Summary Genome-scale studies have revealed extensive, cell type-specific co-localization of transcription factors, but the mechanisms underlying this phenomenon remain poorly understood. Here we demonstrate in macrophages and B cells that collaborative interactions of the common factor PU.1 with small sets of macrophage- or B celllineage-determining transcription factors establish cell-specific binding sites that are associated with the majority of promoter-distal H3K4me1-marked genomic regions. PU.1 binding initiates nucleosome remodeling followed by H3K4 monomethylation at large numbers of genomic regions associated with both broadly and specifically expressed genes. These locations serve as beacons for additional factors, exemplified by liver X receptors, which drive both cell-specific gene expression and signal-dependent responses. Together with analyses of transcription factor binding and H3K4me1 patterns in other cell types, these studies suggest that simple combinations of lineage-determining transcription factors can specify the genomic sites ultimately responsible for both cell identity and cell type-specific responses to diverse signaling inputs.

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The helix-loop-helix (HLH) family of transcriptional regulatory proteins are key players in a wide array of developmental processes. Over 240 HLH proteins have been identified to date in organisms ranging from the yeast Saccharomyces cerevisiae to humans (6). Studies in Xenopus laevis, Drosophila melanogaster, and mice have convincingly demonstrated that HLH proteins are intimately involved in developmental events such as cellular differentiation, lineage commitment, and sex determination. In yeast, HLH proteins regulate several important metabolic pathways, including phosphate uptake and phospholipid biosynthesis (19,67,112). In multicellular organisms, HLH factors are required for a multitude of important developmental processes, including neurogenesis, myogenesis, hematopoiesis, and pancreatic development (12,86,127,179). The purpose of this review is to examine the structure and functional properties of HLH proteins.E-box sites: elements mediating cell-type-specific gene transcription. Gene transcription of the immunoglobulin heavychain (IgH) gene has long been known to be regulated, in part, by a cis-acting DNA element known as the IgH intronic enhancer (109,156). By in vivo methylation protection assays, a number of sites were identified in both the IgH and the kappa light-chain gene enhancers which were specifically protected in B cells but not in nonlymphoid cells (41). These elements shared a signature motif which consisted of the core hexanucleotide sequence, CANNTG, and were subsequently dubbed E boxes (41). A total of five E-box elements are present in the IgH gene enhancer: E1, E2, E3, E4, and E5. The Ig kappa enhancer also contains three cannonical E boxes, designated E1, E2, and E3. E-box sites have been subsequently found in B-cell-specific promoter and enhancer elements, including a subset of Ig light-chain gene promoters, the IgH and Ig light-chain 3Ј enhancers, and, more recently, the 5 promoter (110,118,156).E-box elements have also been identified in promoter and enhancer elements that regulate muscle-, neuron-, and pancreas-specific gene expression. For example, in muscle, the muscle creatine kinase gene, acetylcholine receptor genes ␣ and ␦, and the myosin light-chain gene all require E-box elements for full activity (27,51,85). A number of genes whose expression is limited to the pancreas also require E-box sites for proper expression. The insulin and somatostatin genes, for example, contain E-box sites that, when multimerized, are sufficient to regulate pancreatic ␤-cell-specific gene expression (168). More recently, E-box regulatory sites have been identified in a number of neuron-specific genes, including the opsin, hippocalcin, beta 2 subunit of the neuronal nicotinic acetylcholine receptor, and muscarinic acetylcholine receptor genes (1, 21, 52, 125).E-box sites: cognate recognition sequence for HLH proteins. Two proteins, termed E12 and E47, were originally identified as binding to the E2/E5 site (65, 102). They have a region of homology with the Drosophila Daughterless protein, the...

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It is now established that the transcription factors E2A, EBF1 and Foxo1 play critical roles in B cell development. Here we show that E2A and EBF1 bound regulatory elements present in the Foxo1 locus. E2A and EBF1 as well as E2A and Foxo1, in turn, were wired together by a vast spectrum of cis-regulatory codes. These associations were dynamic during developmental progression. Occupancy by the E2A isoform, E47, directly elevated the abundance as well as the pattern of histone H3K4 monomethylation across putative enhancer regions. Finally, the pro-B cell epigenome was divided into clusters of loci that show E2A, EBF and Foxo1 occupancy. From this analysis a global network consisting of transcriptional regulators, signaling and survival factors, was constructed that we propose orchestrates the B cell fate.

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The E2A gene products, E12 and E47, are critical for proper early B-cell development and commitment to the B-cell lineage. Here we reveal a new role for E2A in T-lymphocyte development. Loss of E2A activity results in a partial block at the earliest stage of T-lineage development. This early T-cell phenotype precedes the development of a T-cell lymphoma which occurs between 3 and 9 months of age. The thymomas are monoclonal and highly malignant and display a cell surface phenotype similar to that of immature thymocytes. In addition, the thymomas generally express high levels of c-myc. As assayed by comparative genomic hybridization, each of the tumor populations analyzed showed a nonrandom gain of chromosome 15, which contains the c-myc gene. Taken together, the data suggest that the E2A gene products play a role early in thymocyte development that is similar to their function in B-lineage determination. Furthermore, the lack of E2A results in development of T-cell malignancies, and we propose that E2A inactivation is a common feature of a wide variety of human T-cell proliferative disorders, including those involving the E2A heterodimeric partners tal-1 and lyl-1.The E2A gene encodes two basic helix-loop-helix (HLH) transcription factors, E12 and E47 (32). E12 and E47, members of the class I HLH proteins, are characterized by their broad expression pattern and their ability to bind DNA either as homodimers or as heterodimers with tissue-specific HLH proteins (9,22,33,40,43). Class I HLH proteins share several highly conserved domains. The HLH domain mediates homoand/or heterodimerization, and the basic region constitutes the sequence-specific DNA binding domain (11,22,32,50). In addition, two distinct domains located in the N-terminal portion of the class I HLH proteins have been shown to be required for transactivation (1,26,39).E12 and E47 arise through differential splicing to the exon that encodes for the HLH domain. Within the HLH domain, their amino acid sequences differ by 20% (32). Both E12 and E47 have the ability to form heterodimers with class II HLH members, including the myogenic regulators (9, 23). However, E12 and E47 have distinct biochemical properties. E47 homodimers bind with high affinity to DNA, whereas an inhibitory domain present in E12 prevents those homodimers from high-affinity DNA binding (46).E2A polypeptides bind to E-box sites present in a wide variety of tissue-specific enhancers, including the insulin, muscle creatine kinase, and immunoglobulin (Ig) intronic and 3Ј enhancers (16,22,28,32,35,38,52). In B cells, it is homodimers of the E2A gene products that bind to E2-box sites present in the Ig enhancers (2,34,43). That E2A gene products play a crucial role in B lymphocyte development has been demonstrated recently by the generation of E2A-deficient mice and transgenic mice overexpressing Id1, an inhibitor of E2A (3,47,56). In the absence of E2A activity, even the earliest committed B-cell precursors are undetectable in the bone marrow (3,47,56). In addition, E2A-deficient mice l...