Nuclear factor one transcription factors as epigenetic regulators in cancer.pdf

Nuclear factor one transcription factors as epigeneticregulators in cancerMitchell Fane1,2,Lachlan Harris1,Aaron G.Smith2,3and Michael Piper11The School of Biomedical Sciences,The University of Queensland,Brisbane,QLD,Australia2School of Biomedical Sciences,Institute of Health and Biomedical Innovation at the Translational Research Institute,Queensland University ofTechnology,Woolloongabba,QLD,Australia3Dermatology Research Centre,The University of Queensland,School of Medicine,Translational Research Institute,Brisbane,QLD,AustraliaTumour heterogeneity poses a distinct obstacle to therapeutic intervention.While the initiation of tumours across various physiologicalsystems is frequently associated with signature mutations in genes that drive proliferation and bypass senescence,increasing evidencesuggests that tumour progression and clonal diversity is driven at an epigenetic level.The tumour microenvironment plays a key role indriving diversity as cells adapt to demands imposed during tumour growth,and is thought to drive certain subpopulations back to astem cell-like state.This stem cell-like phenotype primes tumour cells to react to external cues via the use of developmental pathwaysthat facilitate changes in proliferation,migration and invasion.Because the dynamism of this stem cell-like state requires constantchromatin remodelling and rapid alterations at regulatory elements,it is of great therapeutic interest to identify the cell-intrinsic factorsthat confer these epigenetic changes that drive tumour progression.The nuclear factor one(NFI)family are transcription factors thatplay an important role in the development of many mammalian organ systems.While all four family members have been shown to actas either oncogenes or tumour suppressors across various cancer models,evidence has emerged implicating them as key epigeneticregulators during development and within tumours.Notably,NFIs have also been shown to regulate chromatin accessibility at distalregulatory elements that drive tumour cell dissemination and metastasis.Here we summarize the role of the NFIs in cancer,focusinglargely on the potential mechanisms associated with chromatin remodelling and epigenetic modulation of gene expression.IntroductionEpigenetics has historically been defined as heritable cellularphenotypes of organisms that are independent of alterationsin DNA sequence,however,as the term has continued toevolve,it is now used more to describe changes in cellularand molecular phenotypes that are associated with alterationsto chromatin structure and accessibility.1,2A major substrateof epigenetic change is chromatin,the macromolecular com-plex made up of DNA and histone proteins.Chromatin canbe modified by four distinct mechanisms;these are DNAmethylation,histone modification,nucleosome remodellingand RNA-mediated targeting.While epigenetic processingwas first described as an important process in development,3its misregulation is now thought to also be a key componentin driving cancer formation and tumour progression.4,5Forexample,factors associated with tumour progression such asaging,6chronic inflammation7and environmental exposuresuch as to cigarette smoke8all alter the epigenome of cells.One key aspect associated with cancer progression istumour heterogeneity.9This heterogeneity is not only definedby the various cell types often found within tumours(fibro-blasts,endothelial cells,pericytes and immune cells),but also atthe genetic and phenotypic level within individual cancercells.10Despite many tumours originating via irreversible muta-tions in oncogenes and tumour suppressors derived from a sin-gle cell,many cancers display large levels of clonal diversity.11While the mechanisms behind tumour heterogeneity at thegenetic and phenotypic level are continually being defined,there is a clear epigenetic component that drives this process.12The highly proliferative nature of many malignancies makesthe tumour microenvironment very dynamic during growthand,as such,cells must continuously adapt to environmentalAbbreviations:ADAM12:disintegrin and metalloprotease domain-containing protein 12;ATAC-seq:assay for transposase-accessiblechromatin with high throughput sequencing;ChIP-seq:chromatinimmunoprecipitation sequencing;DNMT:DNA methyltransferase;ECM:extracellular matrix;EZH2:enhancer of zeste homolog 2;GR:glucocorticoidreceptor;HDAC:histonedeacetylase;LTR:long terminal repeats;MeCP2:methyl CpG binding protein 2;miRNA:micro RNA;MMTV:mouse mammary tumour virus;NFI:nuclear factor one;NFIA:nuclear factor one A;NFIB:nuclear factor one B;NFIC:nuclear factor one C;NFIX:nuclearfactor one X;OCT1a:octamer factor 1;RNA-seq:RNA sequencing;SCLC:small cell lung cancer;SV40:simian virus 40Grant sponsor:Cancer Council Queensland Grant(MP)and anAustralian Research Council;Grant number:DP160100368 to MP;Grant sponsor:Australian Research Council Future Fellowship;Grant number:FT120100170;Grant sponsor:MF