histone deacetylases transcriptional control, and cancer

REVIEW ARTICLE

Histone Deacetylases, TranscriptionalControl, and Cancer

W. DOUGLAS CRESS AND EDWARD SETO*Molecular Oncology Program, H. Lee Moffitt Cancer Center and Research Institute,

University of South Florida, Tampa, Florida

A key event in the regulation of eukaryotic gene expression is the posttranslationalmodification of nucleosomal histones, which converts regions of chromosomesinto transcriptionally active or inactive chromatin. The most well studied post-translational modification of histones is the acetylation of e-amino groups onconserved lysine residues in the histones’ amino-terminal tail domains. Signifi-cant advances have been made in the past few years toward the identification ofhistone acetyltransferases and histone deacetylases. Currently, there are over adozen cloned histone acetyltransferases and at least eight cloned human histonedeacetylases. Interestingly, many histone deacetylases can function as transcrip-tional corepressors and, often, they are present in multi-subunit complexes. Moreintriguing, at least some histone deacetylases are associated with chromatin-remodeling machines. In addition, several studies have pointed to the possibleinvolvement of histone deacetylases in human cancer. The availability of thecloned histone deacetylase genes has provided swift progress in the understand-ing of the mechanisms of deacetylases, their role in transcription, and theirpossible role in health and disease. J. Cell. Physiol. 184:1–16, 2000.© 2000 Wiley-Liss, Inc.

For many years, studies aimed at elucidating themechanisms of transcriptional activation sought to de-termine how DNA-bound transcription factors couldaffect transcription initiation and elongation by RNApolymerase II. A major leap forward in this endeavorwas the discovery that some DNA-bound transcrip-tional activation domains function, at least in part, bybinding so-called coactivator complexes that possesshistone acetyltransferase activity. Almost in parallel,investigators seeking to decipher the mechanisms oftranscriptional repression identified corepressor com-plexes that possess histone deacetylase activity. Thephysiological significance of these findings is under-scored by the fact that some human cancers have beenassociated with malfunctions of coactivator or corepres-sor components. The histone deacetylases, their rolesin transcriptional repression, and their involvement inhuman cancer are the central topics of this review.

DNA PACKAGING ANDHISTONE-MODIFYING ENZYMES

Eukaryotic DNA is packaged into chromatin. It isbecoming increasingly clear that the transcription ofthese tightly packaged genes is regulated, at least inpart, by chromatin-remodeling events, which can ren-der the DNA either more or less accessible to transcrip-tion factors. The most basic element of DNA packagingis the nucleosome, which consists of DNA wrappedtwice around a histone octamer, which contains twocopies each of four different histone proteins (H2A,H2B, H3, and H4). Nucleosomes form on DNA at ap-proximately every 200 base pairs, with 146 base pairs

tightly wound around the histone octamer and theremaining 54 base pairs intervening. This arrange-ment forms what appears, under electron microscopy,to be a series of beads on a string. The nucleosomesthemselves can interact with one another to form aspiral that has been termed the solenoid structure. Thisstructure is 30 nm in diameter, contains six nucleo-somes per turn, and, in some eukaryotic cells, requiresone copy of the histone H1 protein to interact with eachnucleosome. It is thought that DNA packaged into thiscompact structure is transcriptionally inert.

Histone acetyltransferasesCrystallography studies revealed that the N-termi-

nal tails of histone proteins protrude from the nucleo-some complex (Luger et al., 1997). These extensionsappear to be involved in higher-order packaging bymediating contacts between adjacent nucleosomes. His-tone tails contain highly conserved lysine residues thatcan be acetylated on their e-amino groups. Each acet-ylation event eliminates another positive charge andpotentially weakens the electrostatic interactions thattether the octamer tails to the DNA phosphate back-bone. Histone acetylation may affect chromatin struc-

Contract grant sponsor: National Institutes of Health.

*Correspondence to: Edward Seto, Molecular Oncology Program,H. Lee Moffitt Cancer Center and Research Institute, Universityof South Florida, 12902 Magnolia Drive, Tampa, FL 33612.E-mail: [email protected]

Received 10 February 2000; Accepted 21 February 2000

JOURNAL OF CELLULAR PHYSIOLOGY 184:1–16 (2000)

© 2000 WILEY-LISS, INC.

ture by at least two different mechanisms. First, thenet reduction in positive charge could lead to destabi-lization and consequent dissociation of nucleosomes,thus allowing access of transcription factors and RNApolymerase to the DNA. Second, histone acetylationmay inhibit the stacking of nucleosomes into the sole-noid structure and thus the formation of higher-orderstructure. With these phenomena in mind, it is easy toenvision a model in which the transcription of genes inany given region is essentially controlled by accessibil-ity. The disruption of higher-order structures by acet-ylation would be expected to stimulate transcription;conversely, the facilitation of higher-order structuresby deacetylation would inhibit transcription. This con-cept seems to work well in providing a theoretical plat-form from which to approach numerous model systems,but is actually an oversimplification. Indeed, it is quitepossible that acetylation may have either positive ornegative effects, depending on the particular gene in-volved (Sun and Hampsey, 1999). For example, a thirdmechanism by which histone acetylation could regulatetranscription is by affecting the binding of regulatoryproteins to the histones themselves. Furthermore, it ispossible that some “histone” acetyltransferases anddeacetylases, in fact, act on other transcription factorsinstead of, or in addition to, the histones (Gu andRoeder, 1997; Gu et al., 1997; Boyes et al., 1998; Zhangand Bieker, 1998; Sartorelli et al., 1999).

The first eukaryotic transcription factor recognizedto encode an acetyltransferase was the protein GCN5.Yeast GCN5 was initially identified as a global tran-scriptional coactivator via genetic techniques (Georga-kopoulos and Thireos, 1992; Marcus et al., 1994); how-ever, its biological function was not apparent until themomentous cloning of Tetrahymena histone acetyl-transferase A (Brownell et al., 1996). The deducedamino acid sequence of the Tetrahymena enzyme re-vealed obvious sequence homology with yeast GCN5and established, for the first time, a direct link betweentranscriptional coactivators and histone acetylation.

This link was firmly reinforced by the demonstrationthat GCN5 possesses acetyltransferase activity, andthat this activity is essential for transcriptional activa-tion in vivo (Candau et al., 1997).

It is now clear that numerous yeast and mammaliantranscriptional coactivators are, in fact, histone acetyl-transferases (for recent reviews see Hampsey, 1997;Struhl, 1998). Initially described as a transcriptionalcoactivator for a number of enhancer-binding proteins(Janknecht and Hunter, 1996), p300/CBP (CREB-bind-ing protein) is one of the best understood examples ofhuman histone acetyltransferases (Bannister andKouzarides, 1996; Ogryzko et al., 1996b). The p300/CBP protein forms complexes with P/CAF (p300/CBP-associated factor), which is also a well-characterizedhistone acetyltransferase (Yang et al., 1996b). Onetranscription factor that utilizes the p300/PCAF com-plex as a coactivator is the retinoic acid receptor alpha(RARa). RARa forms a DNA-binding heterodimer witha retinoid-X receptor (RXR). The RAR/RXR dimer re-presses transcription in the absence of the hormoneligand retinoic acid. Upon hormone binding, the dimerreleases the transcriptional inhibitory complex andsubsequently binds to the p300/PCAF complex. Thiscomplex can activate the transcription of a number ofdevelopmentally regulated genes, such as those in-volved in hematopoiesis (Lenny et al., 1997).

Histone deacetylasesBroadly speaking, histone deacetylase proteins from

various species can be divided into three categories: (1)the class I RPD3-like proteins; (2) the class II HDA1-like proteins; and (3) the class III maize HD2 protein(Table 1).

Class I. The first breakthrough in the identificationof the histone deacetylases came with the cloning ofHDAC1 (initially termed HD1). HDAC1 and an associ-ated protein, RbAp48 (Rb-associated protein 48), werecopurified by affinity chromatography using a modifiedform of the HDAC inhibitor trapoxin as an affinity

TABLE 1. Classification of histone deacetylases

SpeciesClass I:

RPD3-likeClass II:

HDA1-likeClass III:HD2-like Reference

Yeast RPD3 HDA1HOS1HOS2HOS3

Vidal and Gaber, 1991; Rundlett et al., 1996

C. elegans HDA1HDA2HDA3

Shi and Mello, 1998

Drosophila dHDAC1 (dRPD3, RPD3)dHDAC2 (DmHDAC2)dHDAC3 (DmHDAC3)

dHDA2 DeRubertis et al., 1996; Johnson et al., 1998

Xenopus HDm (RPD3) Ladomery et al., 1997Chicken HDAC1 (HDAC1A)

HDAC2 (HD2)HDAC3 (HD3)

Takami et al., 1999

Mouse HDAC1 (HD1)HDAC2 (mRPD3)HDAC3

mHDA1mHDA2

Yang et al., 1996; Bartl et al., 1997;Mahlknecht et al., 1999; Verdel andKhochbin, 1999

Human HDAC1 (HD1)HDAC2 (hRPD3)HDAC3

HDAC4 (HDAC-A)HDAC5 (HDAC-B)HDAC6HDAC7HDAC8

Taunton et al., 1996; Yang et al., 1996, 1997;Dangond et al., 1998; Emiliani et al., 1998;Fischle et al., 1999; Grozinger et al., 1999;Miska et al., 1999

Maize RPD3 HD2 Lusser et al., 1997; Aravind and Koonin, 1998

2 CRESS AND SETO

reagent (Taunton et al., 1996). Sequence analyses re-vealed that HDAC1 was very similar to the yeast RPD3protein (reduced potassium dependency), a knownyeast transcriptional regulator (Vidal et al., 1996).These findings provided the first direct experimentalevidence linking histone deacetylation and transcrip-tional control. Almost concurrently, a second HDAC(HDAC2) was identified as a protein that binds to thetranscription factor YY1 via use of the yeast two-hybridsystem (Yang et al., 1996a). Human HDAC1 andHDAC2 are 75% identical in DNA sequence and 85%identical in protein sequence, respectively.

Three groups have independently cloned an addi-tional member of the class I human histone deacetylasefamily (Yang et al., 1997; Dangond et al., 1998; Emil-iani et al., 1998). Analysis of the predicted amino acidsequence of this protein, HDAC3, revealed an openreading frame of 428 amino acids with a predictedmolecular mass of 49 kDa. HDAC3 and the clonedhuman HDAC1 share 50% sequence identity at theDNA level, and 53% identity at the protein level. Com-parisons of the HDAC3 protein and DNA sequenceswith those from human HDAC2 yielded similar results,with 51% identity in DNA sequence and 52% identityin protein sequence.

Class II. Yeast cells have at least two distinguish-able histone deacetylase activities catalyzed by differ-ent enzymatic complexes (Carmen et al., 1996;Rundlett et al., 1996). The active component of thehistone deacetylase A complex is the HDA1 catalyticsubunit. The histone deacetylase A complex has a na-tive molecular mass of approximately 350 kDa. Thecatalytic subunit of the 600-kDa histone deacetylase Bcomplex is RPD3 or the RPD3-related proteins HOS1,HOS2, or HOS3 (where HOS stands for HDA one sim-ilar) (Rundlett et al., 1996). Mammalian deacetylasesare found in much larger complexes (2,000 kDa) thatcontain an estimated 20 proteins, many of which havenot yet been identified. RPD3 and HDA1 share signif-icant sequence homology as well as homology withprokaryotic proteins known to interact with acetylatedsubstrates. A highly conserved region termed theRPD3 homology domain is clearly involved in catalysis.Deacetylase activity can be abolished by the introduc-tion of mutations within this domain. The class IIHDA1-like protein has at least five homologs in verte-brates; mHDA1 and mHDA2 in mouse; and HDAC4,HDAC5, HDAC6, HDAC7, and HDAC8 in human (Fis-chle et al., 1999; Grozinger et al., 1999; Miska et al.,1999; Verdel and Khochbin, 1999; Wang et al., 1999).Mouse HDA2 (human HDAC6) is particularly interest-ing in that it contains two yeast HDA1 homology do-mains.

Class III. The first two classes of histone deacety-lases have considerable homology with each other; forexample, RPD3 and HDA1 are 25% identical and 50%similar over a region of 490 amino acids. In contrast,the only Class III histone deacetylase, maize HD2, doesnot share obvious or extended regions of sequence sim-ilarities with the Class I or Class II enzymes (Lusser etal., 1997). Maize HD2, however, does share regions ofsequence homology with acidic nucleolar phosphopro-teins (e.g., UBF1, UBF2, nucleolin, and B23), and theimmunophilin FK506-binding proteins. Whether thenucleolar phosphoproteins or the FK506-binding pro-

teins can function as histone deacetylases remains tobe determined.

Within each class of deacetylase, homologs from dif-ferent species share relatively high sequence similari-ties. For example, human HDAC1 and yeast RPD3share 60% sequence identity and 80% sequence simi-larity over a region of 450 amino acids. Many addi-tional members of each deacetylase class have recentlybeen uncovered by searching EST databases for homol-ogous sequence elements, and have not yet been exten-sively characterized.

HISTONE DEACETYLASES ANDTRANSCRIPTIONAL COREPRESSORS

A major breakthrough in our understanding of tran-scriptional repression came from the recent discoveriesthat indicated that transcriptional repression, by atleast some proteins, is directly linked to the recruit-ment of multiprotein complexes containing histonedeacetylases. We discuss three specific examples oftranscriptional repressors that have served as para-digms: YY1, Mad/Max, and the nuclear hormone recep-tors.

YY1As its name suggests, the transcription factor YY1

(Yin Yang 1) can act as either a transcriptional activa-tor or a transcriptional repressor. This evolutionarilyconserved protein is ubiquitously expressed in manycells. A multitude of promoters and enhancers containpotential YY1-binding sites and many of the genescontaining these elements have been shown to be reg-ulated by YY1 (recently reviewed by Thomas and Seto,1999). In an effort to identify YY1-binding proteins(and thus better understand its mechanisms of action)a mouse RPD3 homolog, now known as HDAC2, wasidentified using a yeast two-hybrid screen (Yang et al.,1996a). A GAL4 DNA-binding domain HDAC2-fusionprotein strongly repressed transcription from a pro-moter containing GAL4-binding sites, suggesting thatYY1 negatively regulates transcription by tetheringHDAC2 to DNA as a corepressor, and that this tran-scriptional mechanism is highly conserved from yeastto human. This was the first demonstration that amammalian histone deacetylase can have a direct ef-fect on transcriptional control. Subsequently, it wasfound that HDAC1 and HDAC3 can also bind YY1 andrepress transcription when targeted to promoters(Yang et al., 1997).

