Most nucleosomes are well-organized at the 5Ј ends of S. cerevisiae genes where "−1" and "+1" nucleosomes bracket a nucleosome-free promoter region (NFR). How nucleosomal organization is specified by the genome is less clear. Here we establish and inter-relate rules governing genomic nucleosome organization by sequencing DNA from more than one million immunopurified S. cerevisiae nucleosomes (displayed at http://atlas.bx.psu.edu/). Evidence is presented that the organization of nucleosomes throughout genes is largely a consequence of statistical packing principles. The genomic sequence specifies the location of the −1 and +1 nucleosomes. The +1 nucleosome forms a barrier against which nucleosomes are packed, resulting in uniform positioning, which decays at farther distances from the barrier. We present evidence for a novel 3Ј NFR that is present at >95% of all genes. 3Ј NFRs may be important for transcription termination and anti-sense initiation. We present a high-resolution genome-wide map of TFIIB locations that implicates 3Ј NFRs in gene looping.

Comparative genomics of nucleosome positions provides a powerful means for understanding how the organization of chromatin and the transcription machinery co-evolve. Here we produce a high resolution reference map of H2A.Z and bulk nucleosome locations across the genome of the fly D. melanogaster, and compare it to that from the yeast S. cerevisiae. Like Saccharomyces, Drosophila nucleosomes are organized around active transcription start sites in a canonical −1, NFR (nucleosome-free region), +1 arrangement. However, Drosophila does not incorporate H2A.Z into the −1 nucleosome and does not bury its transcriptional start site in the +1 nucleosome. At thousands of genes, RNA polymerase II engages the +1 nucleosome and pauses. How the transcription initiation machinery contends with the +1 nucleosome appears to be fundamentally different between lower and higher eukaryotes.Knowledge of the precise location of nucleosomes in a genome is essential in order to understand the context in which chromosomal processes such as transcription and DNA replication operate. A common theme to emerge from recent genome-wide maps of nucleosome locations is a general deficiency of nucleosomes in promoter regions and an enrichment of certain histone modifications towards the 5′ end of genes [1][2][3][4][5][6][7] . A high resolution genomic map of nucleosome locations in the budding yeast S. cerevisiae has further revealed Correspondence and request for material should be addressed to B.F.P. (bfp2@psu.edu). * These authors contributed equally to this work.Author Information Sequence data deposition is through NCBI Trace Archives TI SRA000283, Sequencing Center = "CCGB", and microarray deposition through ArrayExpress, Accession numbers E-MEXP-1515 and -1519. Reprints and permissions information is available at www.nature.com/reprints. The authors declare no competing financial interest.Author Contributions T.M. prepared and purified the nucleosomes including Pol II-bound nucleosomes; C.J. analyzed the nucleosome mapping data and its relationship to other genomic features; I.P.I. performed computational analyses related to nucleosome positioning sequences; X.L. conducted ChIP-chip on Pol II; B.J.V. conducted ChIP-chip on nucleosome-Pol II interactions; S.J.Z. provided bioinformatics support; L.T. constructed libraries and sequenced nucleosomal DNA; J.Q. mapped sequencing reads to the yeast genome; RG provided H2A.Z antibodies; SCS directed the DNA sequencing phase; DSG directed embryo preparations and helped interpret the data; I.A. developed computational approaches to derive nucleosome maps from the read locations and developed the associated browser; B.F.P. directed the project, interpreted the data, and wrote the paper. S6). Those 112,750 nucleosomes detected three or more times were further analyzed, although patterns were identical when all nucleosomes were analyzed. The internal median error of the data was 4 bp (Fig. S7). H2A.Z nucleosomes were predominantly distributed at 175 bp intervals from the TSS (compared to 165 ...

