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. 2011 Jun;18(6):742-6.
doi: 10.1038/nsmb.2060. Epub 2011 May 29.

The nucleosome map of the mammalian liver

Zhaoyu Li  1 , Jonathan Schug, Geetu Tuteja, Peter White, Klaus H Kaestner
Affiliations

The nucleosome map of the mammalian liver

Zhaoyu Li et al. Nat Struct Mol Biol. 2011 Jun.

Abstract

Binding to nucleosomal DNA is critical for 'pioneer' transcription factors such as the winged-helix transcription factors Foxa1 and Foxa2 to regulate chromatin structure and gene activation. Here we report the genome-wide map of nucleosome positions in the mouse liver, with emphasis on transcriptional start sites, CpG islands, Foxa2 binding sites and their correlation with gene expression. Despite the heterogeneity of liver tissue, we could clearly discern the nucleosome pattern of the predominant liver cell, the hepatocyte. By analyzing nucleosome occupancy and the distributions of heterochromatin protein 1 (Hp1), CBP (also known as Crebbp) and p300 (Ep300) in Foxa1- and Foxa2-deficient livers, we find that the maintenance of nucleosome position and chromatin structure surrounding Foxa2 binding sites is independent of Foxa1 and Foxa2.

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Conflict of interest statement

COMPETING INTERESTS STATEMENT

The authors declare no competing financial interests.

Figures

Figure 1
Figure 1
Mapping of nucleosome positions by Micrococcal nuclease is more informative than the use of H3K4me1 ChIP-Seq data. (a) Blue, nucleosome positions mapped by Micrococcal nuclease digestion followed by ultra-high throughput sequencing; orange, H3K4me1 ChIP-Seq data from mouse liver based on a cutoff of more than six reads per position. Nucleosome positions estimated from H3K4me1 ChIP-Seq data are unable to resolve closely spaced nucleosomes and also miss multiple nucleosomes detected by mapping of nucleosomes with micrococcal nuclease. Near the 5′ end of the Alb1 gene, H3K4me1 ChIP-Seq data miss two important nucleosomes just downstream of the transcriptional start site, most likely because here H3 was methylated further to the me2 and me3 forms. (b) The well-known Foxa binding site in the albumin enhancer was called as “bi-modal”, i.e. nucleosome-free, by Hoffman and colleagues, however, our data clearly show the site occupied by a nucleosome.
Figure 2
Figure 2
Nucleosome dynamics in the mouse liver. (a–c) Nucleosome map of the mouse liver. Stack height profiles of nucleosome reads represent nucleosome occupancy in the gene body (a), enhancer (b) and surrounding the transcriptional start site (TSS) (c) of the Albumin gene. Blue ovals represent putative nucleosome positions. (d) Nucleosome distribution surrounding transcriptional start sites (‘All’, all annotated genes from the UCSC genome database; ‘Active’, 1,000 genes with the highest expression in the liver; ‘Silent’, 1,000 genes with no expression in the liver. (e) Nucleosome distribution surrounding TSS of genes associated with (CpG+) and without (CpG−) CpG islands. (f) Nucleosome distribution surrounding CpG islands. Either all CpG islands or those located more than 5 Kb distant to the nearest gene (Intergenic) were analyzed. (g) Sizes of nucleosome-free regions at Foxa2 binding sites, surrounding transcriptional start sites, or in the whole genome (All).
Figure 3
Figure 3
Genome-wide distributions of Foxa2 and nucleosomes in the adult mouse liver. (a) Examples of Foxa2 occupancy at a nucleosome-bound (top) or nucleosome-free (bottom) binding site. The 10-bp sequence with the best match to the Foxa2 positional weight matrix is denoted as “Foxa2 binding site”. (b) Nucleosome occupancy surrounding Foxa2 binding sites. 0 indicates the center of 10-bp sequence with the best match to the Foxa2 positional weight matrix in each Foxa2 bound region. Note the relative nucleosome depletion at the center of Foxa2 bound sites. (c) Consensus binding sites of nucleosome-free and nucleosome-occupied Foxa2 do not differ significantly. (d) Direct interactions between Foxa2 and histones. Sequential ChIPs (sChIP) were performed with MNase-digested chromatin and analyzed by qPCR with 18 primers pairs. Nine of these corresponded to Foxa2 binding sites in nucleosome-free genomic regions (left side of graph), while nine corresponded to nucleosome-bound Foxa2 binding sites (right side of graph). The order of sChIP: sChIP-1 (anti-Foxa2 → anti-Histone H3) and sChIP-2 (anti-Histone H3 → anti-Foxa2). Regardless of ChIP order, all nucleosome-bound Foxa2 targets were confirmed as such by sChIP. Insert is the western blot using sChIP samples with anti-Foxa2 and anti-Histone H3 antibodies. (e) Distributions of Foxa2 binding sites at upstream and downstream of transcriptional start sites.
Figure 4
Figure 4
The maintenance of nucleosome position and chromatin structure is independent of Foxa1/2 in the adult liver. (a) Promoter-proximal Foxa2 binding sites are frequently surrounded by CpG islands. Distribution of CpG islands and nucleosomes surrounding Foxa2 binding sites. (b) Distribution of CpG bounded Foxa2 binding sites upstream and downstream of transcriptional start sites. (c) Intersection of differentially expressed genes in Foxa1/2 mutant liver and Foxa2 ChIP-Seq data. Gene expression data were differentially expressed genes from microarray analysis between Foxa1loxP/loxP;Foxa2loxP/loxP;AlfpCre (Mutant) and control mice; Foxa2 targets were Foxa2-targeted genes from ChIP-Seq analysis. (d) Distribution of Foxa2 binding sites upstream and downstream of transcriptional start sites for those sites associated with directly-regulated genes. A total of 369 Foxa2 binding sites were associated with the 197 differentially expressed genes. (e) Nucleosome occupancy surrounding Foxa2 binding sites associated with genes including thymocyte selection-associated high mobility group box (Tox), cAMP specific phosphodiesterase 4B (Pde4b), hydroxy-delta-5-steroid dehydrogenase, 3 beta- and steroid delta-isomerase 5 (Hsd3b5), transthyretin (Ttr), glutathionine S-transferase m1 (Gstm1), albumin promoter (Alb_p), alpha fetoprotein enhancer (Afp_e), sulfotransferase family 3A, member 1 (Sult3a1), the albumin enhancer (Alb1_e), gene model 129 (GM129), ribosomal protein S27A (Rps27a), protein phosphatase 1, regulatory (inhibitor) subunit 3C (Ppp1r3c), COBW domain containing 1 (Cbwd1), poly A polymerase alpha (Pap), ribonuclease L (Rnasel) and PR domain containing 6 (Prdm6), in livers from Foxa1loxP/loxP;Foxa2loxP/loxP;AlfpCre (Mutant) and control mice, mouse heart and mouse embryonic stem cells (ES Cell). (f, g) Distribution of heterochromatin protein 1 (Hp1), CBP, and p300 surrounding Foxa2 binding sites, or Hp1 surrounding H3K9me3 sites in control (f) and Foxa1/2 mutant (g) livers. 0 corresponds to the center of Foxa2 binding sites or H3K9me3 sites, respectively.
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