In addition to HDAC1, HDAC2, and HDAC3, YY1also interacts with the nucleolar phosphoprotein B23(Inouye and Seto, 1994). Because the maize deacety-lase HD2 is closely homologous to nucleolar proteinsUBF1, UBF2, nucleolin, and, most interestingly, B23,it is tempting to speculate that YY1 recruits B23as part of the deacetylase enzymatic activities.The finding that YY1 interacts with three humanHDACs closely related to yeast RPD3 does not excludethe possibility that additional mammalian histonedeacetylases, perhaps with limited sequence homologyto yeast RPD3, may also be recruited by YY1 to represstranscription. Ribosomal RNA genes are localized dis-tinctly in the nucleolus and have their own specificsubset of transcription factors for transcription. Giventhat a subpopulation of YY1 is also localized within the

3HISTONE DEACETYLASES AND CANCER

nucleolus (Guo et al., 1995), it seems reasonable tospeculate that YY1 may direct a distinct family ofdeacetylases devoted to the regulation of rRNA synthe-sis.

Of the many other proteins that have been reportedto interact with YY1, an exceptionally interesting caseis the closely related coactivators CBP and p300 (Lee etal., 1995). Both of these coactivators are HAT enzymes.It is, therefore, quite appealing to speculate that YY1activates transcription by the recruitment of HAT ac-tivities. Perhaps it is the selective binding of eitherHAT or HDAC that determines whether YY1 will me-diate the activation or repression of transcription of agiven gene. Further studies aimed at uncovering thesignals that favor the recruitment of HDAC over HATmay shed some light on the mechanisms of YY1-medi-ated repression.

Although a number of different assays have beenused to confirm that YY1 interacts with class I HDACs,both in vitro and in vivo, it is noteworthy that YY1 hasnot yet been copurified with any of the HDAC com-plexes. It is conceivable that the YY1-HDAC interac-tion is direct and occurs in the absence of other inter-mediate proteins. In this regard, YY1 may be similar tothe Drosophila corepressor Groucho, which also inter-acts directly with the Drosophila RPD3 protein (Chenet al., 1999). Alternatively, it is possible that YY1 ispresent in a substoichiometric quantity compared withthat of other proteins in an HDAC complex. In anycase, further studies will be required to determine howthe YY1-HDAC interaction is regulated and to identifyadditional candidate components of the YY1-HDACcomplex.

Mad-Max and the Sin3/HDAC complexIn growing cells, Myc-Max heterodimers activate

transcription of growth stimulatory genes, such asE2F-2, that are regulated by E-box elements in theirpromoters (Sears et al., 1997; McArthur et al., 1998).However, upon cellular differentiation Myc is replacedby Mad and the Mad/Max heterodimer represses tran-scription of growth stimulatory genes (McArthur et al.,1998). Using the yeast two-hybrid system, the tran-scriptional repression domain of Mad was shown tobind to two mouse proteins, mSin3A and mSin3B, bothof which exhibit significant sequence homology withthe yeast SIN3 general transcriptional repressor (Ayeret al., 1995; Schreiber-Agus et al., 1995). The obviousquestion of “what does Sin3 do?” was quickly answeredwith the discoveries that mSin3A can be coimmunopre-cipitated with HDAC1 and HDAC2, as well as a nu-clear hormone transcriptional corepressor called NCoR(nuclear corepressor) (Alland et al., 1997; Hassig et al.,1997; Heinzel et al., 1997; Zhang et al., 1997). Theseobservations tied the transcriptional repressor Mad toa histone deacetylase as a potential mechanism forrepression, reminiscent of the case with YY1 (Heinzelet al., 1997; Laherty et al., 1997). However, unlike YY1,mSin3A can repress transcription in the absence of theHDAC interaction region, suggesting that mSin3 con-tains an alternative repression domain that is active inthe absence of bound HDAC.

Subsequent work demonstrated that the Sin3A/HDAC complex contains additional components, in-cluding RbAp46 and RbAp48 (Qian et al., 1993; Qian

and Lee, 1995), as well as SAP18 and SAP30 (Zhang etal., 1997; Laherty et al., 1998; Zhang et al., 1998b).RbAp46 and RbAp48 are related histone-binding pro-teins; they are thought to function at least in part byfacilitating interactions of the complex with the histonesubstrate (Parthun et al., 1996; Verreault et al., 1996,1998). The roles of SAP18 and SAP30 are not wellunderstood. GST-SAP18 binds mSin3 and also inter-acts weakly with HDAC1 (Zhang et al., 1997). GST-SAP30 binds in vitro translated mSin3A, mSin3B, andNCoR, but does not bind directly to HDAC1, HDAC2,or RbAp48. SAP30 has a structural and functionalhomolog in yeast. Its deletion in yeast confers a phe-notype very similar to those obtained subsequent todeletion of RPD3 or SIN3. This result suggests thatSAP30 is a critical component of a transcriptional reg-ulatory pathway in yeast. In mammalian cells, micro-injection of anti-SAP30 antibodies demonstrates thatSAP30 is required for repression by the estrogen recep-tor, but not by thyroid or retinoic acid receptors, sug-gesting that it may have a gene-specific role in core-pressor function (Laherty et al., 1998).

Nuclear hormone receptorsThe thyroid hormone (TH) and retinoic-acid recep-

tors (RARs) are ligand-dependent transcription factorsthat control cell development and homeostasis as theresult of both transcriptional repression and activation(Chambon, 1994). These factors activate transcriptionin the presence of ligand by associating with the his-tone acetyltransferases p300/PCAF, whereas in the ab-sence of ligand these receptors repress transcription.The ligand-binding domains of these receptors are ca-pable of transferring active repression to the GAL4DNA-binding domain (Glass et al., 1988; Baniahmad etal., 1992). The purification of a 240-kDa protein thatbound to this modular repression domain identifiedNCoR (Horlein et al., 1995; Kurokawa et al., 1995;Zamir et al., 1996) and also a related protein, SMRT(Chen and Evans, 1995). Receptor mutations that abol-ish NCoR binding also block transcriptional repression,but do not block activation, suggesting that NCoR me-diates transcription by the hormone receptors. Subse-quent experiments using NCoR antibodies demon-strated that Sin3 and histone deacetylase are alsocomponents of this complex (Alland et al., 1997; Hein-zel et al., 1997). It was suggested that the repressorcomplex involved in Mad-mediated transcriptional re-pression is identical to the complex used by the nuclearhormone receptors and that this complex containsSin3, HDAC1/2, RbAp46/48, SAP30, and SAP18. Theemerging model for transcriptional regulation by thehormone receptors is that, in the absence of ligand, theRAR/RXR dimer binds what many refer to as the “Sin3complex”; this complex represses transcription. In thepresence of ligand, the repressor complex is dissoci-ated. The receptor is then free to form a complex withthe histone acetyltransferase, which activates tran-scription.

HISTONE DEACETYLASE-INTERACTINGPROTEINS

In addition to their roles in the Sin3 complex,HDAC1 and HDAC2 also exist in a separate compositeknown as the Mi2 complex. In addition to two proteins

4 CRESS AND SETO

that are also present in the Sin3 complex (RbAp46 andRbAp48), the Mi2 complex contains Mi2, MTA2, andMBD3. Proteins such as NCoR and Mad interact withHDAC1/2 through the Sin3 complex, whereas somerepressors interact with HDAC1/2 through the Mi2complex. In addition, many other proteins interactwith HDAC1/2 either directly or through yet unidenti-fied proteins (summarized in Fig. 1).

Although nearly all HDAC-interacting proteins dis-covered to date are known to interact with HDAC1/2,there are clearly other cellular proteins that could in-

teract with other members of the HDAC family. Forexample, the myocyte enhancer factor MEF2A associ-ates exclusively with HDAC4 (Miska et al., 1999).Binding of HDAC4 to MEF2A results in the repressionof MEF2A transcriptional activation, a function thatrequires the deacetylase domain of HDAC4. Naturally,many more examples of proteins that interact withHDACs will be expected to emerge if a local alterationin chromatin structure is a general means of regulatinggene expression. Perhaps future studies will even re-veal that most transcriptional repressors function, inpart, by recruiting histone deacetylases; those devoid ofthis recruiting activity may turn out to be the interest-ing exceptions.

HISTONE DEACETYLASES AND CANCER—GUILT BY ASSOCIATION

During the past few years, results from basic re-search studies of histone deacetylases have addedgreatly to our general understanding of the regulationof transcription in eukaryotic cells. Increasing impor-tance is now being given to assess the relevance ofdeacetylases in health and disease. The interaction ofMad and Max is important in tumor suppression,whereas the thyroid hormone and retinoic acid recep-tors are indispensable in the control of developmentand homeostasis. The finding that histone deacetylasesare key components in the regulation of gene expres-sion by Mad and by hormone receptors underscores theimportant implications of deacetylases in human dis-ease. No direct alterations in the histone deacetylasegenes have yet been demonstrated in human oncogen-esis. However, it is now known that histone deacety-lases associate with a number of well-characterizedcellular oncogenes and tumor-suppressor genes. Thus,histone deacetylases may represent candidate targetsfor anticancer drugs and therapies.

In the following sections, we review the potentialroles that HDACs play in human cancer. In general,there are three separate known levels that are depen-dent on histone deacetylases and are likely linked totumorigenesis. On the first level are cell-cycle–re-straining transcriptional repressors including Mad andRb. On a second level are transcriptional repressorsthat normally block the process of differentiation incertain cellular lineages, for example, the nuclear hor-mone receptors such as RARa in hematopoietic differ-entiation. A final level of cell biology controlled byhistone deacetylase–containing complexes is thestrong correlation between genomic methylation andthe transcriptional silencing of various tumor-suppres-sor genes, such as p21WAF/CIP1.

Disruption of the Mad/Sin3/HDAC by v-SkiThe roles of the Mad/Sin3A/HDAC complex in

growth control and differentiation were discussed in anearlier section. The most commonly known alterationof this pathway in human cancer is the overexpressionof the alternative Max binding partner Myc, whichoccurs as a result of translocations, gene amplifica-tions, or activating point mutations (see Dang, 1999and references therein). When Myc is overexpressed,Mad/Max heterodimers are excluded and Mad-medi-ated transcriptional repression is blocked. However, inthis event it is not clear whether transformation occurs

Fig. 1. HDAC-associated proteins. Proteins that interact withHDAC1/2 through the Sin3 complex: MeCP2 (Jones et al., 1998; Nanet al., 1998; Wade et al., 1998a), Ikaros (Koipally et al., 1999), UME6(Kadosh and Struhl, 1997), Ski (Nomura et al., 1999), p53 (Murphy etal., 1999), NCoR/SMRT (Alland et al., 1997; Heinzel et al., 1997),MAD (Ayer et al., 1995; Schreiber-Agus et al., 1995); and through theMi2 complex: Hunchback (Kehle et al., 1998), PcG (van der Vlag andOtte, 1999), Ikaros (Kim et al., 1999b), HPV E7 (Brehm et al., 1999b),MBD2 (Ng et al., 1999; Zhang et al., 1999b) are displayed on the upperleft and upper center panels, respectively. All other HDAC-interact-ing proteins bind HDACs either directly or through unknown mech-anisms: YY1 (Yang et al., 1996a, 1997), Groucho (Chen et al., 1999),p107/p130/pRB (Brehm et al., 1998; Ferreira et al., 1998; Luo et al.,1998; Magnaghi-Jaulin et al., 1998a,b; Stiegler et al., 1998; Lai et al.,1999), TRb1 (Sasaki et al., 1999), LIM (Bach et al., 1999), REST(Huang et al., 1999), CBF (Hsieh et al., 1999), menin (Gobl et al.,1999), MBP-1 (Ghosh et al., 1999), Sp1 (Sowa et al., 1997; Doetzlhoferet al., 1999; Sowa et al., 1999; Xiao et al., 1999), LAZ3 (BCL-6)(Dhordain et al., 1998), Net (Criqui-Filipe et al., 1999), PLZF (PLZF-RARa) (Grignani et al., 1998; He et al., 1998; Lin et al., 1998), BRCA1(Yarden and Brody, 1999), ATM (Kim et al., 1999a), TGIF (Wotton etal., 1999), Rbp1 (Lai et al., 1999). References for HDAC3 interactionsinclude: HDAC4 and HDAC5 (Grozinger et al., 1999), YY1 (Yang etal., 1997), NCoR/SMRT (Kao et al., 2000), Rbp1 (Lai et al., 1999).References for HDAC4 interactions include: HDAC3 (Grozinger et al.,1999), MEF2A (Miska et al., 1999; Wang et al., 1999), NCoR/SMRT(Huang et al., 2000; Kao et al., 2000). References for HDAC5 interac-tions are: HDAC3 (Grozinger et al., 1999), NCoR/SMRT (Huang et al.,2000; Kao et al., 2000). Reference for HDAC7: Kao et al., 2000.


as a result of a blockage of repression per se, or ifadditional transcriptional activation by Myc/Max isalso required. Recent studies of the cellular proto-on-cogene c-Ski may address this issue.

Over 10 years ago, the oncogenic form of Ski, v-Ski,was found to transform chicken embryo fibroblasts (Liet al., 1986). However, the mechanism of this transfor-mation has remained unclear. A recent study has dem-onstrated that the cellular Ski protein is a componentof the Sin3/NCoR/HDAC complex (Nomura et al.,1999). Yeast two-hybrid and coimmunoprecipitationexperiments demonstrate that Ski may interact simul-taneously with both NCoR and Sin3A. Domain-map-ping experiments determined that c-Ski binds to NCoRvia its N-terminal region, and Sin3A through its C-terminal domain. It therefore seems possible that c-Skimay function, at least in part, as a bridging proteinthat tethers Sin3A and NCoR. It has been shown thatmutations within the N-terminal region of v-Ski (whichlacks the Sin3A-binding C-terminus) blocks transfor-mation. These observations suggested that v-Ski exertsits transforming effects by blocking the function orassembly of the Sin3/HDAC/NCoR/c-Ski complex. Totest this hypothesis, mutants of c-Ski lacking the C-terminal Sin3A-interacting domain were examined forthe ability to block transcriptional repression mediatedby Mad. As anticipated, v-Ski and C-terminally trun-cated forms of c-Ski blocked repression by Mad in adominant-negative fashion.

