activity give rise to yet another recognizable CD phenotype in


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Fig. 2. Epigenetic network in cognitive disorders. A) Protein network of the epigenetic regulators that are listed in Table 1 shows that they are highly connected. The network was generated using Genemania (www.genemania.org) and displays direct physical protein interactions. B) Magnification of the epigenetic network that comprises the SWI/SNF complex and genes causing the Kleefstra syndrome spectrum. The associated human disorders and the kind of the type of interaction are color-coded. Manual curation reveals further links between ID genes, illustrating that further connections can be expected to be uncovered in the future.

male patients. Several other mechanisms can attribute to the clinical variability observed for mutations in single genes. The genetic context of individual patients may introduce clinical variability due to the presence of additional independent mutations that add more features or increase the severity of the CD phenotype. The genetic background can also modulate the core phenotype by genetic (SNPs) and genomic polymorphisms (CNVs) that enhance or suppress features of the core phenotype (Coe et al., 2012). In addition, pre- and

postnatal environmental stressors might affect the phenotype as well. Finally, some caution is warranted in the diagnostic labels associated with the clinical classification of phenotypes. The clinical diagnosis appears to be highly dependent on the medical specialist who is making the diagnosis (i.e. psychiatrist, neurologist, pediatrician, clinical geneticist). In addition, we must recognize that mental illnesses are spread along a spectrum, and that splitting them into subcategories ignores their substantial clinical overlap.


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A remarkable twist in the genotypeephenotype discussion is introduced by the notion that somatic mutations in many of the epigenetic CD genes are associated with cancer (Shih et al., 2012). This begs the question whether patients with CD due to a germline mutation in an epigenetic gene have a greater risk to develop tumors. Indeed this seems to be the case for some disorders, for example Wilms tumors are relatively common in Sotos syndrome (EZH2 mutations). Also RubinsteineTaiby syndrome patients (CREBBP and EP300 mutations) have an increased risk of tumor formation (5%), including tumors located in the nervous system such us oligodendrogioma, medulloblastoma, neuroblastoma and meningioma (Miller and Rubinstein, 1995). In addition, germline missense mutations and splice site mutations of SMARCB1 predispose to familial meningiomas and schwannomas (Boyd et al., 2008; van den Munckhof et al., 2012). Surprisingly, there is no report of CD in these families and mutation carriers do have offspring in many cases. There are only incidental reports of brain tumors in CoffineSiris syndrome and it is not known whether these patients had a SMARCB1 mutation, however follow-up of these patients is certainly warranted. In any case, for most of the reported genes there is no documentation for an increased incidence of cancer. It thus seems that the high frequency of mutations in specific genes in specific cancer types, e.g. ASXL1 in myelodysplasia, may represent secondary hits contributing to tumorigenesis rather than the primary cause of the neoplasm. 5.3. Perspectives for pharmacological intervention Phenotypic defects induced by disruptions of the epigenetic machinery have a great potential for therapeutic intervention (Millan, 2013). For example, histone deacteylase inhibitors (HDAC inhibitors) such as valproic acid, have long been used for the treatment of psychiatric disorders and epilepsy. Furthermore, HDAC inhibitors have been shown to alleviate neurodegeneration in a Drosophila model of Huntington’s disease (Steffan et al., 2001) and memory deficits in a mouse model for ID (Alarcón et al., 2004). The identification of networks of epigenetic genes underlying CDs, such as the nBAF complex and the chromatin-modification module underlying Kleefstra syndrome, is of particular interest given recent evidence showing that epigenetic processes are not only important for neurodevelopmental processes but also for acute cognitive functioning (Day and Sweatt, 2011). Adult rescue of cognitive deficits has been accomplished in several animal ID models, including Mecp2 mouse models for Rett syndrome (Cobb et al., 2010) and EHMT-mutant flies (Kramer et al., 2011), raising hope that cognition can be improved postnatally. Further dissection of the epigenetic networks underlying ID and the mechanisms through which they influence cognitive function may generate strategies for the identification of drugs that alleviate phenotypic features in groups of patients with a different genetic etiology. Uncited reference Ansari and Morse, 2013; Leonard and Wen, 2002; Michaud et al., 2013; Slobbe et al., 2011; Smith and Meissner, 2013; Vissers et al., 2010. Acknowledgments Research in the groups of the authors is supported by the EU FP7 Large-Scale Integrating Project Genetic and Epigenetic Networks in Cognitive Dysfunction (241995 to A.S. and H.v.B.) and a grant from the Dutch Organization for Health Research and Development (ZON-MW 907-00-365 to TK).

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Genetics. Genetics driving epigenetics.

Genetics and epigenetics of gliomas.

Genetics and Epigenetics of Eating Disorders.

Genetics and epigenetics of adrenocortical tumors.

Genetics and epigenetics of primary biliary cirrhosis.

Epigenetics and Genetics of Viral Latency.

The role of epigenetics in cognitive ageing.

Genetics and Epigenetics of Diabetic Nephropathy.

Genetics and epigenetics of aging and longevity.

Genetics and epigenetics of endocrine neoplasia.

Inflammatory Bowel Disease: Genetics, Epigenetics, and Pathogenesis.

Epigenetics: where environment, society and genetics meet.

Interplay between Epigenetics and Genetics in Cancer.

Congenital heart disease: the crossroads of genetics, epigenetics and environment.

The genetics of loneliness: linking evolutionary theory to genome-wide genetics, epigenetics, and social science.

Genetics and epigenetics of arrhythmia and heart failure.

Carcinogenic mechanisms of endometrial cancer: involvement of genetics and epigenetics.

Genetics, epigenetics, and regulation of drug-metabolizing cytochrome p450 enzymes.

Understanding type 2 diabetes: from genetics to epigenetics.

Chemical genetics approaches for selective intervention in epigenetics.

Osteosarcoma Genetics and Epigenetics: Emerging Biology and Candidate Therapies.

From Genetics to Epigenetics: New Perspectives in Tourette Syndrome Research.

Antipsychotic induced weight gain: genetics, epigenetics, and biomarkers reviewed.

Genetics of specific cognitive abilities.