Active chromatin domains are defined by acetylation islands revealed by genome-wide mapping

Abstract

The identity and developmental potential of a human cell is specified by its epigenome that is largely defined by patterns of chromatin modifications including histone acetylation. Here we report high-resolution genome-wide mapping of diacetylation of histone H3 at Lys 9 and Lys 14 in resting and activated human T cells by genome-wide mapping technique (GMAT). Our data show that high levels of the H3 acetylation are detected in gene-rich regions. The chromatin accessibility and gene expression of a genetic domain is correlated with hyperacetylation of promoters and other regulatory elements but not with generally elevated acetylation of the entire domain. Islands of acetylation are identified in the intergenic and transcribed regions. The locations of the 46,813 acetylation islands identified in this study are significantly correlated with conserved noncoding sequences (CNSs) and many of them are colocalized with known regulatory elements in T cells. TCR signaling induces 4045 new acetylation loci that may mediate the global chromatin remodeling and gene activation. We propose that the acetylation islands are epigenetic marks that allow prediction of functional regulatory elements.

Figure 1.

High levels of the H3 acetylation are detected in the promoter regions. (A) A schematic showing the human genome arbitrarily separated into three parts: 2-kb promoter regions, intergenic regions, and gene body regions. (B, left) Calculated percentage of each region in the genome. (Right) The percentage of the tags detected in the GMAT library from each region. (C) A 10-kb region of 21,355 genes was aligned relative to their transcription initiation sites (X-axis). The Y-axis shows the tag density that was obtained by normalizing the total number of detected tags with the number of expected NlaIII sites in a 50-bp window. The green line represents the calculated value, provided the tags were detected randomly. The black line represents the tag density of all of the 21,355 genes. The pink line represents the tag density of the 8000 highly active genes. The blue line represents the rest of the genes that are either silent or expressed at lower levels.

Figure 2.

CpG islands are highly acetylated. (A) CpG islands are highly acetylated. “Active” and “silent” indicate the CpG islands identified between the -10- and +10-kb regions in active genes and the genes whose expression was not detected, respectively. “Intergenic” indicates the CpG islands identified in the intergenic region beyond -10 kb upstream of the transcription initiation site. (B) Sharp boundaries of histone H3 acetylation are detected surrounding CpG islands. The detected GMAT tags within the 4973 CpG islands that have a size ranging from 1 to 2 kb were counted in a window of 10% of the CpG island length and normalized to the number of NlaIII sites in the window to get the tag density. The GMAT tags outside of the CpG islands were counted in a 0.2-kb window and normalized to the number of NlaIII sites in the window to get the tag density. Each bar represents 10% of the sequence length within the CpG islands and 0.2 kb outside of the CpG islands. Black bars indicate the tag density within the CpG islands and the gray bars indicate the tag density outside of the CpG islands.

Figure 3.

Active domains are not uniformly acetylated. (A) A gene-rich region on chromosome 12. The transcribed regions and gene orientations are indicated by the arrows. The acetylation levels of the locus in resting T cells are shown. The Y-axis indicates the detection frequency and the X-axis indicates the chromosome coordinate. The numbers above the broken lines indicate the detection frequency. (B) BAF53A locus. (C) The Sp1 locus. Acetylation islands are highlighted and numbered. (D) The STAT5B locus.

Figure 4.

Colocalization of acetylation islands with known regulatory elements. (A) CD4 locus. The upper panel shows the acetylation data, above which gene positions and known functional regulatory elements are indicated. The lower panel shows the VISTA human and mouse sequence comparison. (DE) Distal enhancer; (PE) proximal enhancer; (Pr) promoter; (Sil) silencer; (LCR) locus control region; (TE) thymocyte enhancer. The acetylation islands colocalized with known regulatory elements are highlighted in pink. The acetylation islands with no known functions are highlighted in green. (B) CD8 locus. The acetylation data and VISTA sequence analysis are shown as in A. The positions of the six clusters of DNase hypersensitive sites (HS) are indicated below the genes.

Figure 5.

Genome-wide changes of acetylation induced by TCR signaling. The average tag density was derived by normalizing the total number of detected tags to the number of NlaIII site in a 3-kb window. The average tag densities from resting and activated T cells were compared directly to obtain the fold change. Changes of threefold or more between activated and resting T cells were plotted along the chromosome coordinates. The data for chromosome 12 are shown. The other chromosomes are shown in Supplementary Figure 6.

Figure 6.

TCR signaling-induced acetylation islands activate transcription in a chromatin-dependent manner. (A) IL13/IL4 locus. The upper and middle panels show the acetylation data from resting and activated T cells, respectively. The lower panel shows the VISTA human and mouse sequence comparison. The gene locations are indicated above the acetylation data. Also indicated is the CNS-1 identified by Loots et al. (2000). (B) TCR signaling induces the expression of the IL13 and IL4 genes in T cells. Total RNA was isolated from resting T cells treated with anti-CD3 and anti-CD28 for 24 h. The expression of the IL13 and IL4 genes were analyzed by RT-PCR with specific primers. The 18s RNA was used as a control. (C) Acetylation Islands 1 and 2 have constitutive enhancer activity in a nonchromatin vector. The 1.5-kb DNA containing Acetylation Islands 1 and 2 in A or a control sequence from the neighboring unacetylated region (chromosome 5: 132,080,058-132,081,677) were inserted upstream of a minimal GM-CSF1 promoter in the pGL3 luciferase reporter vector. The constructs were transfected into Jurkat cells for 48 h, followed by stimulation with 1 μg/mL ionomycin and 10 ng/mL PMA for 15 h. The luciferase activity was analyzed with the dual luciferase system from Promega as described (Liu et al. 2001). (PI) PMA and Ionomycin. (D) The enhancer activity of Acetylation Islands 1 and 2 is dependent on TCR signaling in a chromatin-forming vector. The 1.5-kb DNA containing Acetylation Islands 1 and 2 in A or the control sequence from the neighboring unacetylated region were inserted upstream of a minimal GM-CSF1 promoter in the pREP4 luciferase reporter vector. The constructs were similarly transfected into Jurkat cells and analyzed as above.