The observations discussed earlier demonstrate thatv-Ski is oncogenic, at least in part, because of its abilityto block Mad-mediated repression in the absence ofMyc overexpression. Furthermore, they suggest abroad potential for disruptions of the Sin3A/NCoR/HDAC complex in tumorigenesis. The Sin3A/NCoR/HDAC complex is likely involved in other regulatorypathways that may also contribute to v-Ski transfor-mation. For instance, it has been shown that c-Ski mayalso be involved in Rb-mediated transcriptional repres-sion (Tokitou et al., 1999), and other work has demon-strated that c-Ski can modulate RARa-mediated tran-scriptional repression (Dahl et al., 1998). Is c-Ski auniversal component of the complex, or is it necessaryonly in a subset of Sin3A/NCoR/HDAC-regulated path-ways? Future work is required to characterize the bio-logical roles of c-Ski.

Histone deacetylases interact with the Rbtumor-suppressor protein

Although disruptions of the Myc/Mad pathway con-tribute significantly to human cancer, they are notpresent in all tumors. In contrast, the Rb/E2F tran-scriptional regulatory pathway is disrupted in virtuallyevery human tumor, making it a nearly universal tar-get for anticancer drug and therapy development (Sell-ers and Kaelin, 1997). The 105-kDa Rb protein isknown to interact with numerous other proteins. Itsbest-understood binding partner is the dimeric tran-scription factor E2F, which is central in the control ofcell-cycle progression (for reviews see Sellers and Kae-lin, 1997; Dyson, 1998; Johnson and Schneider-Brous-sard, 1998; Nevins, 1998; Brehm et al., 1999a). E2F-regulated promoters are involved in the expression oftwo classes of activities: (1) enzymes required for DNAsynthesis, such as dihydrofolate reductase and thymi-

dine kinase; and (2) cell-cycle regulatory proteins, suchas cyclin A and cyclin E.

The current Rb/E2F paradigm is highlighted in Fig.2. In this model, an E2F family member binds DNA asa heterodimer with a DP family partner (Helin et al.,1993). In quiescent cells, this heterodimeric complex isbound by a member of the Rb family, which convertsthis potential transcriptional activator to a transcrip-tional repressor (Adnane et al., 1995; Weintraub et al.,1995; Luo et al., 1998). This Rb-mediated transcrip-tional repression is critical for the active downregula-tion of many E2F/Rb-regulated promoters (Lam andWatson, 1993; Johnson et al., 1994b; Lam et al., 1995;Sellers et al., 1995; Zwicker et al., 1996). Upon growthstimulation, the Rb protein is phosphorylated by thecyclin-dependent kinases (cdks); and the hyperphos-phorylated Rb then releases the E2F heterodimer,which goes on to activate transcription. The mecha-nism of E2F/DP-mediated transcription activation isnot fully understood, but is thought to involve therecruitment of HAT activity. Specifically, it has beendemonstrated that the E2F-1 activation domain asso-ciates with an ATM-related protein, TRRAP (McMahonet al., 1998; McMahon et al., 2000), and that TRRAPassociates with HAT activity (Vassilev et al., 1998).Rb’s phosphorylation state, and thus its growth inhib-itory activity, is tightly regulated by the activities ofthe cdks. The cdks are in turn regulated by cyclin-dependent kinase inhibitor proteins such as p21WAF/CIP1.Both active transcriptional repression and transcrip-

Fig. 2. Model for cell-cycle–specific transcriptional regulation byE2F and Rb. In quiescent cells, Rb interacts with DNA-bound E2F. Rbrepresses transcription of E2F-regulated promoters by recruitingHDAC1, HDAC2, or HDAC3. Rb may interact directly with HDACs,or may require the bridging factor Rbp1. Growth stimulation acti-vates G1 cyclin-dependent kinases to phosphorylate Rb and dissociateit from E2F and thus the promoter. Recent evidence suggests thatE2F may then associate with TRRAP, which likely possesses HATactivity (McMahon et al., 1998; Vassilev et al., 1998). Basal transcrip-tion factors and RNA polymerase may then associate with the pro-moter.

6 CRESS AND SETO

tional activation via E2F are likely to be critical forproper cell-cycle regulation (Dobrowolski et al., 1994;Sellers et al., 1995; Ishizaki et al., 1996; Wu et al.,1996; Zhang et al., 1999a). Evidence that the G0 Rb/E2F complex serves as a critical transcriptional repres-sor involved in cell-cycle control includes the following:

1. Rb overexpression in Rb2/2 cells blocks the tran-scription of E2F-regulated genes (Hiebert et al.,1992) and can, in some Rb-negative cell lines, leadto growth arrest (Qin et al., 1992).

2. In some Rb-regulated promoters, such as B-myb,an E2F binding site has been shown to be occupiedonly during G0 (when the promoter is off) and notin S phase, implying that the E2F site functionsexclusively as a repressor in this context (Zwickeret al., 1996).

3. Numerous transfection/reporter experiments havedemonstrated that E2F binding sites can serve asRb-dependent repressing elements in cis for bothartificial and natural promoters (Weintraub et al.,1992; Lam and Watson, 1993; Johnson et al.,1994b).

4. When fused to the DNA-binding domain of GAL4,the Rb protein represses transcription of artificialpromoters containing GAL4 binding sites (Adnaneet al., 1995; Weintraub et al., 1995).

5. When fused to the DNA-binding domain of E2F-1,the Rb protein blocks cell-cycle progression (Sellerset al., 1995).

6. Transcriptional repression by Rb is reversed by itsphosphorylation (Weintraub et al., 1995).

7. In serum-starved fibroblasts from Rb knockout an-imals, certain E2F-regulated promoters are consti-tutively activated (Hurford et al., 1997).

As mentioned earlier, the Rb protein binds to a hostof cellular proteins (Taya, 1997). One of the proteinsfound to bind to Rb was RbAp48 (Rb-associated protein48) (Huang et al., 1991; Qian et al., 1993). Both of theseproteins are ubiquitously expressed in the nucleus andthey form complexes with Rb both in vitro and in vivo.RbAp48 shares sequence homology with MSI1, a neg-ative regulator of the yeast Ras-cyclic AMP pathway(Qian et al., 1993; Qian and Lee, 1995). The earlyobservation that RbAp48 was also present in com-plexes containing HDAC1 (Taunton et al., 1996) ledseveral groups to specifically test whether the Rb pro-tein associates with histone deacetylases in vivo. Notsurprisingly, several laboratories have provided evi-dence that Rb associates with HDACs in vivo (Brehmet al., 1998; Luo et al., 1998; Magnaghi-Jaulin et al.,1998). However, it is now known that Rb’s interactionwith HDACs is not dependent on the binding toRbAp46 or RbAp48, but rather, is mediated throughthe bridging protein Rbp1 (Lai et al., 1999). The follow-ing evidence from three initial studies (Brehm et al.,1998; Luo et al., 1998; Magnaghi-Jaulin et al., 1998)firmly established the existence of a bona fide interac-tion between Rb and HDAC1/2:

1. A GST-Rb fusion protein was demonstrated to spe-cifically associate with HDAC1 and HDAC2. Fur-thermore, the association of GST-Rb with histonedeacetylases was dependent on Rb domains known

to be critical for cell-growth control and transcrip-tional repression.

2. Rb antibodies were used to coimmunoprecipitatehistone deacetylase activity from crude extracts.

3. Viral oncoproteins known to cause the dissociationof Rb complexes had similar effects on the Rb/HDAC interaction. As expected, inactive mutantsof these oncoproteins did not result in the dissoci-ation of Rb/HDAC.

4. An E2F/Rb/histone deacetylase complex could beformed in vitro from recombinant proteins.

5. GST-E2F associated with histone deacetylase fromcrude extracts when Rb protein was provided as atether.

6. Transfection of HDAC1 together with Rb resultedin collaborative repression of an E2F-regulatedpromoter.

7. The histone deacetylase inhibitor trichostatin A(TSA) activated the transcription of several E2F-regulated promoters, including thymidine kinaseand dihydrofolate reductase in Rb-expressing cells,but not in cells lacking Rb.

8. Recruitment of Rb to a promoter resulted in de-creased histone acetylation of nucleosomes boundto that promoter.

HDAC1 and HDAC2 contain an IXCXE sequence,similar to the LXCXE motif, through which severalviral and cellular proteins interact with the A/B pocketof Rb (Nevins, 1994). This domain may be sufficient forinteraction with Rb since its deletion abolished theHDAC1/Rb interaction in vitro. Although there is po-tential for a direct interaction between Rb and HDAC1and HDAC2, recent evidence suggests that HDAC3also associates with Rb in vivo (Lai et al., 1999). SinceHDAC3 lacks an LXCXE-like motif, it seems likely thatit may be tethered to Rb via interaction with a bridgingprotein. A prime bridging protein candidate is Rbp1,which contains a consensus LXCXE motif and is knownto interact with HDAC1, -2, and -3 (Lai et al., 1999).Rbp1 possesses two transcriptional repression do-mains, one that binds HDACs and a second that re-presses transcription in an HDAC-independent man-ner. It is not yet known whether other HDAC-associated proteins such as Sin3A, NCoR, RbAp46, andRbAp48 are components of the Rb/HDAC/Rbp1 com-plex. In one study, immunoprecipitations were utilizedto demonstrate that Rb associates with c-Ski, a relatedprotein Sno, and Sin3A. However, interactions betweenRb and NCoR were not detected in this effort. In an-other study, the expression of v-Ski was correlated withdiminished interaction between Rb and HDACs, aswell as lower levels of Rb-mediated transcriptional re-pression (Tokitou et al., 1999). Taken together, theseresults suggest a complex scenario in which Rb associ-ates with HDACs through a variety of mechanisms.Additional work will be required for a more completeunderstanding of the regulation and functions of theRb family of proteins. The molecular mechanisms un-derlying the fine-tuning of cell growth control will un-doubtedly be the subject of future studies for years tocome.


Alterations of the Rb/HDAC interaction inhuman cancer

There are numerous genetic alterations that resultin loss of Rb/HDAC interactions in cancer. The mostobvious genetic alteration is simply the loss of a func-tional Rb gene, by deletions and point mutations,which occurs in many solid tumors in both sporadic andheritable patterns. A second common mechanism of Rbinactivation occurs in cervical cancers caused by high-risk strains of the human papilloma virus. These vi-ruses express the E7 oncoprotein, which binds Rb andblocks its interaction with E2F and with HDACs(Phelps et al., 1991; Brehm et al., 1998). In a thirdmechanism, numerous genetic alterations not involv-ing the Rb gene directly lead to constitutive Rb phos-phorylation (inactivation). For example, the loss of cdkinhibitor proteins (most commonly p16) or the overex-pression of cyclin D1 or cdk4 can each result in consti-tutive Rb phosphorylation and enhanced tumor growth(Hall and Peters, 1996). Adding further to the complex-ity of the Rb/E2F pathway is the fact that Rb, E2F, andDP are not single polypeptides but are each a memberof protein families. Rb has two close protein relatives,p107 (Ewen et al., 1991) and p130 (originally namedpRb2/p130) (Hannon et al., 1993; Li et al., 1993; Mayolet al., 1993). Collectively, these proteins are referred toas the pocket proteins, since each shares a large con-served structural domain (the “pocket”) required forinteraction with E2F and with viral oncogenes such asE7 (Kaelin et al., 1991). Both p107 and p130 possessthe ability to repress transcription and to block cellgrowth (Zhu et al., 1993; Claudio et al., 1994; Zhu et al.,1995). Furthermore, recent work has shown that bothp107 and p130 bind to HDACs (Ferreira et al., 1998;Stiegler et al., 1998; Iavarone and Massague, 1999).

Although they share many structural and functionalsimilarities, the members of the Rb family clearly havedifferent functional roles. Rb is the only member of itsfamily that is known to be commonly mutated in hu-man cancer; it is also the only protein of its class thathas been shown to be required for mouse viability (Leeet al., 1992; Cobrinik et al., 1996; Lee et al., 1996; Helinet al., 1997). However, p130 is also a bone fide tumor-suppressor gene, since it has been found to be mutatedin primary nasopharyngeal carcinoma (Claudio et al.,2000a) and in lung tumors (Claudio et al., 2000b), aswell as in cell lines derived from small cell lung carci-noma (Helin et al., 1997). Replacement of the wild-typep130 allele by retrovirus leads to suppression of tumorgrowth. It is not yet clear why Rb has such a specialrole. All members of the Rb family interact with mem-bers of the E2F family; however, there is some speci-ficity, in that p107 and p130 do not physiologicallyinteract with E2F-1, E2F-2, or E2F-3. In contrast, Rbinteracts with the five known E2F proteins (Beijersber-gen et al., 1994; Ginsberg et al., 1994; Hijmans et al.,1995; Sardet et al., 1995). Perhaps the unique role ofRb lies in its ability to interact with all members of theE2F family. Alternatively, Rb-family knockout micereveal that Rb, p107, and p130 regulate slightly differ-ent subsets of E2F-driven promoters (Hurford et al.,1997). Specifically, Rb appears to be uniquely able toregulate the cyclin E and p107 promoters. Thus, Rb’sunique roles may be a function of its ability to regulate

a relatively small number of critical cell-cycle regula-tory factors.

