Master regulatory genes require stable silencing by the polycomb group (PcG) to prevent misexpression during differentiation and development. Some PcG proteins covalently modify histones, which contributes to heritable repression. The role for other effects on chromatin structure is less understood. We characterized the organization of PcG target genes in ESCs and neural progenitors using 5C and super-resolution microscopy. The genomic loci of repressed PcG targets formed discrete, small (20-140 Kb) domains of tight interaction that corresponded to locations bound by canonical polycomb repressive complex 1 (PRC1). These domains changed during differentiation as PRC1 binding changed. Their formation depended upon the Polyhomeotic component of canonical PRC1 and occurred independently of PRC1-catalyzed ubiquitylation. PRC1 domains differ from topologically associating domains in size and boundary characteristics. These domains have the potential to play a key role in transmitting epigenetic silencing of PcG targets by linking PRC1 to formation of a repressive higher-order structure.

ATP-dependent chromatin-remodeling complexes are conserved among all eukaryotes and function by altering nucleosome structure to allow cellular regulatory factors access to the DNA. Mammalian SWI-SNF complexes contain either of two highly conserved ATPase subunits: BRG1 or BRM. To identify cellular genes that require mammalian SWI-SNF complexes for the activation of gene expression, we have generated cell lines that inducibly express mutant forms of the BRG1 or BRM ATPases that are unable to bind and hydrolyze ATP. The mutant subunits physically associate with at least two endogenous members of mammalian SWI-SNF complexes, suggesting that nonfunctional, dominant negative complexes may be formed. We determined that expression of the mutant BRG1 or BRM proteins impaired the ability of cells to activate the endogenous stress response gene hsp70 in response to arsenite, a metabolic inhibitor, or cadmium, a heavy metal. Activation of hsp70 by heat stress, however, was unaffected. Activation of the heme oxygenase 1 promoter by arsenite or cadmium and activation of the cadmium-inducible metallothionein promoter also were unaffected by the expression of mutant SWI-SNF components. Analysis of a subset of constitutively expressed genes revealed no or minimal effects on transcript levels. We propose that the requirement for mammalian SWI-SNF complexes in gene activation events will be specific to individual genes and signaling pathways.