Enhancers are short regions of DNA that are

Enhancers are short regions of DNA that are bound by DNA-binding proteins (TFs) to increase the likelihood of a particular gene to be expressed. Importantly, enhancers can be identified, and classified, by their patterns of chromatin accessibility and of enrichment in certain histone modifications. Enhancers are often far away from their target genes and need to be brought in proximity to their target promoters via DNA looping. During development, both TF activity and gene expression programs as well as cell type-specifc enhancers vary in a spatiotemporal manner. Recent reports have clarified the role of AP-1 in establishing cell type-specific enhancers and gene expression programs in macrophages and fibroblasts. Moreover, parallel studies reveal that disruption of these programs is necessary for the successful reprogramming of fibroblasts to induced pluripotent stem Endoxifen with Yamanaka factors or for chemical induction of pluripotency , , , , . AP-1, a ubiquitously expressed heterodimeric TF comprising proteins belonging to the JUN, FOS, ATF, MAF, and CREB families, plays a pivotal role in differentiation, apoptosis, and proliferation and is activated by Ras–MAPK signaling in almost all cell types. However, in cooperation with other cell type-specific TFs, AP-1 is also involved in establishing cell type-specific enhancers and cell identities. This role is well recognized in macrophages, where AP-1 cooperatively binds to the DNA together with PU.1 (SPI1) and C/EBP family factors . This cooperative binding of a small set of cell type-specific TFs is likely to be a general mechanism that gives rise to cell type-specific enhancers . Importantly, the combination of factors, rather than the individual factors themselves, is specific to a given cell type. For example, PU.1 is also important in establishing the B cell enhancers, where it collaborates with other factors. Despite the established role of AP-1 in macrophage enhancer selection, the molecular mechanisms by which AP-1 contributes to cell type-specific enhancer formation and how these enhancers are brought in contact with their distal target genes has remained elusive. A recent study by Phanstiel . investigated DNA loops during macrophage differentiation from human monocytic leukemia THP-1 cells using Hi-C , which maps contact frequencies between genomic regions. Phanstiel . found more loops gained than lost during differentiation, suggesting that gained and preformed chromatin loops acquire enhancer activity during differentiation, connecting single promoters to multiple distal enhancers. Those interactive hubs that link multiple enhancers to gene promoters could bring multiple accessible chromatin regions into close proximity during differentiation. TF footprinting analysis of assay of transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) in this experimental system detected enrichment of sequence motifs of FOS, JUN, and other AP-1 related proteins, which paralleled upregulation in gene expression. When TF footprints were correlated with anchors of different classes of DNA loops detected by Hi-C, activated and gained DNA loops were far more enriched (compared with lost, deactivated, or static classes) for AP-1 than in the CTCF motif, a multifunctional zinc-finger protein that regulates chromatin architecture. However, due to uncertainty in bioinformatic footprint identification, future work should focus on determining which specific members of the AP-1 complex (JUN, FOS, or MAF protein families) are involved in loop formation. Furthermore, it is likely that other TFs in addition to AP-1 are capable of DNA loop formation in different cell types and contexts, as highlighted by studies of the developmentally controlled beta-globin locus . In addition to macrophages, AP-1 might also contribute to cell type-specific enhancer selection in fibroblasts . By using ATAC-seq and H3K27ac ChIP-seq, Vierbuchen . characterized active versus primed enhancers during differentiation of G-phase synchronized mouse embryonic fibroblasts (MEFs) after serum stimulation and associated them with cell-identity and late-response genes (LRGs); 82% of LRG enhancers and 67% of cell-identity enhancers contained the AP-1 motif, and the enrichment was much higher than that found for motifs of known fibroblast-specific TFs (e.g., TEAD, ETS, CREB/ATF). Vierbuchen . then performed an unbiased genetic approach using two different mouse strains to identify SNPs that might disrupt TF binding associated with enhancer selection. More SNPs in AP-1 motifs were present in strain-specific and LRG enhancers. Not only did disruption of the AP-1 motif inhibit or decrease chromatin accessibility, H3K4me1, and H3K27ac in strain-specific enhancers, but AP-1 also collaborated with cell type-specific TFs in LRG enhancers in fibroblasts. AP-1 established accessible chromatin by recruiting the BAF ATP-dependent chromatin-remodeling complex. Moreover, a subset of AP-1 binding occurred in nucleosome-occluded enhancers, suggesting that AP-1 acts as a pioneer TF that is able to displace nucleosomes and establish new enhancers .