The E2F family itself is theoretically a target formutations that would activate cell growth. For exam-ple, a mutation that would increase E2F expression ormake E2F unable to interact with Rb (and thus unableto tether Rb and associated HDACs to promoters)would be predicted to be oncogenic. This principle hasbeen demonstrated in numerous experiments designedto overexpress E2F or to express altered Rb-insensitiveforms of E2F (Beijersbergen et al., 1994; Johnson et al.,1994a; Singh et al., 1994; Jooss et al., 1995; Wang etal., 1995; Xu et al., 1995; Guy et al., 1996; Pierce et al.,1998a,b). There is also limited evidence that alter-ations in E2F occur in human tumors. The E2F-1 genehas been shown to be amplified in a human erythro-leukemia cell line (Saito et al., 1995) and E2F-4 isfrequently altered within a serine repeat motif near itsC-terminus (Yoshitaka et al., 1996; Souza et al., 1997).Although there is little evidence that known membersof the E2F family themselves are commonly altered inhuman oncogenesis, it should be noted that the field isstill relatively young, and the list of known E2F vari-ants involved in oncogenesis may very well remain tobe discovered.

Paradoxical observationsIn the cases of both Mad and Rb, transcriptional

repression is critical in evoking growth arrest. Becausethe capacity of these proteins to repress transcriptionis dependent on HDAC association, one would predictthat histone deacetylase inhibitors would prevent cell-cycle arrest. Although histone deacetylase inhibitorsactivate certain Rb/E2F-regulated promoters, they donot promote cell growth, but rather lead to growtharrest. These observations seem paradoxical, especiallyconsidering the central role that Rb/E2F is thought toplay in growth control. The most general way to recon-cile these observations is to point out that histonedeacetylase inhibitors may affect various transcrip-tional pathways in addition to Mad and Rb. The preciseidentity of the critical pathway or pathways is notknown, but one would speculate that HDAC inhibitorswould affect events upstream of Myc and E2F in the G1phase of the cell cycle. A putative target would be thetumor-suppressor protein p21WAF/CIP1, which has re-cently been shown to be strongly stimulated at the levelof transcription by the addition of histone deacetylaseinhibitors (Archer et al., 1998); p21WAF/CIP1 is a cyclin-dependent kinase inhibitor, whose expression leads tohypophosphorylated (growth-suppressing) Rb. Nor-mally, p21WAF/CIP1 is induced by p53 in response toDNA damage and, thus, it is also known as WAF1 orCIP1 (wild-type p53-activated factor or cdk inhibitorprotein-1) (Di Leonardo et al., 1994). In this model,addition of histone deacetylase inhibitor inducesp21WAF/CIP1, which in turn leads to the activation of Rband growth arrest. Although the mechanism by whichHDAC inhibitors activate p21 is not clear, it is clearlyp53 independent (Xiao et al., 1997) and is dependent onSp1-binding sites within the promoter (Sowa et al.,1997, 1999).

An obvious difficulty with this model is that theHDAC inhibitor might block Rb’s transcriptional re-pression function and, thus, p21WAF/CIP1 induction

8 CRESS AND SETO

would be pointless. However, there are several exam-ples of promoters that are transcriptionally repressedby Rb/E2F independent of histone deacetylase. For ex-ample, among genes controlled by well-characterizedE2F-regulated promoters, TSA induces p107 andDHFR transcription, but does not affect the transcrip-tion of ribonucleotide reductase, thymidine kinase, orPCNA (Luo et al., 1998). Thus, it is clear that Rb hastranscriptional-repressing properties that are notblocked by histone deacetylase inhibitors. This may beexpected since Rb binds to many proteins, which po-tentially may function as HDAC-independent tran-scriptional repressors, such as Rbp1 (Lai et al., 1999).Furthermore, it has been known for some time that Rbcan inhibit transcriptional activation by E2F in vitro inthe absence of chromatin (Dynlacht et al., 1994). Usinghighly purified components, it was shown that Rb di-rectly blocks the interaction between E2F and thebasal transcription factors TFIID and TFIIA, thus pre-venting the formation of a preinitiation complex invitro (Ross et al., 1999). These findings strongly sug-gest that Rb utilizes a number of mechanisms to re-press transcription. The exact mechanism used may beeither promoter dependent or contingent on the mech-anisms stimulating growth arrest. Future work is nec-essary to characterize the mechanisms underlying Rb’sgrowth-restraining properties.

Histone deacetylases and interaction withfusion proteins in leukemia

Acute leukemia results when the differentiation ofimmature hematopoietic cells is blocked and the cellsproliferate without restraint. Acute promyelocytic leu-kemia (APL) is most often associated with chromo-somal translocation t(15;17), which fuses a large por-tion of the retinoic acid receptor a (RARa) to the codingsequence of a second gene, PML (promyelocytic leuke-mia). In a smaller percentage of APL cases, a similartranslocation t(11;17) occurs in which the same codingregion of RARa is fused to a different protein, PLZF(promyelocytic leukemia zinc finger). PLZF is aKrupple-like DNA-binding protein containing ninezinc-finger motifs. It normally plays a role in centralnervous system development and in hematopoiesis.RARa also plays a role in hematopoiesis. Specifically,RARa binds to DNA as a heterodimer with an RXR(retinoid-X receptor) protein. In the absence of retinoicacid, which normally induces differentiation of promy-elocytic cells, the RARa/RXR heterodimer binds to atranscriptional corepressor that contains NCoR,Sin3A, and histone deacetylase (Grignani et al., 1998).In the presence of retinoic acid, the corepressor com-plex is displaced and is replaced by a coactivator com-plex that contains the histone acetyltransferases p300/CBP and PCAF. However, when the RARa protein ispresent in either of these fusion proteins it is no longerresponsive to physiological levels of retinoic acid. Thus,under these conditions, RARa becomes a constitutivetranscriptional repressor, which blocks normal differ-entiation and leads to leukemia.

Although both of the RARa fusion proteins discussedearlier are insensitive to physiological levels of retinoicacid, the PML–RARa fusion (and thus patients withthis translocation) will respond to pharmacologicaldoses of all-trans-retinoic acid (ATRA). In contrast, the

PLZF–RARa fusion protein is completely insensitive toretinoic acid; patients with this translocation do notbenefit from retinoic acid treatment. This differenceresults from the fact that the PLZF protein itself con-tains an interaction domain that can bind to the NCoR/Sin3A/HDAC complex. Thus, the fusion protein inter-acts with corepressors through two domains, one ofwhich is not sensitive to retinoic acid. Consistent withthis model, the histone deacetylase inhibitor trichosta-tin A restores RA sensitivity to PLZF–RARa and al-lows these leukemic cells to respond to ATRA (Grignaniet al., 1998; He et al., 1998; Lin et al., 1998). Theseobservations clearly demonstrate a role for the histonedeacetylases in oncogenesis and suggest that histonedeacetylase inhibitors may be valuable in treating cer-tain forms of leukemia.

AMLThe AML1 gene is disrupted by the t(8;21) translo-

cation in acute myeloid leukemia (for review see Fen-rick and Hiebert, 1998). Normally, AML1 functions asthe DNA-binding component of CBF (the enhancercore-binding factor). This complex apparently activatesexpression of genes required for myeloid differentiation(Lenny et al., 1997). In the DNA-binding complex,AML1 forms a heterodimer with a second proteinCBFb; this complex appears to function as a transcrip-tional activator via interaction with the p300/PCAFhistone acetyltransferase complex (Kitabayashi et al.,1998). In the t(8;21) translocation, the DNA-bindingdomain of AML1 is fused to a second protein referred toas ETO (“8;21” or eight, twenty-one).

Numerous AML translocations in addition to t(8;21)occur in acute myeloid leukemia. The common elementof these fusion proteins is that they all fuse the DNA-binding domain of AML1 with a second protein thatinterferes with AML-dependent transcriptional activa-tion (Fenrick and Hiebert, 1998). The t(12;21) translo-cation fuses most of the AML protein to a second tran-scription factor TEL (translocation, ets, leukemia). Thet(16;21) translocation fuses AML1 to MTG16 (myeloidtumor gene 16), a protein very similar in structure toETO (Gamou et al., 1998). Finally, the t(3;21) translo-cation fuses AML1 to Evi I, a known transcriptionalrepressor (Nucifora and Rowley, 1995). The inv(16)inversion fuses the CBFb protein to a smooth musclemyosin heavy-chain gene. Although this does not affectthe AML1 gene directly, the inv(16) retains its abilityto interact with AML1 and, thus, is thought to induceleukemia through the disruption of AML1-mediatedtranscriptional activation.

Each of the fusion proteins mentioned previously hasbeen shown to inhibit AML1-dependent transcription(Fenrick and Hiebert, 1998). So far, the histonedeacetylases have been directly implicated in only theAML–ETO translocation. However, recent evidencesuggests that ETO, like RARa and PLZF, binds toNCoR and to mSin3A and recruits histone deacetylases(Lutterbach et al., 1998). Because of its similarity instructure to ETO, it is predicted that the AML–MTG16fusion will also bind constitutively to the NCoR,mSin3A, and histone deacetylase complex; however,this has yet to be demonstrated experimentally. If his-tone deacetylases are commonly involved in AML fu-


sions, then histone deacetylase inhibitors will no doubthave value in AML therapy.

Another link between HDAC and AML1 came re-cently with the discovery that the Drosophila corepres-sor Groucho interacts functionally with the Droso-philia HDAC protein (Chen et al., 1999). HDACpotentiates repression by Groucho, and histonedeacetylase inhibitors were used to show that deacety-lase activity is required for efficient Groucho-mediatedrepression. Mutations in Groucho and HDAC geneshave synergistic effects on embryonic lethality and pat-tern formation in flies. Given that the human homologof Groucho is known to bind to AML1, it is tempting tospeculate that it plays roles in hematopoiesis and mayeven be involved in leukemia.

Chromatin remodeling, histone deacetylases,and methylation: potential roles in cancer

Recent work has demonstrated the existence of cel-lular complexes with both histone deacetylase andATP-dependent nucleosome-remodeling activity (Tonget al., 1998; Wade et al., 1998b; Zhang et al., 1998a,1999b). One of these complexes contains at least sevensubunits and has been named NRD or NuRD (nucleo-some-remodeling histone deacetylase complex; identi-cal to the Mi2 complex). The Mi2 complex does notcontain Sin3A or SAP30 and is thus distinct from theSin3A/HDAC complex. Four of the NuRD subunits, thehistone deacetylases HDAC1/2 and the histone-bindingproteins RbAp48 and RbAp46 are common to both theMi2 complex and the Sin3A/HDAC complex. The threeremaining subunits have apparent molecular masses of230, 70, and 32 kDa, respectively, and appear to beunique to the Mi2 complex. Sequence analyses revealedthat the largest of these subunits is identical to thedermatomyositis-specific autoantigen Mi2 (which isidentical to the protein CHD3) (Targoff and Reichlin,1985). CHD3 is closely related to another polypeptideknown as CHD4 (Woodage et al., 1997). The CHD3/4subunit of the Mi2 complex contains a helicase/ATPasedomain and possesses nucleosome-remodeling activity.Peptide sequence analyses of the 70-kDa subunit of theMi2 complex revealed that it is related to the MTA1(metastasis-associated protein 1); accordingly, it wastermed MTA2. The MTA2 subunit is essential for highlevels of histone deacetylase activity. The overexpres-sion of MTA1 correlates with the metastatic potentialof numerous cancer cells; it will therefore be of greatinterest to determine whether MTA1 overexpressionalters Mi2 complex function. The 32-kDa subunit cor-responds to MBD3, a member of a family of proteinscontaining methyl-CpG–binding domains (Zhang etal., 1999b). MBD3 is required for association of MTA2with the core histone deacetylase complex. Despite pos-sessing a methyl-CpG-binding domain, the MBD3 sub-unit is apparently unable to bind methylated DNA.Rather, the Mi2 complex can be tethered to methylatedDNA via an interaction with an eighth protein, MBD2,which is highly homologous to MBD3. Although able totether NuRD to methylated DNA, MBD2 was not foundto be stably associated with either HDAC1 or HDAC2.

These observations suggest that some chromatin re-modeling is mediated by cellular complexes with his-tone deacetylase activity. It is speculated that the cou-pling of remodeling to deacetylation may be required to

allow access of the RbAp46 and RbAp48 subunits to thecore histones. Furthermore, this complex may bind di-rectly to methylated genomic DNA. The methylation ofpromoter CpG dinucleotides has been tightly corre-lated with transcriptional repression (Siegfried and Ce-dar, 1997; Siegfried et al., 1999). Results from recentstudies suggest that methylation-mediated transcrip-tional silencing of tumor-suppressor genes may be acritical event in the formation of certain cancers. Al-though various mechanisms may underlie this repres-sion (dependent on the promoter), recent work hasdemonstrated that one mechanism mediating this re-pression may involve the recruitment of histonedeacetylases to methylated CpG dinucleotides. MeCP2is the best-characterized member of a family of methyl-CpG–binding proteins (Meehan et al., 1989; Lewis etal., 1992; Cross et al., 1997; Hendrich and Bird, 1998).It has recently been demonstrated that MeCP2 is atranscriptional repressor that recruits histone deacety-lase via the Sin3 complex (Nan et al., 1997, 1998; Joneset al., 1998). Four proteins with homology to theMeCP2 methyl-CpG–binding domain, MBD1–MBD4(Hendrich and Bird, 1998), have been identifiedthrough searches of EST databases. Of these four,MBD2 appears to be a demethylase (Bhattacharya etal., 1999) and MBD4 is an endonuclease likely involvedin DNA repair (Bellacosa et al., 1999).

The roles of methyl-CpG–binding/HDAC complexesin human cancer are not yet fully understood. Onepossibility is the strong correlation between methyl-ation and transcriptional silencing of tumor-suppres-sor genes. Aberrant CpG methylation is observed in anumber of tumor-suppressor gene promoters includingRb, the cdk inhibitors p15 and p16, and the DNA repairgene MLH1 (reviewed in Jones and Laird, 1999). Rb isthe most extensively characterized example to date. Ina number of tumor types, one Rb allele is abolished bya structural mutation, and the otherwise normal alleleis transcriptionally silenced by aberrant methylation(Stirzaker et al., 1997). The methylation of the silencedRb promoter is extensive, and encompasses a pair ofcritical E2F-binding elements. Recent work has dem-onstrated that methylation of these E2F sites in the Rbpromoter abolishes E2F binding (and thus E2F-medi-ated activation of the promoter) and creates a bindingsite for MeCP2, thus silencing the allele (Di Fiore et al.,1999). An important question that certainly needs to beaddressed is: “what events lead to the hypermethyl-ation of the Rb-promoter?”

