Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2011 May 15;474(7351):390-4.
doi: 10.1038/nature10006.

Reprogramming transcription by distinct classes of enhancers functionally defined by eRNA

Dong Wang  1 , Ivan Garcia-Bassets, Chris Benner, Wenbo Li, Xue Su, Yiming Zhou, Jinsong Qiu, Wen Liu, Minna U Kaikkonen, Kenneth A Ohgi, Christopher K Glass, Michael G Rosenfeld, Xiang-Dong Fu
Affiliations

Reprogramming transcription by distinct classes of enhancers functionally defined by eRNA

Dong Wang et al. Nature. .

Abstract

Mammalian genomes are populated with thousands of transcriptional enhancers that orchestrate cell-type-specific gene expression programs, but how those enhancers are exploited to institute alternative, signal-dependent transcriptional responses remains poorly understood. Here we present evidence that cell-lineage-specific factors, such as FoxA1, can simultaneously facilitate and restrict key regulated transcription factors, exemplified by the androgen receptor (AR), to act on structurally and functionally distinct classes of enhancer. Consequently, FoxA1 downregulation, an unfavourable prognostic sign in certain advanced prostate tumours, triggers dramatic reprogramming of the hormonal response by causing a massive switch in AR binding to a distinct cohort of pre-established enhancers. These enhancers are functional, as evidenced by the production of enhancer-templated non-coding RNA (eRNA) based on global nuclear run-on sequencing (GRO-seq) analysis, with a unique class apparently requiring no nucleosome remodelling to induce specific enhancer-promoter looping and gene activation. GRO-seq data also suggest that liganded AR induces both transcription initiation and elongation. Together, these findings reveal a large repository of active enhancers that can be dynamically tuned to elicit alternative gene expression programs, which may underlie many sequential gene expression events in development, cell differentiation and disease progression.