and LH weresupported by Australian Postgraduate Awards.DOI:10.1002/ijc.30603History:Received 27 Oct 2016;Accepted 29 Dec 2016;Online 11Jan 2017Correspondence to:Michael Piper,The School of BiomedicalSciences,The University of Queensland,Brisbane,4072,Australia,E-mail:m.piperuq.edu.au;Tel:161 733654484Mini ReviewInt.J.Cancer:140,26342641(2017)VC2017 UICCInternational Journal of CancerIJCstressors.Conditions such as nutrient deprivation,hypoxia,immune responses and even treatments aimed at destroyingtumours have been shown to induce considerable changes incancer cell DNA methylation status and chromatin remodel-ling.13,14Cross talk between cellular populations(both cancerand other stromal populations),either through direct interaction,or via secretory factors,can also induce changes in cell motilityand proliferation underpinned by epigenetic changes.An inter-esting observation commonly associated with cellular invasion isthe idea that many cancer cells undergo a process of de-differentiation to attain a stem cell-like phenotype.15,16Such acellstate isextremely dynamic and primed to respond to externalcues,allowing it to quickly alter its proliferative and migratorycharacteristics throughout various developmental stages.12,17This dynamism relies heavily on chromatin remodelling andchanges to histone structure and function,through the combina-torial effect of a myriad of histone modifications that can be rap-idly altered to facilitate immediate and reversible changes togene expression.18Micro RNAs(miRNAs)are another key epigenetic regulatorthat potentiate tumour progression.They can act as eitheroncogenes or tumour suppressors and,as such,can be regulat-ed in a manner whereby their expression is increased ordecreased via transcriptional regulation.There are variousmechanisms contributing to miRNA deregulation in cancercells.19Interestingly,while the regulation of cancer cell pheno-types via miRNAs is often an intrinsic cellular process,thereare various mechanisms by which miRNAs can be secreted inthe microenvironment to induce autocrine and paracrineeffects on other tumour cell populations.19They can also play adistal role in regulating tumour angiogenesis,immunopheno-type and extracellular matrix(ECM)remodelling.19,20Deregu-lation of miRNAs within stromal cell populations has also beenshown to play a large role in promoting initial tumour forma-tion and progression,20while other microenvironmental factorssuch as hypoxia,nutrient deprivation and increased acidity alsodrive aberrant miRNA expression in cancer cells.21,22Tumour heterogeneity has a clear therapeutic impact.2325The clonal diversity observed in many developed tumours isvast and frequently small clonal subsets will confer selectiveadvantages that drive tumour progression or remain refracto-ry to therapies that are effective against the majority of cellsin the tumour.It is clear that modulation of the genome atan epigenetic level plays a major role in heterogeneity and istherefore of great therapeutic interest to identify key regula-tors that confer these epigenetic changes throughout tumourprogression.This review will focus one such family of regulators,theNFIs,and their role in tumour progression.First described asa host protein required for adenovirus replication(Nagataet al.1982),there have been four NFI transcription factorsidentified in vertebrates,NFIA,NFIB,NFIC and NFIX.26,27NFIs are developmentally important proteins that in broadterms drive progenitor cell differentiation within the centralnervous system,2830lung31,32and muscle.33,34After earlyembryonic and postnatal development,NFIs continue to haveimportant roles in regulating progenitor cell biology.In adultprogenitors cells,within neural tissue,the skin and skeletalmuscle,NFIs promote differentiation35,36but also regulatethe balance between cell-cycle entry and exit,3638and there-by regulate the homeostasis that exists between progenitorpool expansion and tissue regeneration.NFIs have also beenestablished as key tumour suppressors and oncogenes acrossa range of cancers,with recent evidence suggesting thatthey may function as key drivers of tumour progression bycoordinating changes in the epigenome.39NFIs As Epigenetic RegulatorsTranscription factors are traditionally defined by their abilityto coordinate gene expression by binding directly to regulato-ry regions within DNA alone,or via protein interactions withtranscription co-factors that bind DNA indirectly throughinteraction with transcription factors at cis-regulatory sites.NFIs for example,can both activate or repress gene expres-sion by binding to the DNA dyad symmetric consensus siteor half-site sequence TTGGC(N5)GCCAA on double strand-ed DNA,4042forming homodimers or heterodimers.Whilethis is a key feature of transcription factors,evidence revealsthat many families are able to alter gene expression by mod-ulating chromatin structure directly through nucleosomeremodelling,or indirectly via interactions with,or the regula-tion of,epigenetic modifiers.We argue