Recent evidence suggests that fos may transformcells by a mechanism involving activation of DNA5-methylcytosine transferase (Bakin and Curran,1999). Perhaps activation of CpG-methylation, whetherpromoter-specific or not, will be found to be an impor-tant oncogenic activity. Equally important, blockingthis activity pharmacologically may prove to be aneffective anticancer therapy in the future.

HISTONE DEACETYLASE INHIBITORS ASCANCER CHEMOTHERAPEUTIC AND

CHEMOPREVENTIVE AGENTSThe initial cloning of histone deacetylases and the

subsequent rapid advances in understanding themechanisms of histone deacetylases and transcrip-tional repression have been greatly attributed to the

10 CRESS AND SETO

availability of histone deacetylase inhibitors. Early on,it was clear that in addition to alterations in deacety-lation activity and transcription, many histonedeacetylase inhibitors alter cellular functions. Treat-ment of mammalian cell cultures with deacetylase in-hibitors causes cell-cycle arrest at either stage G1 orstage G2, consistent with the fact that histone deacety-lation is tightly linked to cell-cycle control (Ogryzko etal., 1996a). Furthermore, a large number of studieshave shown that histone deacetylase inhibitors caneffectively arrest and revert transformation of somecells (e.g, Yoshida et al., 1990; Yoshida and Sugita,1992; Kijima et al., 1993; Futamura et al., 1995; Richonet al., 1996; McBain et al., 1997; Richon et al., 1998)and can block the formation of tumors in rodent models(Cohen et al., 1998; Desai et al., 1999). These observa-tions suggest that histone deacetylase inhibitors maybe effective chemotherapeutic agents in human cancer.The histone deacetylase inhibitor butyrate has beentested in clinical trials for b-thalassemia (Collins et al.,1995) and for prostate and brain cancers (Samid et al.,1997).

HDAC inhibitors target critical cell-cycle regulatorypathways and may induce the expression of varioussilenced tumor-suppressor genes. Thus, these agentsmay have a broad application in chemotherapy andmay hold special promise for the treatment of acuteleukemias. As discussed in detail previously, fusionproteins that utilize HDACs have been implicated inmany acute leukemias (Fenrick and Hiebert, 1998).These fusion proteins block the differentiation of im-mature hematopoietic cells, resulting in unstrainedgrowth. Histone deacetylase inhibitors block the abilityof these fusion proteins to repress transcription ofgenes required for differentiation. This repression canoften be relieved by addition of high levels of all-trans-retinoic acid, which allows these cells to differentiate(thus the term differentiation therapy). However, inother cases these fusion proteins interact with HDACsvia retinoic acid–insensitive domains (Guidez et al.,1998). In these cases, HDAC inhibitors are capable ofrestoring RA responsiveness, suggesting they mayserve to augment retinoic acid differentiation therapy.This principle has been demonstrated in a mousemodel in which trichostatin A was shown to be aneffective treatment for leukemias induced by trans-genic RARa-fusion proteins (He et al., 1998). Further-more, because of the central role of HDACs in hemato-poiesis, it is likely that HDAC inhibitors will be oftherapeutic value, regardless of the exact nature of thefusion causing the leukemia.

Butyrate and colon cancerHigh-fiber diets are associated with a significant de-

crease in the incidence of colon cancer (Trock et al.,1990a,b). Butyrate, a four-carbon fatty acid, which isproduced in millimolar quantities by the bacterial fer-mentation of fiber, is thought to mediate a substantialportion of dietary fiber’s chemopreventive activity(McIntyre et al., 1991). The growth-inhibiting proper-ties of butyrate have been demonstrated both in vitro,using colorectal cancer cell lines (Barnard and War-wick, 1993; Whitehead et al., 1986), and in vivo, usingchemically induced colorectal tumors in rat (McIntyreet al., 1993).

Although the mechanism of butyrate’s action is notyet fully understood, early work demonstrated that oneresult of exposure to butyrate was the induction ofhistone hyperacetylation (Riggs et al., 1977). This ef-fect can now be explained from the understanding thatbutyrate is a noncompetitive inhibitor of histonedeacetylases (Sealy and Chalkley, 1978). HDAC5 wasinitially described as an antigen in human colon cancerand named NY-CO-9 (Scanlan et al., 1998). Moreover,recent work may provide a direct connection betweenbutyrate exposure and stimulation of Rb tumor-sup-pressor gene function (Archer et al., 1998). Specifically,expression of the cyclin-dependent kinase inhibitorp21WAF/CIP1 mRNA is sharply upregulated followingbutyrate treatment. This induction occurs within 2 h oftreatment, even in the presence of cycloheximide. Anexperiment comparing p21 (2/2) human colon carci-noma cells with isogenic p21 (1/1) cells demonstratedthat p21 is absolutely required for butyrate-inducedgrowth arrest. It is likely that this therapeutic effortwould be irrelevant to tumor cells that have alreadylost p21 or Rb function. However, these experimentssuggest that butyrate and other more specific HDACinhibitors may have a potent antitumor effect if admin-istered at appropriate doses over the lifetime of suscep-tible individuals. Such preventative treatment may bebroadly applicable to various types of cancer, assumingthat future clinical trials do not reveal broad side ef-fects or toxicity problems.

CONCLUSIONSThe purification, cloning, and partial characteriza-

tion of histone deacetylases in the past few years haveprovided important insights into the transcriptionalregulation of chromatin. Undoubtedly, these lines ofstudy will continue to hone our understanding of enzy-matic machines in gene regulation. As with the discov-ery of any molecules related to cancer, the initial linkbetween HDACs and cancer is greeted with great ex-citement and overwhelming expectations. The futurehope and challenge will be to convert this tenuous linkinto a substantiated, firm connection, and to discoverways to destroy cancer through inhibitors of HDACs.

ACKNOWLEDGMENTSWe thank Tanya Butler, Rhonda Croxton, Nancy

Olashaw, Matt Thomas, Bill Tsai, Wen-Ming Yang,and Alice Yao for their helpful suggestions regardingthe manuscript. We apologize to the many colleagueswhose work could not be referenced because of spacelimitations.

LITERATURE CITEDAdnane J, Shao Z, Robbins PD. 1995. The retinoblastoma susceptibil-

ity gene product represses transcription when directly bound to thepromoter. J Biol Chem 270:8837–8843.

Alland L, Muhle R, Hou H Jr, Potes J, Chin L, Schreiber-Agus N,DePinho RA. 1997. Role for N-CoR and histone deacetylase inSin3-mediated transcriptional repression. Nature 387:49–55.

Aravind L, Koonin EV. 1998. Second family of histone deacetylases.Science 280:1167.

Archer SY, Meng S, Shei A, Hodin RA. 1998. p21(WAF1) is requiredfor butyrate-mediated growth inhibition of human colon cancercells. Proc Natl Acad Sci USA 95:6791–6796.

Ayer DE, Lawrence QA, Eisenman RN. 1995. Mad-Max transcrip-tional repression is mediated by ternary complex formation withmammalian homologs of yeast repressor Sin3. Cell 80:767–776.


Bach I, Rodriguez-Esteban C, Carriere C, Bhushan A, Krones A, RoseDW, Glass CK, Andersen B, Izpisua Belmonte JC, Rosenfeld MG.1999. RLIM inhibits functional activity of LIM homeodomain tran-scription factors via recruitment of the histone deacetylase complex.Nat Genet 22:394–399.

Bakin AV, Curran T. 1999. Role of DNA 5-methylcytosine transferasein cell transformation by fos. Science 283:387–390.

Baniahmad A, Kohne AC, Renkawitz R. 1992. A transferable silenc-ing domain is present in the thyroid hormone receptor, in thev-erbA oncogene product and in the retinoic acid receptor. EMBO J11:1015–1023.

Bannister AJ, Kouzarides T. 1996. The CBP co-activator is a histoneacetyltransferase. Nature 384:641–643.

Barnard JA, Warwick G. 1993. Butyrate rapidly induces growth in-hibition and differentiation in HT-29 cells. Cell Growth Differ4:495–501.

Bartl S, Taplick J, Lagger G, Khier H, Kuchler K, Seiser C. 1997.Identification of mouse histone deacetylase 1 as a growth factor-inducible gene. Mol Cell Biol 17:5033–5043.

Beijersbergen RL, Kerkhoven RM, Zhu L, Carlee L, Voorhoeve PM,Bernards R. 1994. E2F-4, a new member of the E2F gene family,has oncogenic activity and associates with p107 in vivo. Genes Dev8:2680–2690.

Bellacosa A, Cicchillitti L, Schepis F, Riccio A, Yeung AT, MatsumotoY, Golemis EA, Genuardi M, Neri G. 1999. MED1, a novel humanmethyl-CpG-binding endonuclease, interacts with DNA mismatchrepair protein MLH1. Proc Natl Acad Sci USA 96:3969–3974.

Bhattacharya SK, Ramchandani S, Cervoni N, Szyf M. 1999. A mam-malian protein with specific demethylase activity for mCpG DNA.Nature 397:579–583.

Boyes J, Byfield P, Nakatani Y, Ogryzko V. 1998. Regulation ofactivity of the transcription factor GATA-1 by acetylation. Nature396:594–598.

Brehm A, Miska EA, McCance DJ, Reid JL, Bannister AJ, KouzaridesT. 1998. Retinoblastoma protein recruits histone deacetylase torepress transcription. Nature 391:597–601.

Brehm A, Miska E, Reid J, Bannister A, Kouzarides T. 1999a. The cellcycle-regulating transcription factors E2F-RB. Br J Cancer80(Suppl 1):38–41.

Brehm A, Nielsen SJ, Miska EA, McCance DJ, Reid JL, Bannister AJ,Kouzarides T. 1999b. The E7 oncoprotein associates with Mi2 andhistone deacetylase activity to promote cell growth. EMBO J 18:2449–2458.

Brownell JE, Zhou J, Ranalli T, Kobayashi R, Edmondson DG, RothSY, Allis CD. 1996. Tetrahymena histone acetyltransferase A: ahomolog to yeast Gcn5p linking histone acetylation to gene activa-tion. Cell 84:843–851.

Candau R, Zhou JX, Allis CD, Berger SL. 1997. Histone acetyltrans-ferase activity and interaction with ADA2 are critical for GCN5function in vivo. EMBO J 16:555–565.

Carmen AA, Rundlett SE, Grunstein M. 1996. HDA1 and HDA3 arecomponents of a yeast histone deacetylase (HDA) complex. J BiolChem 271:15837–15844.

Chambon P. 1994. The retinoid signaling pathway: molecular andgenetic analyses. Semin Cell Biol 5:115–125.

Chen G, Fernandez J, Mische S, Courey AJ. 1999. A functional inter-action between the histone deacetylase Rpd3 and the corepressorgroucho in Drosophila development. Genes Dev 13:2218–2230.

Chen JD, Evans RM. 1995. A transcriptional co-repressor that inter-acts with nuclear hormone receptors. Nature 377:454–457.

Claudio PP, Howard CM, Baldi A, DeLuca A, Fu Y, Condorelli G, SunY, Colburn N, Calabretta B, Giordano A. 1994. p130/pRb2 hasgrowth suppressive properties similar to yet distinctive from thoseof retinoblastoma family members pRb and p107. Cancer Res 54:5556–5560.

Claudio P, Howard C, Fu Y, Cinti C, Califano L, Micheli P, Mercer E,Caputi M, Giordano A. 2000a. Mutations in the retinoblastoma-related gene RB2/p130 in primary nasopharyngeal carcinoma. Can-cer Res 60:8–12.

Claudio PP, Howard CM, Pacilio C, Cinti C, Romano G, Minimo C,Maraldi NM, Minna JD, Gelbert L, Leoncini L, Tosi GM, Hicheli P,Caputi M, Giordano GG, Giordano A. 2000b. Mutations in theretinoblastoma-related gene RB2/p130 in lung tumors and suppres-sion of tumor growth in vivo by retrovirus-mediated gene transfer.Cancer Res 60:372–382.

Cobrinik D, Lee MH, Hannon G, Mulligan G, Bronson RT, Dyson N,Harlow E, Beach D, Weinberg RA, Jacks T. 1996. Shared role of thepRB-related p130 and p107 proteins in limb development. GenesDev 10:1633–1644.

Cohen L, Richon V, Rifkind R, Marks P, Desai D, Amin S. 1998.Inhibition of N-methylnitrosourea-induced tumors by the cytodif-

ferentiating agent, suberanilohydroxamic acid [abstract 736]. ProcAm Assoc Cancer Res 39:108.

Collins AF, Pearson HA, Giardina P, McDonagh KT, Brusilow SW,Dover GJ. 1995. Oral sodium phenylbutyrate therapy in homozy-gous beta thalassemia: a clinical trial. Blood 85:43–49.

Criqui-Filipe P, Ducret C, Maira SM, Wasylyk B. 1999. Net, a nega-tive Ras-switchable TCF, contains a second inhibition domain, theCID, that mediates repression through interactions with CtBP andde-acetylation. EMBO J 18:3392–3403.

Cross SH, Meehan RR, Nan X, Bird A. 1997. A component of thetranscriptional repressor MeCP1 shares a motif with DNA methyl-transferase and HRX proteins. Nat Genet 16:256–259.

Dahl R, Kieslinger M, Beug H, Hayman MJ. 1998. Transformation ofhematopoietic cells by the Ski oncoprotein involves repression ofretinoic acid receptor signaling. Proc Natl Acad Sci USA 95:11187–11192.

Dang CV. 1999. c-Myc target genes involved in cell growth, apoptosis,and metabolism. Mol Cell Biol 19:1–11.

Dangond F, Hafler DA, Tong JK, Randall J, Kojima R, Utku N,Gullans SR. 1998. Differential display cloning of a novel humanhistone deacetylase (HDAC3) cDNA from PHA-activated immunecells. Biochem Biophys Res Commun 242:648–652.

De Rubertis F, Kadosh D, Henchoz S, Pauli D, Reuter G, Struhl K,Spierer P. 1996. The histone deacetylase RPD3 counteracts genomicsilencing in Drosophila and yeast. Nature 384:589–591.

