Due to advances in genomic technologies, our understanding of estrogen receptor (ER)-mediated transcription in breast cancer cells has evolved significantly in recent years. Genome-wide mapping experiments revealed thousands of ER-binding events, but linking them to the target genes has been an ongoing struggle. A recent paper describes a new technique, called ChIA-PET (chromatin interaction analysis using paired-end tag sequencing), that can directly address these questions. ChIA-PET is an unbiased approach for simultaneously identifying all genome-wide binding events of a transcription factor and those involved in long-range chromatin loops

Carroll, Jason S

Theodorou, Vasiliki

English

PubMed

Apollo (Cambridge)

BackgroundEstrogen receptor (ER) has been one of the most intensively studied transcription factors, primarily due to its importance in breast cancer. Much eff ort has been invested in identifying the target genes of ER, mainly by expression arrays in breast cancer cell lines and in human tumour samples [1,2]. In the past few years, the evolution of genomic technologies has permitted mapping of ER-binding events in an unbiased manner [3,4]. Th is involves combining chromatin immunoprecipitation with micro arrays or high-throughput sequencing in order to identify ER contact points within the genome. Th ese genome-wide experiments revealed a number of surprising features about ER biology, the most unexpected one being that ER regulates transcription mostly from distal enhancers. Th is fi nding raised the subsequent question: which of the thousands of ER-binding events are the functional ones that regulate expression of the few hundred ER target genes? Fullwood and colleagues [5] have now established a method for addressing this question, allowing researchers to assign ER-binding events to transcriptional hubs and target genes.The articleResearchers at the Genome Institute of Singapore have developed a technique called ChIA-PET (chromatin interaction analysis using paired-end tag sequencing) [5], which couples chromosome conformation capture [6], a method for identifying interacting chromatin regions, with high-throughput sequencing. Th is permits the global identifi cation of chromatin loops formed at a specifi c time point. Th e authors used a well-established system whereby MCF-7 breast cancer cells are cultured in hormone-deprived media and subsequently treated with estrogen for a short time period, resulting in synchronisation of transcriptional events [7]. Using this system, the authors could capture the high-confi dence chromatin interactions that form during the fi rst wave of estrogen-ER-mediated transcription. Th ese interactions were extensively validated, supporting their conclusions. Th e implications of this investigation are profound. It is revealed that chromatin interactions can occur in clusters of contact points and that the actual anchor points, in a linear two-dimensional represen ta tion, can be found separ ated by hundreds of kilobases and in some cases megabases. Contrary to previous sugges tions [8], inter-chromosomal interactions do not appear to be a common or experimentally verifi able pheno menon. Th is suggests that ER-binding events regulate genes within mostly a spatially confi ned environ ment, which in many cases involves the folding of multiple regulatory regions into one transcriptional hub.In summary, the authors fi nd 689 ER-associated chromatin interaction complexes made up of duplexes and more complex interactions. Th ese tend to involve the stronger ER-binding events, which are biased toward specifi c histone marks and other transcriptional regula-tors important for ER function, such as FoxA1 [9]. By combining the ChIA-PET data with gene expression data, they could discover individual transcriptional hubs and the target genes localised in the vicinity.The viewpointUntil now, researchers have been linking enhancers and cis-regulatory elements with the target genes one at a time [10,11]. ChIA-PET circumvents this limitation and AbstractDue to advances in genomic technologies, our understanding of estrogen receptor (ER)-mediated transcription in breast cancer cells has evolved signifi cantly in recent years. Genome-wide mapping experiments revealed thousands of ER-binding events, but linking them to the target genes has been an ongoing struggle. A recent paper describes a new technique, called ChIA-PET (chromatin interaction analysis using paired-end tag sequencing), that can directly address these questions. ChIA-PET is an unbiased approach for simultaneously identifying all genome-wide binding events of a transcription factor and those involved in long-range chromatin loops.