Mammalian promoters can be separated into two classes, conserved TATA box-enriched promoters, which initiate at a well-defined site, and more plastic, broad and evolvable CpG-rich promoters. We have sequenced tags corresponding to several hundred thousand transcription start sites (TSSs) in the mouse and human genomes, allowing precise analysis of the sequence architecture and evolution of distinct promoter classes. Different tissues and families of genes differentially use distinct types of promoters. Our tagging methods allow quantitative analysis of promoter usage in different tissues and show that differentially regulated alternative TSSs are a common feature in protein-coding genes and commonly generate alternative N termini. Among the TSSs, we identified new start sites associated with the majority of exons and with 3' UTRs. These data permit genome-scale identification of tissue-specific promoters and analysis of the cis-acting elements associated with them.

Genome-wide analysis of mammalian promoter architecture and evolution

Neutrophil kinetics in health and disease

Toll-Like Receptors on Hematopoietic Progenitor Cells Stimulate Innate Immune System Replenishment

Molecular interactions between protein complexes and DNA carry out essential gene regulatory functions. Uncovering such interactions by means of chromatin-immunoprecipitation coupled with massively parallel sequencing (ChIP-Seq) has recently become the focus of intense interest. We here introduce QuEST (Quantitative Enrichment of Sequence Tags), a powerful statistical framework based on the Kernel Density Estimation approach, which utilizes ChIP-Seq data to determine positions where protein complexes come into contact with DNA. Using QuEST, we discovered several thousand binding sites for the human transcription factors SRF, GABP and NRSF at an average resolution of about 20 base-pairs. MEME-based motif analyses on the QuEST-identified sequences revealed DNA binding by cofactors of SRF, providing evidence that cofactor binding specificity can be obtained from ChIP-Seq data. By combining QuEST analyses with gene ontology (GO) annotations and expression data, we illustrate how general functions of transcription factors can be inferred.

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Genome-wide analysis of transcription factor binding sites based on ChIP-Seq data.

Bifurcation dynamics in lineage-commitment in bipotent progenitor cells.

GA-binding protein (GABP) is an ets transcription factor that controls gene expression in several important biological settings. It is unique among ets factors, since the transcriptionally active complex is an obligate heterotetramer that is composed of two distinct proteins. GABPalpha includes an ets DNA binding domain (DBD), while a distinct protein, GABPbeta, contains ankyrin repeats and the transcriptional activation domain (TAD). GABP was first identified as a regulator of viral genes and nuclear respiratory factors. However, GABP is now recognized to be a key transcriptional regulator of dynamically regulated, lineage-restricted genes, especially in myeloid cells and at the neuromuscular junction. Furthermore, it regulates genes that are intimately involved in cell cycle control, protein synthesis, and cellular metabolism. GABP acts as an integrator of cellular signaling pathways by regulating key hormones and transmembrane receptors. In addition, GABP itself, is a target of phosphorylation events that lie downstream of signal transduction pathways. The physical and functional interactions of GABPalpha and GABPbeta with each other and with other transcription factors and co-activators are key to its ability to regulate gene expression. Its role in regulating genes involved in fundamental cellular processes places GABP at the nexus of key cellular pathways and functions.

GA-binding protein transcription factor: a review of GABP as an integrator of intracellular signaling and protein-protein interactions.

Transcriptional regulation in myelopoiesis: Hematopoietic fate choice, myeloid differentiation, and leukemogenesis

Centrins in vertebrates have traditionally been associated with microtubule-nucleating centers such as the centrosome. Unexpectedly, we found centrin 2 to associate biochemically with nucleoporins, including the Xenopus laevis Nup107-160 complex, a critical subunit of the vertebrate nuclear pore in interphase and of the kinetochores and spindle poles in mitosis. Immunofluorescence of Xenopus cells and in vitro reconstituted nuclei indeed revealed centrin 2 localized at the nuclear pores. Use of the mild detergent digitonin in immunofluorescence also allowed centrin 2 to be clearly visualized at the nuclear pores of human cells. Disruption of nuclear pores using RNA interference of the pore assembly protein ELYS/MEL-28 resulted in a specific decrease of centrin 2 at the nuclear rim of HeLa cells. Functionally, excess expression of either the Nor C-terminal calcium-binding domains of human centrin 2 caused a dominant-negative effect on both mRNA and protein export, leaving protein import intact. The mRNA effect mirrors that found for the Saccharomyes cerevisiae centrin Cdc31p at the yeast nuclear pore, a role until now thought to be unique to yeast. We conclude that in vertebrates, centrin 2 interacts with major subunits of the nuclear pore, exhibits nuclear pore localization, and plays a functional role in multiple nuclear export pathways.

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Centrin 2 Localizes to the Vertebrate Nuclear Pore and Plays a Role in mRNA and Protein Export

GA-binding protein (GABP) and Sp1 are required, along with retinoid receptors, to mediate retinoic acid responsiveness of CD18 (beta 2 leukocyte integrin): a novel mechanism of transcriptional regulation in myeloid cells.

Gene transcription plays a critical role in the differentiation of myeloid cells. However, there is no single, master regulator of all myeloid genes. Rather, myeloid gene transcription is regulated by the combinatorial effects of a limited number of key transcription factors. Sp1 is a powerful activator of gene transcription in many cell types. Although it is wildly expressed, Sp1 binds and activates the promoters of a large number of important myeloid genes. This presents the paradox of how a widely expressed transcription factor can regulate lineage-specific gene transcription. This review discusses the structure, function, and expression patterns of Sp1 and its related Sp family members. Illustrative examples of the tissue-specific regulation of myeloid target genes are presented. The roles of post-translational modifications of Sp1, alterations in target gene chromatin structure, and important cooperating transcription factors are discussed. Thus, Sp1 serves as a model of how a widely expressed transcription factor regulates the expression of tissue-specific genes.

Karen K. Resendes

Papers

GA-binding protein transcription factor: a review of GABP as an integrator of intracellular signaling and protein-protein interactions.

Transcriptional regulation in myelopoiesis: Hematopoietic fate choice, myeloid differentiation, and leukemogenesis

Centrin 2 Localizes to the Vertebrate Nuclear Pore and Plays a Role in mRNA and Protein Export

GA-binding protein (GABP) and Sp1 are required, along with retinoid receptors, to mediate retinoic acid responsiveness of CD18 (beta 2 leukocyte integrin): a novel mechanism of transcriptional regulation in myeloid cells.

Sp1 control of gene expression in myeloid cells.