Desai D, El-Bayoumy K, Amin S. 1999. Chemopreventative efficacy ofsuberanilohydroxamic acid (SAHA), a cytodifferentiating agent,against tobacco-specific nitrosamine 4-(methylinitros-amino)-1-(3-pyridyl)-1-butanone (NNK)-induced lung tumorigenesis in femaleA/J mice [abstract 362]. Proc Am Assoc Cancer Res 40:2396.

Dhordain P, Lin RJ, Quief S, Lantoine D, Kerckaert JP, Evans RM,Albagli O. 1998. The LAZ3(BCL-6) oncoprotein recruits a SMRT/mSIN3A/histone deacetylase containing complex to mediate tran-scriptional repression. Nucleic Acids Res 26:4645–4651.

Di Fiore B, Guarguaglini G, Palena A, Kerkhoven RM, Bernards R,Lavia P. 1999. Two E2F sites control growth-regulated and cellcycle-regulated transcription of the Htf9-a/RanBP1 gene throughfunctionally distinct mechanisms. J Biol Chem 274:10339–10348.

Di Leonardo A, Linke SP, Clarkin K, Wahl GM. 1994. DNA damagetriggers a prolonged p53-dependent G1 arrest and long-term induc-tion of Cip1 in normal human fibroblasts. Genes Dev 8:2540–2551.

Dobrowolski SF, Stacey DW, Harter ML, Stine JT, Hiebert SW. 1994.An E2F dominant negative mutant blocks E1A induced cell cycleprogression. Oncogene 9:2605–2612.

Doetzlhofer A, Rotheneder H, Lagger G, Koranda M, Kurtev V, BroschG, Wintersberger E, Seiser C. 1999. Histone deacetylase 1 canrepress transcription by binding to Sp1. Mol Cell Biol 19:5504–5511.

Dynlacht BD, Flores O, Lees JA, Harlow E. 1994. Differential regu-lation of E2F transactivation by cyclin/cdk2 complexes. Genes Dev8:1772–1786.

Dyson N. 1998. The regulation of E2F by pRB-family proteins. GenesDev 12:2245–2262.

Emiliani S, Fischle W, Van Lint C, Al-Abed Y, Verdin E. 1998. Char-acterization of a human RPD3 ortholog, HDAC3. Proc Natl Acad SciUSA 95:2795–2800.

Ewen ME, Xing YG, Lawrence JB, Livingston DM. 1991. Molecularcloning, chromosomal mapping, and expression of the cDNA forp107, a retinoblastoma gene product-related protein. Cell 66:1155–1164.

Fenrick R, Hiebert SW. 1998. Role of histone deacetylases in acuteleukemia. J Cell Biochem Suppl 31:194–202.

Ferreira R, Magnaghi-Jaulin L, Robin P, Harel-Bellan A, Trouche D.1998. The three members of the pocket proteins family share theability to repress E2F activity through recruitment of a histonedeacetylase. Proc Natl Acad Sci USA 95:10493–10498.

Fischle W, Emiliani S, Hendzel MJ, Nagase T, Nomura N, Voelter W,Verdin E. 1999. A new family of human histone deacetylases relatedto Saccharomyces cerevisiae HDA1p. J Biol Chem 274:11713–11720.

Futamura M, Monden Y, Okabe T, Fujita-Yoshigaki J, Yokoyama S,Nishimura S. 1995. Trichostatin A inhibits both ras-induced neu-rite outgrowth of PC12 cells and morphological transformation ofNIH3T3 cells. Oncogene 10:1119–1123.

Gamou T, Kitamura E, Hosoda F, Shimizu K, Shinohara K, HayashiY, Nagase T, Yokoyama Y, Ohki M. 1998. The partner gene of AML1in t(16;21) myeloid malignancies is a novel member of theMTG8(ETO) family. Blood 91:4028–4037.

Georgakopoulos T, Thireos G. 1992. Two distinct yeast transcriptionalactivators require the function of the GCN5 protein to promotenormal levels of transcription. EMBO J 11:4145–4152.

12 CRESS AND SETO

Ghosh AK, Steele R, Ray RB. 1999. MBP-1 physically associates withhistone deacetylase for transcriptional repression. Biochem Bio-phys Res Commun 260:405–409.

Ginsberg D, Vairo G, Chittenden T, Xiao ZX, Xu G, Wydner KL,DeCaprio JA, Lawrence JB, Livingston DM. 1994. E2F-4, a newmember of the E2F transcription factor family, interacts with p107.Genes Dev 8:2665–2679.

Glass CK, Holloway JM, Devary OV, Rosenfeld MG. 1988. The thyroidhormone receptor binds with opposite transcriptional effects to acommon sequence motif in thyroid hormone and estrogen responseelements. Cell 54:313–323.

Gobl AE, Berg M, Lopez-Egido JR, Oberg K, Skogseid B, Westin G.1999. Menin represses JunD-activated transcription by a histonedeacetylase-dependent mechanism. Biochim Biophys Acta 1447:51–56.

Grignani F, De Matteis S, Nervi C, Tomassoni L, Gelmetti V, Cioce M,Fanelli M, Ruthardt M, Ferrara FF, Zamir I, Seiser C, Grignani F,Lazar MA, Minucci S, Pelicci PG. 1998. Fusion proteins of theretinoic acid receptor-alpha recruit histone deacetylase in promy-elocytic leukaemia. Nature 391:815–818.

Grozinger CM, Hassig CA, Schreiber SL. 1999. Three proteins definea class of human histone deacetylases related to yeast Hda1p. ProcNatl Acad Sci USA 96:4868–4873.

Gu W, Roeder RG. 1997. Activation of p53 sequence-specific DNAbinding by acetylation of the p53 C-terminal domain. Cell 90:595–606.

Gu W, Shi XL, Roeder RG. 1997. Synergistic activation of transcrip-tion by CBP and p53. Nature 387:819–823.

Guidez F, Ivins S, Zhu J, Soderstrom M, Waxman S, Zelent A. 1998.Reduced retinoic acid-sensitivities of nuclear receptor corepressorbinding to PML- and PLZF-RARalpha underlie molecular patho-genesis and treatment of acute promyelocytic leukemia. Blood 91:2634–2642.

Guo B, Odgren PR, van Wijnen AJ, Last TJ, Nickerson J, Penman S,Lian JB, Stein JL, Stein GS. 1995. The nuclear matrix proteinNMP-1 is the transcription factor YY1. Proc Natl Acad Sci USA92:10526–10530.

Guy CT, Zhou W, Kaufman S, Robinson MO. 1996. E2F-1 blocksterminal differentiation and causes proliferation in transgenicmegakaryocytes. Mol Cell Biol 16:685–693.

Hall M, Peters G. 1996. Genetic alterations of cyclins, cyclin-depen-dent kinases, and Cdk inhibitors in human cancer. Adv Cancer Res68:67–108.

Hampsey M. 1997. A SAGA of histone acetylation and gene expres-sion. Trends Genet 13:427–429.

Hannon GJ, Demetrick D, Beach D. 1993. Isolation of the Rb-relatedp130 through its interaction with CDK2 and cyclins. Genes Dev7:2378–2391.

Hassig CA, Fleischer TC, Billin AN, Schreiber SL Ayer, DE. 1997.Histone deacetylase activity is required for full transcriptional re-pression by mSin3A. Cell 89:341–347.

He LZ, Guidez F, Tribioli C, Peruzzi D, Ruthardt M, Zelent A, PandolfiPP. 1998. Distinct interactions of PML-RARalpha and PLZF-RARalpha with co-repressors determine differential responses toRA in APL. Nat Genet 18:126–135.

Heinzel T, Lavinsky RM, Mullen TM, Soderstrom M, Laherty CD,Torchia J, Yang WM, Brard G, Ngo SD, Davie JR, Seto E, EisenmanRN, Rose DW, Glass CK, Rosenfeld MG. 1997. A complex containingN-CoR, mSin3 and histone deacetylase mediates transcriptionalrepression. Nature 387:43–48.

Helin K, Wu C-L, Lees JA, Dynlacht BD, Ngwu C, Harlow E. 1993.Heterodimerization of the transcription factors E2F-1 and DP-1leads to cooperative. trans-activation. Genes Dev 7:1850–1861.

Helin K, Holm K, Niebuhr A, Eiberg H, Tommerup N, Hougaard S,Poulsen HS, Spang-Thomsen M, Norgaard P. 1997. Loss of theretinoblastoma protein-related p130 protein in small cell lung car-cinoma. Proc Natl Acad Sci USA 94:6933–6938.

Hendrich B, Bird A. 1998. Identification and characterization of afamily of mammalian methyl-CpG binding proteins. Mol Cell Biol18:6538–6547.

Hiebert SW, Chellappan SP, Horowitz JM, Nevins JR. 1992. Theinteraction of RB with E2F coincides with an inhibition of thetranscriptional activity of E2F. Genes Dev 6:177–185.

Hijmans EM, Voorhoeve PM, Beijersbergen RL, van ’t Veer LJ, Ber-nards R. 1995. E2F-5, a new E2F family member that interacts withp130 in vivo. Mol Cell Biol 15:3082–3089.

Horlein AJ, Naar AM, Heinzel T, Torchia J, Gloss B, Kurokawa R,Ryan A, Kamei Y, Soderstrom M, Glass CK, Rosenfeld MG. 1995.Ligand-independent repression by the thyroid hormone receptormediated by a nuclear receptor co-repressor. Nature 377:397–404.

Hsieh JJ, Zhou S, Chen L, Young DB, Hayward SD. 1999. CIR, a

corepressor linking the DNA binding factor CBF1 to the histonedeacetylase complex. Proc Natl Acad Sci USA 96:23–28.

Huang EY, Zhang J, Miska EA, Guenther MG, Kouzarides T, LazarMA. 2000. Nuclear receptor corepressors partner with class II his-tone deacetylases in a Sin3-independent repression pathway. GenesDev 14:45–54.

Huang S, Lee WH, Lee EY. 1991. A cellular protein that competeswith SV40 T antigen for binding to the retinoblastoma gene prod-uct. Nature 350:160–162.

Huang Y, Myers SJ, Dingledine R. 1999. Transcriptional repressionby REST: recruitment of Sin3A and histone deacetylase to neuronalgenes. Nat Neurosci 2:867–872.

Hurford RK Jr, Cobrinik D, Lee MH, Dyson N. 1997. pRB and p107/p130 are required for the regulated expression of different sets ofE2F responsive genes. Genes Dev 11:1447–1463.

Iavarone A, Massague J. 1999. E2F and histone deacetylase mediatetransforming growth factor beta repression of cdc25A during kera-tinocyte cell cycle arrest. Mol Cell Biol 19:916–922.

Inouye CJ, Seto E. 1994. Relief of YY1-induced transcriptional repres-sion by protein-protein interaction with the nucleolar phosphopro-tein B23. J Biol Chem 269:6506–6510.

Ishizaki J, Nevins JR, Sullenger BA. 1996. Inhibition of cell prolifer-ation by an RNA ligand that selectively blocks E2F function. NatMed 2:1386–1389.

Janknecht R, Hunter T. 1996. Transcription: a growing coactivatornetwork [news; comment]. Nature 383:22–23.

Johnson CA, Barlow AL, Turner BM. 1998. Molecular cloning ofDrosophila melanogaster cDNAs that encode a novel histonedeacetylase dHDAC3. Gene 221:127–134.

Johnson DG, Cress WD, Jakoi L, Nevins JR. 1994a. Oncogenic capac-ity of the E2F1 gene. Proc Natl Acad Sci USA 91:12823–12827.

Johnson DG, Ohtani K, Nevins JR. 1994b. Autoregulatory control ofE2F1 expression in response to positive and negative regulators ofcell cycle progression. Genes Dev 8:1514–1525.

Johnson DG, Schneider-Broussard R. 1998. Role of E2F in cell cyclecontrol and cancer. Front Biosci 3:447–448.

Jones PA, Laird PW. 1999. Cancer epigenetics comes of age. NatGenet 21:163–167.

Jones PL, Veenstra GJ, Wade PA, Vermaak D, Kass SU, LandsbergerN, Strouboulis J, Wolffe AP. 1998. Methylated DNA and MeCP2recruit histone deacetylase to repress transcription. Nat Genet 19:187–191.

Jooss K, Lam EW, Bybee A, Girling R, Muller R, La Thangue NB.1995. Proto-oncogenic properties of the DP family of proteins. On-cogene 10:1529–1536.

Kadosh D, Struhl K. 1997. Repression by Ume6 involves recruitmentof a complex containing Sin3 corepressor and Rpd3 histone deacety-lase to target promoters. Cell 89:365–371.

Kaelin WG Jr, Pallas DC, DeCaprio JA, Kaye FJ, Livingston DM.1991. Identification of cellular proteins that can interact specificallywith the T/E1A-binding region of the retinoblastoma gene product.Cell 64:521–532.

Kao HY, Downes M, Ordentlich P, Evans RM. 2000. Isolation of anovel histone deacetylase reveals that class I and class II deacety-lases promote SMRT-mediated repression. Genes Dev 14:55–66.

Kehle J, Beuchle D, Treuheit S, Christen B, Kennison JA, Bienz M,Muller J. 1998. dMi-2, a hunchback-interacting protein that func-tions in polycomb repression. Science 282:1897–1900.

Kijima M, Yoshida M, Sugita K, Horinouchi S, Beppu T. 1993. Trap-oxin, an antitumor cyclic tetrapeptide, is an irreversible inhibitor ofmammalian histone deacetylase. J Biol Chem 268:22429–22435.

Kim GD, Choi YH, Dimtchev A, Jeong SJ, Dritschilo A, Jung M.1999a. Sensing of ionizing radiation-induced DNA damage by ATMthrough interaction with histone deacetylase. J Biol Chem 274:31127–31130.

Kim J, Sif S, Jones B, Jackson A, Koipally J, Heller E, Winandy S, VielA, Sawyer A, Ikeda T, Kingston R, Georgopoulos K. 1999b. IkarosDNA-binding proteins direct formation of chromatin remodelingcomplexes in lymphocytes. Immunity 10:345–355.