PubMed Disclaimer

Figures

Figure 1
Figure 1. FoxA1 contributes to the enhancer code in prostate cancer cells
(a) Distribution of histone marks within ±2Kb windows around distinct genomic regions (n=43,565) marked by H3K4me1, but not H3K4me3, in androgen (DHT)-stimulated LNCaP cells. The ChIP-seq datasets for H3K4me1, H3K4me2, H3K4me3, H3K27ac, H4K5ac and p300 were each aligned with respect to the center of theH3K4me1 signal and sorted by the length of H3K4me1-marked regions. (b) Top-enriched DNA motifs with significant P-value and prospective families of DNA binding transcription factors identified by de novo motif analysis of non-promoter regions marked by H3K4me1. (c) Percentage of H3K4me1-marked regions that show FoxA1 binding events (top panel) and percentage of FoxA1 binding sites that are marked by H3K4me1 (bottom panel). Note that H3K4me1-marked regions tend to be broad, but FoxA1 binding sites are discrete, and as a result, many H3K4me1-positive regions may contain more than one FoxA1 binding site. (d) Genomic distance from FoxA1/H3K4me1-positive loci to the nearest TSS of genes in response to FOXA1 knockdown. Outliers were omitted from box plots. P-values indicate the significance in pair-wise comparisons. (e-g) Three classes of FoxA1/H3K4me1-positive loci according to the response in levels of H3K4me1 to FOXA1 knockdown: >1.5-fold decrease (e), no significant change (f), and >1.5-fold increase (g). (h) Ratio (log2) of up- and down-regulated genes in each H3K4me1 responsive category in e-g.
Figure 2
Figure 2. AR reprogramming and induced alternative hormonal response
(a)FOXA1 siRNA-induced cell progression to S phase. Relative numbers of propidium iodide (PI)-labeled cells in S-phase at different DH) concentrations were determined by FASCan. The P-value for the difference detected at each hormonal level is indicated; mean±s.e.m. is based on three independent experiments. (b) Top-enriched motifs associated with AR-occupied loci (n=3,115). (c) Comparison between genome-wide AR binding programs before and after FOXA1 knockdown in DHT-treated LNCaP cells. (d) Quantitative levels of AR binding in the “lost”, “conserved”, and “gained” programs. Outliers were omitted from box plots. (e) ChIP-qPCR validated AR binding events on randomly selected loci from the lost (n=22), conserved (n=27), and gained (n=16) programs. (f) Microarray analysis of DHT-induced genes before and after FOXA1 knockdown. (g) Quantitative analysis of gained androgen up-regulated genes based on microarray analysis in f. Outliers were omitted from box plots. (h) Genomic distance of androgen-responsive genes from TSS to the nearest AR binding site in the original and gained AR binding programs.
Figure 3
Figure 3. Transcriptional response on individual enhancer programs to FOXA1 down-regulation
(a) Display of nascent RNA detected by GRO-seq on the KLK3 locus. The DHT-induced AR binding is shown at bottom as a reference. (b,c) Induction of eRNA by DHT (b) or FOXA1 knockdown in DHT-treated LNCaP cells (c). The eRNA levels under different conditions (indicated at bottom) are separately displayed on three AR binding programs. (d,e) Effects of FoxA1 on binding of p300 (d) and Med12 (e) in each AR program in DHT-treated LNCaP cells. (f, g) Long-distance interaction between gene promoter and AR bound site (at ~50Kb) was determined by the 3C assay on two representative gene loci selected from the conserved and gained AR programs. Negative controls at shorter distances (~30 and ~40Kb) and a positive control with the corresponding BAC in the region are included in each case.
Figure 4
Figure 4. Distinct classes of AR enhancers in the human genome
(a,b) Profiles of H3K4me1 (a) and H3K27ac (b) associated with the lost (bottom panels), conserved (top panels), and gained (middle panels) AR programs in DHT-treated LNCaP cells in response to FOXA1 knockdown. (c, d) Profiles of H3K4me2 around AR binding loci at the nucleosomal resolution in response to DHT stimulation in control siRNA-treated (c) or FOXA1 siRNA-treated (d) LNCaP cells. (e) Profiles of the histone variant H2A.Z on the three different AR programs. (f) Model for FoxA1-mediated AR targeting and reprogramming in LNCaP cells. In Class I (the lost AR program), FoxA1 licenses liganded AR to bind to ARE in relatively nucleosome-free regions. AR binding does not induce nucleosome remodeling in this class of enhancers. In Class II (the conserved AR program), AR binds independently of FoxA1 to ARE, inducing nucleosome remodeling. In Class III (the gained AR program), FoxA1 restricts AR binding, despite the presence of strong AREs. Although pre-established, these gained loci exhibit a strong central nucleosome and are associated with H2AZ, which is not affected by AR binding. FOXA1 knockdown converted these sites to androgen-responsive sites. In all these three classes, eRNAs were generated or increased after AR binding.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Hu Q, et al. Enhancing nuclear receptor-induced transcription requires nuclear motor and LSD1-dependent gene networking in interchromatin granules. Proc Natl Acad Sci U S A. 2008;105:19199–19204. - PMC - PubMed
    1. Holbro T, et al. The ErbB2/ErbB3 heterodimer functions as an oncogenic unit: ErbB2 requires ErbB3 to derive breast tumor cell proliferation. Proc Natl Acad Sci U S A. 2003;100:8933–8938. - PMC - PubMed
    1. Heinz S, et al. Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. Mol Cell. 2010;38:576–589. - PMC - PubMed
    1. Barski A, et al. High-resolution profiling of histone methylations in the human genome. Cell. 2007;129:823–837. - PubMed
    1. Linhart C, Halperin Y, Shamir R. Transcription factor and microRNA motif discovery: the Amadeus platform and a compendium of metazoan target sets. Genome Res. 2008;18:1180–1189. - PMC - PubMed

Publication types

MeSH terms

Associated data