© 2010 BioMed Central LtdEstrogen receptor action in three dimensions - looping the loopVasiliki Theodorou and Jason S CarrollV I E W P O I N T*Correspondence: Jason.carroll@cancer.org.ukCancer Research UK, Cambridge Research Institute, Robinson Way, Cambridge, CB2 0RE, UKTheodorou and Carroll Breast Cancer Research 2010, 12:303http://breast-cancer-research.com/content/12/1/303© 2010 BioMed Central Ltdcan identify all active loops at a given time point, allowing us to redefi ne the functional ER-binding events. While this constitutes the fi rst snapshot of ER-mediated chroma tin loops, it is a quantum leap in our under-standing of transcription dynamics. However, this pioneer ing work leads to a series of new questions. Th e authors associate the DNA loops that they captured at 45  minutes of estrogen treatment with a time course expression profi le of cells treated with estrogen from 0 to 48 hours of treatment. A challenging concept that needs further testing is whether the chromatin loops (captured at 45  minutes) are stable over time or whether they change, leading to the formation of new loops at later time points to regulate transcription of the ‘late’ target genes. Moreover, what is the role of co-operating transcription factors in loop formation? Also, since many of the ER functional binding events do not directly involve promoter regions, one can speculate that some DNA loops might circum vent the need for a transcription factor to be present at the promoter by concentrating the transcription machinery in the three-dimensional hubs.Furthermore, this technology can be used to explore ER chromatin loops and transcriptional hubs in diff erent breast cancer cell lines and treatment conditions. Th e identifi cation of transcriptionally active binding events in the diff erent contexts will illuminate the factors that dictate diff erential binding and transcriptional activity. However, the major limitations of the ChIA-PET tech-nology are the complex technical aspects of the proce-dure, the heavy bioinformatics demand and the require-ment for high-throughput sequencers. Neverthe less, these genomic tools are becoming more readily available, and as such, ChIA-PET will prove to be a powerful tool for delineating mechanisms of transcription in a spatial and temporal manner, applicable to any transcription factor of interest.AbbreviationsChIA-PET = chromatin interaction analysis using paired-end tag sequencing; ER = estrogen receptor.Competing interestsThe authors declare that they have no competing interests.Published: 8 February 2010References1. Perou CM, Sørlie T, Eisen MB, van de Rijn M, Jeff rey SS, Rees CA, Pollack JR, Ross DT, Johnsen H, Akslen LA, Fluge O, Pergamenschikov A, Williams C, Zhu SX, Lønning PE, Børresen-Dale AL, Brown PO, Botstein D: Molecular portraits of human breast tumours. Nature 2000, 406:747-752.2. 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Lupien M, Eeckhoute J, Meyer CA, Wang Q, Zhang Y, Li W, Carroll JS, Liu XS, Brown M: FoxA1 translates epigenetic signatures into enhancer-driven lineage-specifi c transcription. Cell 2008, 132:958-970.10. Barnett DH, Sheng S, Charn TH, Waheed A, Sly WS, Lin CY, Liu ET, Katzenellenbogen BS: Estrogen receptor regulation of carbonic anhydrase XII through a distal enhancer in breast cancer. Cancer Res 2008, 68:3505-3515.11. Pan YF, Wansa KD, Liu MH, Zhao B, Hong SZ, Tan PY, Lim KS, Bourque G, Liu ET, Cheung E: Regulation of estrogen receptor-mediated long range transcription via evolutionarily conserved distal response elements. J Biol Chem 2008, 283:32977-32988.Theodorou and Carroll Breast Cancer Research 2010, 12:303http://breast-cancer-research.com/content/12/1/303doi:10.1186/bcr2470Cite this article as: Theodorou V, Carroll JS: Estrogen receptor action in three dimensions - looping the loop. Breast Cancer Research 2010, 12:303.Page 2 of 2

Estrogen receptor action in three dimensions - looping the loop.

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FoxA1 translates epigenetic signatures into enhancer-driven lineage-specifi c transcription. Cell

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Genome-wide analysis of estrogen receptor binding sites. Nat Gen

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Estrogen receptor action in three dimensions - looping the loop.

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