Kitabayashi I, Yokoyama A, Shimizu K, Ohki M. 1998. Interactionand functional cooperation of the leukemia-associated factorsAML1 and p300 in myeloid cell differentiation. EMBO J 17:2994–3004.

Koipally J, Renold A, Kim J, Georgopoulos K. 1999. Repression byIkaros and Aiolos is mediated through histone deacetylase com-plexes. EMBO J 18:3090–3100.

Kurokawa R, Soderstrom M, Horlein A, Halachmi S, Brown M, Rosen-feld MG, Glass CK. 1995. Polarity-specific activities of retinoic acidreceptors determined by a co-repressor. Nature 377:451–454.


Ladomery M, Lyons S, Sommerville J. 1997. Xenopus HDm, a mater-nally expressed histone deacetylase, belongs to an ancient family ofacetyl-metabolizing enzymes. Gene 198:275–280.

Laherty CD, Yang WM, Sun JM, Davie JR, Seto E, Eisenman RN.1997. Histone deacetylases associated with the mSin3 corepressormediate mad transcriptional repression. Cell 89:349–356.

Laherty CD, Billin AN, Lavinsky RM, Yochum GS, Bush AC, Sun JM,Mullen TM, Davie JR, Rose DW, Glass CK, Rosenfeld MG, Ayer DE,Eisenman RN. 1998. SAP30, a component of the mSin3 corepressorcomplex involved in N-CoR-mediated repression by specific tran-scription factors. Mol Cell 2:33–42.

Lai A, Lee JM, Yang WM, DeCaprio JA, Kaelin WG Jr, Seto E,Branton PE. 1999. RBP1 recruits both histone deacetylase-depen-dent and -independent repression activities to retinoblastoma fam-ily proteins. Mol Cell Biol 19:6632–6641.

Lam EW, Watson RJ. 1993. An E2F-binding site mediates cell-cycleregulated repression of mouse B-myb transcription. EMBO J 12:2705–2713.

Lam EW, Bennett JD, Watson RJ. 1995. Cell-cycle regulation ofhuman B-myb transcription. Gene 160:277–281.

Lee EY-HP, Chang C-Y, Hu N, Wang Y-CJ, Lai C-C, Herrup K, LeeW-H, Bradley A. 1992. Mice deficient for Rb are nonviable and showdefects in neurogenesis and hematopoieses. Nature 359:288–294.

Lee JS, Galvin KM, See RH, Eckner R, Livingston D, Moran E, Shi Y.1995. Relief of YY1 transcriptional repression by adenovirus E1A ismediated by E1A-associated protein p300. Genes Dev 9:1188–1198.

Lee MH, Williams BO, Mulligan G, Mukai S, Bronson RT, Dyson N,Harlow E, Jacks T. 1996. Targeted disruption of p107: functionaloverlap between p107 and Rb. Genes Dev 10:1621–1632.

Lenny N, Westendorf JJ, Hiebert SW. 1997. Transcriptional regula-tion during myelopoiesis. Mol Biol Rep 24:157–168.

Lewis JD, Meehan RR, Henzel WJ, Maurer-Fogy I, Jeppesen P, KleinF, Bird A. 1992. Purification, sequence, and cellular localization ofa novel chromosomal protein that binds to methylated DNA. Cell69:905–914.

Li Y, Turck CM, Teumer JK, Stavnezer E. 1986. Unique sequence, ski,in Sloan-Kettering avian retroviruses with properties of a newcell-derived oncogene. J Virol 57:1065–1072.

Li Y, Graham C, Lacy S, Duncan AM, Whyte P. 1993. The adenovirusE1A-associated 130-kD protein is encoded by a member of theretinoblastoma gene family and physically interacts with cyclins Aand E. Genes Dev 7:2366–2377.

Lin RJ, Nagy L, Inoue S, Shao W, Miller WH Jr, Evans RM. 1998. Roleof the histone deacetylase complex in acute promyelocytic leukae-mia. Nature 391:811–814.

Luger K, Mader AW, Richmond RK, Sargent DF, Richmond TJ. 1997.Crystal structure of the nucleosome core particle at 2.8 Å resolution.Nature 389:251–260.

Luo RX, Postigo AA, Dean DC. 1998. Rb interacts with histonedeacetylase to repress transcription. Cell 92:463–473.

Lusser A, Brosch G, Loidl A, Haas H, Loidl P. 1997. Identification ofmaize histone deacetylase HD2 as an acidic nucleolar phosphopro-tein. Science 277:88–91.

Lutterbach B, Westendorf JJ, Linggi B, Patten A, Moniwa M, DavieJR, Huynh KD, Bardwell VJ, Lavinsky RM, Rosenfeld MG, Glass C,Seto E, Hiebert SW. 1998. ETO, a target of t(8;21) in acute leuke-mia, interacts with the N-CoR and mSin3 corepressors. Mol CellBiol 18:7176–7184.

Magnaghi-Jaulin L, Groisman R, Naguibneva I, Robin P, Lorain S, LeVillain JP, Troalen F, Trouche D, Harel-Bellan A. 1998a. Retino-blastoma protein represses transcription by recruiting a histonedeacetylase. Nature 391:601–605.

Magnaghi-Jaulin L, Groisman R, Naguibneva I, Robin P, Trouche D,Harel-Bellan A. 1998b. Histone deacetylase and retinoblastomaprotein. Bull Cancer 85:606–607.

Mahlknecht U, Hoelzer D, Bucala R, Verdin E. 1999. Cloning andcharacterization of the murine histone deacetylase (HDAC3). Bio-chem Biophys Res Commun 263:482–490.

Marcus GA, Silverman N, Berger SL, Horiuchi J, Guarente L. 1994.Functional similarity and physical association between GCN5 andADA2: putative transcriptional adaptors. EMBO J 13:4807–4815.

Mayol X, Grana X, Baldi A, Sang N, Hu Q, Giordano A. 1993. Cloningof a new member of the retinoblastoma gene family (pRb2) whichbinds to the E1A transforming domain. Oncogene 8:2561–2566.

McArthur GA, Laherty CD, Queva C, Hurlin PJ, Loo L, James L,Grandori C, Gallant P, Shiio Y, Hokanson WC, Bush AC, Cheng PF,Lawrence QA, Pulverer B, Koskinen PJ, Foley KP, Ayer DE, Eisen-man RN. 1998. The Mad protein family links transcriptional re-pression to cell differentiation. Cold Spring Harb Symp Quant Biol63:423–433.

McBain JA, Eastman A, Nobel CS, Mueller GC. 1997. Apoptotic death

in adenocarcinoma cell lines induced by butyrate and other histonedeacetylase inhibitors. Biochem Pharmacol 53:1357–1368.

McIntyre A, Young GP, Taranto T, Gibson PR, Ward PB. 1991. Dif-ferent fibers have different regional effects on luminal contents ofrat colon. Gastroenterology 101:1274–1281.

McIntyre A, Gibson PR, Young GP. 1993. Butyrate production fromdietary fibre and protection against large bowel cancer in a ratmodel. Gut 34:386–391.

McMahon SB, Van Buskirk HA, Dugan KA, Copeland TD, Cole MD.1998. The novel ATM-related protein TRRAP is an essential cofac-tor for the c-Myc and E2F oncoproteins. Cell 94:363–374.

McMahon SB, Wood MA, Cole MD. 2000. The essential cofactorTRRAP recruits the histone acetyltransferase hGCN5 to c-Myc. MolCell Biol 20:556–562.

Meehan RR, Lewis JD, McKay S, Kleiner EL, Bird AP. 1989. Identi-fication of a mammalian protein that binds specifically to DNAcontaining methylated CpGs. Cell 58:499–507.

Miska EA, Karlsson C, Langley E, Nielsen SJ, Pines J, Kouzarides T.1999. HDAC4 deacetylase associates with and represses the MEF2transcription factor. EMBO J 18:5099–5107.

Murphy M, Ahn J, Walker KK, Hoffman WH, Evans RM, Levine AJ,George DL. 1999. Transcriptional repression by wild-type p53 uti-lizes histone deacetylases, mediated by interaction with mSin3a.Genes Dev 13:2490–2501.

Nan X, Campoy FJ, Bird A. 1997. MeCP2 is a transcriptional repres-sor with abundant binding sites in genomic chromatin. Cell 88:471–481.

Nan X, Ng HH, Johnson CA, Laherty CD, Turner BM, Eisenman RN,Bird A. 1998. Transcriptional repression by the methyl-CpG-bind-ing protein MeCP2 involves a histone deacetylase complex. Nature393:386–389.

Nevins JR. 1994. Cell cycle targets of the DNA tumor viruses. CurrOpin Genet Dev 4:130–134.

Nevins JR. 1998. Toward an understanding of the functional complex-ity of the E2F and retinoblastoma families. Cell Growth Differ9:585–593.

Ng HH, Zhang Y, Hendrich B, Johnson CA, Turner BM, Erdjument-Bromage H, Tempst P, Reinberg D, Bird A. 1999. MBD2 is atranscriptional repressor belonging to the MeCP1 histone deacety-lase complex. Nat Genet 23:58–61.

Nomura T, Khan MM, Kaul SC, Dong HD, Wadhwa R, Colmenares C,Kohno I, Ishii S. 1999. Ski is a component of the histone deacetylasecomplex required for transcriptional repression by Mad and thyroidhormone receptor. Genes Dev 13:412–423.

Nucifora G, Rowley JD. 1995. AML1 and the 8;21 and 3;21 translo-cations in acute and chronic myeloid leukemia. Blood 86:1–14.

Ogryzko VV, Hirai TH, Russanova VR, Barbie DA, Howard BH.1996a. Human fibroblast commitment to a senescence-like state inresponse to histone deacetylase inhibitors is cell cycle dependent.Mol Cell Biol 16:5210–5218.

Ogryzko VV, Schiltz RL, Russanova V, Howard BH, Nakatani Y.1996b. The transcriptional coactivators p300 and CBP are histoneacetyltransferases. Cell 87:953–959.

Parthun MR, Widom J, Gottschling DE. 1996. The major cytoplasmichistone acetyltransferase in yeast: links to chromatin replicationand histone metabolism. Cell 87:85–94.

Phelps WC, Bagchi S, Barnes JA, Raychaudhuri P, Kraus V, MungerK, Howley PM, Nevins JR. 1991. Analysis of trans activation byhuman papillomavirus type 16 E7 and adenovirus 12S E1A sug-gests a common mechanism. J Virol 65:6922–6930.

Pierce AM, Fisher SM, Conti CJ, Johnson DG. 1998a. Deregulatedexpression of E2F1 induces hyperplasia and cooperates with ras inskin tumor development. Oncogene 16:1267–1276.

Pierce AM, Gimenez-Conti IB, Schneider-Broussard R, Martinez LA,Conti CJ, Johnson DG. 1998b. Increased E2F1 activity induces skintumors in mice heterozygous and nullizygous for p53. Proc NatlAcad Sci USA 95:8858–8863.

Qian YW, Lee EY. 1995. Dual retinoblastoma-binding proteins withproperties related to a negative regulator of ras in yeast. J BiolChem 270:25507–25513.

Qian YW, Wang YC, Hollingsworth RE Jr, Jones D, Ling N, Lee EY.1993. A retinoblastoma-binding protein related to a negative regu-lator of Ras in yeast. Nature 364:648–652.

Qin XQ, Chittenden T, Livingston DM, Kaelin WG Jr. 1992. Identifi-cation of a growth suppression domain within the retinoblastomagene product. Genes Dev 6:953–964.

Richon VM, Webb Y, Merger R, Sheppard T, Jursic B, Ngo L, Civoli F,Breslow R, Rifkind RA, Marks PA. 1996. Second generation hybridpolar compounds are potent inducers of transformed cell differen-tiation. Proc Natl Acad Sci USA 93:5705–5708.

14 CRESS AND SETO

Richon VM, Emiliani S, Verdin E, Webb Y, Breslow R, Rifkind RA,Marks PA. 1998. A class of hybrid polar inducers of transformed celldifferentiation inhibits histone deacetylases. Proc Natl Acad SciUSA 95:3003–3007.

Riggs MG, Whittaker RG, Neumann JR, Ingram VM. 1977. n-Bu-tyrate causes histone modification in HeLa and Friend erythroleu-kaemia cells. Nature 268:462–464.

Ross JF, Liu X, Dynlacht BD. 1999. Mechanism of transcriptionalrepression of E2F by the retinoblastoma tumor suppressor protein.Mol Cell 3:195–205.

Rundlett SE, Carmen AA, Kobayashi R, Bavykin S, Turner BM,Grunstein M. 1996. HDA1 and RPD3 are members of distinct yeasthistone deacetylase complexes that regulate silencing and tran-scription. Proc Natl Acad Sci USA 93:14503–14508.

Saito M, Helin K, Valentine MB, Griffith BB, Willman CL, Harlow E,Look AT. 1995. Amplification of the E2F1 transcription factor genein the HEL erythroleukemia cell line. Genomics 25:130–138.

Samid D, Hudgins WR, Shack S, Liu L, Prasanna P, Myers CE. 1997.Phenylacetate and phenylbutyrate as novel, nontoxic differentia-tion inducers. Adv Exp Med Biol 400A:501–505.

Sardet C, Vidal M, Cobrinik D, Geng Y, Onufryk C, Chen A, WeinbergRA. 1995. E2F-4 and E2F-5, two members of the E2F family, areexpressed in the early phases of the cell cycle. Proc Natl Acad SciUSA 92:2403–2407.

Sartorelli V, Puri PL, Hamamori Y, Ogryzko V, Chung G, Nakatani Y,Wang JY, Kedes L. 1999. Acetylation of MyoD directed by PCAF isnecessary for the execution of the muscle program. Mol Cell 4:725–734.

Sasaki S, Lesoon-Wood LA, Dey A, Kuwata T, Weintraub BD, Hum-phrey G, Yang WM, Seto E, Yen PM, Howard BH, Ozato K. 1999.Ligand-induced recruitment of a histone deacetylase in the nega-tive-feedback regulation of the thyrotropin beta gene. EMBO J18:5389–5398.

