Swi/Snf chromatin remodeling/tumor suppressor complex establishes nucleosome occupancy at target promoters

Abstract

Precise nucleosome-positioning patterns at promoters are thought to be crucial for faithful transcriptional regulation. However, the mechanisms by which these patterns are established, are dynamically maintained, and subsequently contribute to transcriptional control are poorly understood. The switch/sucrose non-fermentable chromatin remodeling complex, also known as the Brg1 associated factors complex, is a master developmental regulator and tumor suppressor capable of mobilizing nucleosomes in biochemical assays. However, its role in establishing the nucleosome landscape in vivo is unclear. Here we have inactivated Snf5 and Brg1, core subunits of the mammalian Swi/Snf complex, to evaluate their effects on chromatin structure and transcription levels genomewide. We find that inactivation of either subunit leads to disruptions of specific nucleosome patterning combined with a loss of overall nucleosome occupancy at a large number of promoters, regardless of their association with CpG islands. These rearrangements are accompanied by gene expression changes that promote cell proliferation. Collectively, these findings define a direct relationship between chromatin-remodeling complexes, chromatin structure, and transcriptional regulation.

Fig. 1.

Snf5- and Brg1-deficient cells display loss of nucleosome occupancy around the TSS. (A) Key parameters of the nucleosome landscape. We considered the possibility that loss of Swi/Snf could affect position (orange line), occupancy (green line), or spacing of phased nucleosomes (pink line). Genomewide nucleosome occupancy profiles at peri-TSS regions are shown for all genes (B), expressed genes (C), and silent genes (D). Profiles for WT and Snf5- and Brg1-deficient cells are shown with blue, pink, and green lines, respectively. (E) Immunoblot for nonchromatin-associated Histone H3 in WT and Snf5-deficient cells.

Fig. 2.

Snf5- and Brg1-deficient cells display altered nucleosomal phasing. (A) Alignment of WT, Snf5-deficient, and Brg1-deficient nucleosomal profiles. Thin vertical lines mark stable nucleosome positions in WT (solid lines) and Snf5 mutant cells (dotted lines). (B) Fourier analysis reveals internucleosomal distance of 182 bp in WT cells (blue line) and 174 bp in Snf5- and Brg1-deficient cells (pink and green lines, respectively).

Fig. 3.

Brg1 ChIP reveals enrichment of Swi/Snf at peri-TSS regions. (A) Brg1 binding is prominent at TSS. Results are shown for all (black), expressed (red), and silent (blue) genes. (B) Brg1 enrichment in the TSS region (+/−2 kb) displays a bimodal distribution. The red line indicates the threshold of Brg1 enrichment used to identify groups of genes with low and high Brg1 levels. (C and D) Nucleosomal occupancy profiles for Brg1 high and Brg1 low genes.

Fig. 4.

Effects of nucleosome occupancy and Brg1 binding on transcription. (A) Schematic illustration of the relative nucleosome occupancy score definition. (B) Comparison of the score in WT (blue), Snf5 (pink), and Brg1 (green) samples. (C) Average absolute changes in gene expression upon Brg1 inactivation are compared for genes with low and high Brg1 enrichment. The 95% confidence intervals and P values are indicated.

Fig. 5.

Swi/Snf sculpts the promoter-associated nucleosome landscape. Swi/Snf functions to establish the nucleosome landscape at the peri-TSS region by promoting occupancy at the −1 and +1 positions while ensuring relative depletion at the NDR.