that this may also betrue of NFI family members,consistent with a number ofstudies demonstrating that NFIs interact with chromatin andchromatin regulators in a variety of ways.Moreover,recentchromatin immunoprecipitation sequencing(ChIP-seq)andRNA sequencing(RNA-seq)experiments suggest that the neteffect of these NFI-chromatin interactions may be to increasechromatin accessibility and gene expression.39,43NFI activity generally correlates with increased expressionof their target genes and is associated with higher levels ofactive promoter methylation marks such as H3K4me3 andH3K36me3,implicating them predominantly as activators oftranscription.44The first evidence revealing that the NFI fam-ily can act in a transcriptionally independent manner foundthat they were able to bind to GCCAAT recognition sitesand serve as initiation factors during DNA replication.45Moreover,early structural analysis of NFIs identified theirpotential for chromatin regulation as they contained a trans-activation domain that interacted with histones H1 andH3.46,47Further functional analysis demonstrated that NFIswere able to alter the interaction of reconstituted nucleoso-mal cores with DNA in vitro in a growth factor-dependentmanner.47In vivo evidence also revealed that NFIs are ableto alter native chromatin structure at yeast origins of replica-tion48and other promoter regions through direct interactionwith histone proteins.49NFIs were also shown to activatesimian virus 40(SV40)DNA replication in vivo by interact-ing directly with histone H3 and relieving nucleosomalrepression at the SV40 origin.50Indeed,there are many otherMini ReviewFane et al.2635Int.J.Cancer:140,26342641(2017)VC2017 UICCexamples of NFI directly interacting with the nucleosomalarchitecture.Studies in yeast cells for example,show that NFIwas able to delimit chromatin domain boundaries by bindinghistone H3,thereby preventing epigenetic teleomeric silenc-ing by forming a partition between the gene and telomereand blocking silent information regulator(SIR)proteinsinvolved in the de-acetylation of histone tails.51More recent-ly,genome wide mapping analysis of NFI DNA binding siteswithin mouse embryonic fibroblasts confirms that there is aclear association between NFI and histone H3.44Consistentwith this,NFI globally associates with chromatin domainboundaries,separatingpermissiveandsilentchromatinmarkers as defined by localization with H3K27me3 andH3K36me3 boundaries of opposite polarities,and they werefurtherfoundtodirectlyinteractwithpositionednucleosomes.43Crucially,NFIs have also been shown to interact withchromatin in human cells.NFIs are able to prevent thesilencing of transgenes upon chromosomal integration in ahistone dependent manner,however,they were unable toactivate transcription alone in HEK293 cells.52Studies per-formed in HeLa cells investigating mechanisms underlyingthe boundaries between euchromatic(active,permissive chro-matin)and heterochromatic(silent,condensed chromatin)domains confirmed that NFI proteins,or fusions containingthe histone binding domain of NFIs,can partition two genescolocalized at a telomeric locus into active and inactive chro-matin structures.53These findings suggest a mechanismwhereby NFI interacts directly with nucleosomes in a tran-scriptionally independent manner to establish a chromosomalstructure that blocks silencing signals emanating from thetelomere,while maintaining a permissive chromatin state toallow for increased gene expression within these regions.Italso provides further mechanistic evidence to explain the pre-vious findings that NFIs can induce a permissive chromatinstate to reverse chromatin-mediated gene silencing withoutbeing able to activate transcription of these genes alone.52In depth analysis of NFI interactions with chromatin andtheir role in remodelling have also been performed usinglong terminal repeat(LTR)regulatory sequences of themouse mammary tumour virus(MMTV)(for an in depthreview on this,see ref.54).NFI-binding sites were found onB nucleosomes in the MMTV promoter in the 1980s,withresearchshowingthatthereisafunctionalsynergismbetween NFI and the glucocorticoid receptor(GR)in bindingto a chromatin template and activating the promoter,whichwas abrogated in a nucleosome depleted environment.55Fol-low up studies in this model system using linker-scanningmutants of transcription factor binding sites found that NFIswere required for hormone-dependent chromatin remodel-ling,as they found that binding site mutations substantiallydecreased hormone-mediated remodelling of nucleosome Band that NFI was also necessary for the association betweenthe BRG1 chromatin remodelling complex and GR on thepromoter in vivo.56Studies into S.cerevisiae looking atminichromosomes assembled on MMTV LTR sequences areconsistent with a role for NFIs in chromatin remodelling,asthey act as classical transcription factors in a relaxed chroma-tin context,whereas in a wild type chromatin confirmation,NFI cooperates with steroid hormone receptors to stabilize