Scanlan MJ, Chen YT, Williamson B, Gure AO, Stockert E, GordanJD, Tureci O, Sahin U, Pfreundschuh M, Old LJ. 1998. Character-ization of human colon cancer antigens recognized by autologousantibodies. Int J Cancer 76:652–658.

Schreiber-Agus N, Chin L, Chen K, Torres R, Rao G, Guida P, Skoult-chi AI, DePinho RA. 1995. An amino-terminal domain of Mxi1mediates anti-Myc oncogenic activity and interacts with a homologof the yeast transcriptional repressor SIN3. Cell 80:777–786.

Sealy L, Chalkley R. 1978. The effect of sodium butyrate on histonemodification. Cell 14:115–121.

Sears R, Ohtani K, Nevins JR. 1997. Identification of positively andnegatively acting elements regulating expression of the E2F2 genein response to cell growth signals. Mol Cell Biol 17:5227–5235.

Sellers WR, Kaelin WG Jr. 1997. Role of the retinoblastoma protein inthe pathogenesis of human cancer. J Clin Oncol 15:3301–3312.

Sellers WR, Rodgers JW, Kaelin WG Jr. 1995. A potent transrepres-sion domain in the retinoblastoma protein induces a cell cycle arrestwhen bound to E2F sites. Proc Natl Acad Sci USA 92:11544–11548.

Shi Y, Mello C. 1998. A CBP/p300 homolog specifies multiple differ-entiation pathways in Caenorhabditis elegans. Genes Dev 12:943–955.

Siegfried Z, Cedar H. 1997. DNA methylation: a molecular lock. CurrBiol 7:R305–R307.

Siegfried Z, Eden S, Mendelsohn M, Feng X, Tsuberi BZ, Cedar H.1999. DNA methylation represses transcription in vivo. Nat Genet22:203–206.

Singh P, Wong SH, Hong W. 1994. Overexpression of E2F-1 in ratembryo fibroblasts leads to neoplastic transformation. EMBO J13:3329–3338.

Souza RF, Yin J, Smolinski KN, Zou TT, Wang S, Shi YQ, Rhyu MG,Cottrell J, Abraham JM, Biden K, Simms L, Leggett B, Bova GS,Frank T, Powell SM, Sugimura H, Young J, Harpaz N, Shimizu K,Matsubara N, Meltzer SJ. 1997. Frequent mutation of the E2F-4cell cycle gene in primary human gastrointestinal tumors. CancerRes 57:2350–2353.

Sowa Y, Orita T, Minamikawa S, Nakano K, Mizuno T, Nomura H,Sakai T. 1997. Histone deacetylase inhibitor activates the WAF1/Cip1 gene promoter through the Sp1 sites. Biochem Biophys ResCommun 241:142–150.

Sowa Y, Orita T, Minamikawa-Hiranabe S, Mizuno T, Nomura H,Sakai T. 1999. Sp3, but not Sp1, mediates the transcriptional acti-vation of the p21/WAF1/Cip1 gene promoter by histone deacetylaseinhibitor. Cancer Res 59:4266–4270.

Stiegler P, De Luca A, Bagella L, Giordano A. 1998. The COOH-terminal region of pRb2/p130 binds to histone deacetylase 1(HDAC1), enhancing transcriptional repression of the E2F-depen-dent cyclin A promoter. Cancer Res 58:5049–5052.

Stirzaker C, Millar DS, Paul CL, Warnecke PM, Harrison J, Vincent

PC, Frommer M, Clark SJ. 1997. Extensive DNA methylation span-ning the Rb promoter in retinoblastoma tumors. Cancer Res 57:2229–2237.

Struhl K. 1998. Histone acetylation and transcriptional regulatorymechanisms. Genes Dev 12:599–606.

Sun ZW, Hampsey M. 1999. A general requirement for the Sin3-rpd3histone deacetylase complex in regulating silencing in Saccharomy-ces cerevisiae. Genetics 152:921–932.

Takami Y, Kikuchi H, Nakayama T. 1999. Chicken histone deacety-lase-2 controls the amount of the IgM H-chain at the steps of bothtranscription of its gene and alternative processing of its pre-mRNAin the DT40 cell line. J Biol Chem 274:23977–23990.

Targoff IN, Reichlin M. 1985. The association between Mi-2 antibod-ies and dermatomyositis. Arthritis Rheum 28:796–803.

Taunton J, Hassig CA, Schreiber SL. 1996. A mammalian histonedeacetylase related to the yeast transcriptional regulator Rpd3p.Science 272:408–411.

Taya Y. 1997. RB kinases and RB-binding proteins: new points ofview. Trends Biochem Sci 22:14–17.

Thomas MJ, Seto E. 1999. Unlocking the mechanisms of transcriptionfactor YY1: are chromatin modifying enzymes the key? Gene 236:197–208.

Tokitou F, Nomura T, Khan MM, Kaul SC, Wadhwa R, Yasukawa T,Kohno I, Ishii S. 1999. Viral ski inhibits retinoblastoma protein(Rb)-mediated transcriptional repression in a dominant negativefashion. J Biol Chem 274:4485–4488.

Tong JK, Hassig CA, Schnitzler GR, Kingston RE, Schreiber SL. 1998.Chromatin deacetylation by an ATP-dependent nucleosome remod-elling complex. Nature 395:917–921.

Trock B, Lanza E, Greenwald P. 1990a. Dietary fiber, vegetables, andcolon cancer: critical review and meta-analyses of the epidemiologicevidence. J Natl Cancer Inst 82:650–661.

Trock BJ, Lanza E, Greenwald P. 1990b. High fiber diet and coloncancer: a critical review. Prog Clin Biol Res 346:145–157.

van der Vlag J, Otte AP. 1999. Transcriptional repression mediatedby the human polycomb-group protein EED involves histonedeacetylation. Nat Genet 23:474–478.

Vassilev A, Yamauchi J, Kotani T, Prives C, Avantaggiati ML, Qin J,Nakatani Y. 1998. The 400 kDa subunit of the PCAF histone acety-lase complex belongs to the ATM superfamily. Mol Cell 2:869–875.

Verdel A, Khochbin S. 1999. Identification of a new family of highereukaryotic histone deacetylases: coordinate expression of differen-tiation-dependent chromatin modifiers. J Biol Chem 274:2440–2445.

Verreault A, Kaufman PD, Kobayashi R, Stillman B. 1996. Nucleo-some assembly by a complex of CAF-1 and acetylated histonesH3/H4. Cell 87:95–104.

Verreault A, Kaufman PD, Kobayashi R, Stillman B. 1998. Nucleoso-mal DNA regulates the core-histone-binding subunit of the humanHat1 acetyltransferase. Curr Biol 8:96–108.

Vidal M, Gaber RF. 1991. RPD3 encodes a second factor required toachieve maximum positive and negative transcriptional states inSaccharomyces cerevisiae. Mol Cell Biol 11:6317–6327.

Vidal M, Braun P, Chen E, Boeke JD, Harlow E. 1996. Geneticcharacterization of a mammalian protein-protein interaction do-main by using a yeast reverse two-hybrid system. Proc Natl AcadSci USA 93:10321–10326.

Wade PA, Jones PL, Vermaak D, Veenstra GJ, Imhof A, Sera T, TseC, Ge H, Shi YB, Hansen JC, Wolffe AP. 1998a. Histone deacetylasedirects the dominant silencing of transcription in chromatin: asso-ciation with MeCP2 and the Mi-2 chromodomain SWI/SNF ATPase.Cold Spring Harb Symp Quant Biol 63:435–445.

Wade PA, Jones PL, Vermaak D, Wolffe AP. 1998b. A multiple sub-unit Mi-2 histone deacetylase from Xenopus laevis cofractionateswith an associated Snf2 superfamily ATPase. Curr Biol 8:843–846.

Wang AH, Bertos NR, Vezmar M, Pelletier N, Crosato M, Heng HH,Th’ng J, Han J, Yang XJ. 1999. HDAC4, a human histone deacety-lase related to yeast HDA1, is a transcriptional corepressor. MolCell Biol 19:7816–7827.

Wang J, Helin C, Jin P, Nadal-Ginard B. 1995. Inhibition of in vitromyogenic differentiation by cellular transcription factor E2F1. CellGrowth Differ 6:1299–1306.

Weintraub SJ, Prater CA, Dean DC. 1992. Retinoblastoma proteinswitches the E2F site from positive to negative element. Nature358:259–261.

Weintraub SJ, Chow KN, Luo RX, Zhang SH, He S, Dean DC. 1995.Mechanism of active transcriptional repression by the retinoblas-toma protein. Nature 375:812–815.

Whitehead RH, Young GP, Bhathal PS. 1986. Effects of short chainfatty acids on a new human colon carcinoma cell line (LIM1215).Gut 27:1457–1463.


Woodage T, Basrai MA, Baxevanis AD, Hieter P, Collins FS. 1997.Characterization of the CHD family of proteins. Proc Natl Acad SciUSA 94:11472–11477.

Wotton D, Lo RS, Lee S, Massague J. 1999. A Smad transcriptionalcorepressor. Cell 97:29–39.

Wu CL, Classon M, Dyson N, Harlow E. 1996. Expression of domi-nant-negative mutant DP-1 blocks cell cycle progression in G1. MolCell Biol 16:3698–3706.

Xiao H, Hasegawa T, Miyaishi O, Ohkusu K, Isobe K. 1997. Sodiumbutyrate induces NIH3T3 cells to senescence-like state and en-hances promoter activity of p21WAF/CIP1 in p53-independentmanner. Biochem Biophys Res Commun 237:457–460.

Xiao H, Hasegawa T, Isobe K. 1999. Both Sp1 and Sp3 are responsiblefor p21waf1 promoter activity induced by histone deacetylase in-hibitor in NIH3T3 cells. J Cell Biochem 73:291–302.

Xu G, Livingston DM, Krek W. 1995. Multiple members of the E2Ftranscription factor family are the products of oncogenes. Proc NatlAcad Sci USA 92:1357–1361.

Yang WM, Inouye C, Zeng Y, Bearss D, Seto E. 1996a. Transcriptionalrepression by YY1 is mediated by interaction with a mammalianhomolog of the yeast global regulator RPD3. Proc Natl Acad SciUSA 93:12845–12850.

Yang WM, Yao YL, Sun JM, Davie JR, Seto E. 1997. Isolation andcharacterization of cDNAs corresponding to an additional memberof the human histone deacetylase gene family. J Biol Chem 272:28001–28007.

Yang XJ, Ogryzko VV, Nishikawa J, Howard BH, Nakatani Y. 1996b.A p300/CBP-associated factor that competes with the adenoviraloncoprotein E1A. Nature 382:319–324.

Yarden RI, Brody LC. 1999. BRCA1 interacts with components of thehistone deacetylase complex. Proc Natl Acad Sci USA 96:4983–4988.

Yoshida H, Sugita K. 1992. A novel tetracyclic peptide, trapoxin,induces phenotypic change from transformed to normal in sis-onco-gene-transformed NIH3T3 cells. Jpn J Cancer Res 83:324–328.

Yoshida M, Kijima M, Akita M, Beppu T. 1990. Potent and specificinhibition of mammalian histone deacetylase both in vivo and invitro by trichostatin A. J Biol Chem 265:17174–17179.

Yoshitaka T, Matsubara N, Ikeda M, Tanino M, Hanafusa H, Tanaka

N, Shimizu K. 1996. Mutations of E2F-4 trinucleotide repeats incolorectal cancer with microsatellite instability. Biochem BiophysRes Commun 227:553–557.

Zamir I, Harding HP, Atkins GB, Horlein A, Glass CK, Rosenfeld MG,Lazar MA. 1996. A nuclear hormone receptor corepressor mediatestranscriptional silencing by receptors with distinct repression do-mains. Mol Cell Biol 16:5458–5465.

Zhang HS, Postigo AA, Dean DC. 1999a. Active transcriptional re-pression by the Rb-E2F complex mediates G1 arrest triggered byp16INK4a, TGFbeta, and contact inhibition. Cell 97:53–61.

Zhang Y, Iratni R, Erdjument-Bromage H, Tempst P, Reinberg D.1997. Histone deacetylases and SAP18, a novel polypeptide, arecomponents of a human Sin3 complex. Cell 89:357–364.

Zhang W, Bieker JJ. 1998. Acetylation and modulation of erythroidKruppel-like factor (EKLF) activity by interaction with histoneacetyltransferases. Proc Natl Acad Sci USA 95:9855–9860.

Zhang Y, LeRoy G, Seelig HP, Lane WS, Reinberg D. 1998a. Thedermatomyositis-specific autoantigen Mi2 is a component of a com-plex containing histone deacetylase and nucleosome remodelingactivities. Cell 95:279–289.

Zhang Y, Sun ZW, Iratni R, Erdjument-Bromage H, Tempst P, Hamp-sey M, Reinberg D. 1998b. SAP30, a novel protein conserved be-tween human and yeast, is a component of a histone deacetylasecomplex. Mol Cell 1:1021–1031.

Zhang Y, Ng HH, Erdjument-Bromage H, Tempst P, Bird A, ReinbergD. 1999b. Analysis of the NuRD subunits reveals a histone deacety-lase core complex and a connection with DNA methylation. GenesDev 13:1924–1935.

Zhu L, van den Heuvel S, Helin K, Fattaey A, Ewen M, LivingstonDM, Dyson N, Harlow E. 1993. Inhibition of cell proliferation byp107, a relative of the retinoblastoma protein. Genes Dev 7:1111–1125.

Zhu L, Enders G, Lees JA, Beijersbergen RL, Bernards R, Harlow E.1995. The pRB-related protein p107 contains two growth suppres-sion domains: independent interactions with E2F and cyclin/cdkcomplexes. EMBO J 14:1904–1913.

Zwicker J Liu N, Engeland K Lucibello FC, Muller R. 1996. Cell cycleregulation of E2F site occupation in vivo. Science 271:1595–1597.

16 CRESS AND SETO

histone deacetylases transcriptional control, and cancer

Documents

primary nasopharyngeal carcinoma

nucleolar phosphoprotein b23

van der vlag

tetrahymena histone acetyltransferase

constitutive rb phosphorylation

nuclear hormone receptors

histone deacetylase inhibitors

histone deacetylase inhibitor