SUMMARY Hundreds of different proteins regulate and implement transcription in Saccharomyces. Yet their inter-relationships have not been investigated on a comprehensive scale. Here we determined the genome-wide binding locations of 200 transcription-related proteins, under normal and acute heat shock conditions. This study distinguishes binding between distal versus proximal promoter regions as well as the 3' ends of genes for nearly all mRNA and tRNA genes. This study reveals 1) a greater diversity and specialization of regulation associated with the SAGA transcription pathway compared to the TFIID pathway, 2) new regulators enriched at tRNA genes, 3) a global co-occupancy network of >20,000 unique regulator combinations that show a high degree of regulatory interconnections among lowly-expressed genes, 4) regulators of the SAGA pathway located largely distal to the core promoter and regulators of the TFIID pathway located proximally, and 5) distinct mobilization of SAGA- versus TFIID-linked regulators during acute heat shock.

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The Pol II transcription machinery assembles and operates in the context of nucleosomes, which have well‐defined canonical positions at the beginning and end of protein‐coding genes. However, the predominant organizational theme by which the transcription machinery and chromatin regulators are positioned within promoter regions or throughout genes in a genome is largely unknown. We have mapped the genomic location of diverse representative components of the gene regulatory machinery in Saccharomyces cerevisiae to an experimental resolution of <40 bp using ChIP‐chip. Defining how these factors are arranged with respect to each other and to promoter nucleosome positions should provide insight into how the organization of the transcription machinery at promoters regulates gene expression. We find that the ‐1 nucleosome, which establishes the upstream border of the promoter region, is a likely target of substantial regulation (assuming that occupancy is linked to function) since nearly all tested chromatin remodeling complexes selectively occupy the ‐1 nucleosomal region compared to the +1 region. Furthermore, we find that the ‐1 nucleosome is depleted rather than laterally displaced when Pol II is present, but not when only TBP and TFIIB are recruited. This suggests that the ‐1 nucleosome is removed during PIC assembly, after TBP and TFIIB have been recruited, but before or concurrent with Pol II recruitment, which might explain the prevalence of remodeling complexes in the vicinity of the ‐1 nucleosome. This work was supported by NIH grant ES013768.

Summary We report that p73 is expressed in multiciliated cells (MCCs), is required for MCC differentiation, and directly regulates transcriptional modulators of multiciliogenesis. Loss of ciliary biogenesis provides a unifying mechanism for many phenotypes observed in p73 knockout mice including hydrocephalus, hippocampal dysgenesis, sterility and chronic inflammation/infection of lung, middle ear and sinus. Through p73 and p63 ChIP-seq using murine tracheal cells, we identified over 100 putative p73 target genes that regulate MCC differentiation and homeostasis. We validated Foxj1, a transcriptional regulator of multiciliogenesis, and many other cilia-associated genes as direct target genes of p73 and p63. We show p73 and p63 are co-expressed in a subset of basal cells, and suggest that p73 ‘marks’ these cells for MCC differentiation. In sum, p73 is essential for MCC differentiation, functions as a critical regulator of a transcriptome required for MCC differentiation and, like p63, has an essential role in development of tissues.

Linker histone H1 proteins bind to nucleosomes and facilitate chromatin compaction 1 , although their biological functions are poorly understood. Histone H1 ( HIST1H1B-E ) mutations are highly recurrent in B-cell lymphomas, but their cancer relevance and mechanism are unknown. Here we show that lymphoma-associated H1 alleles are genetic driver mutations in lymphomas. Disruption of H1 function results in profound architectural remodeling of the genome characterized by large-scale, yet focal shifts of chromatin from a compacted, to a relaxed state. This decompaction drives distinct changes in epigenetic states, primarily due to gain of histone H3 lysine 36 dimethylation, and/or loss of repressive H3 lysine 27 trimethylation. These changes unlock expression of stem cell genes that are normally silenced during early development. Loss of H1c and H1e alleles in mice conferred enhanced fitness and self-renewal properties to germinal center B-cells, ultimately leading to aggressive lymphoma with enhanced repopulating potential. Collectively, our data indicate that H1 proteins are normally required to sequester early developmental genes into architecturally inaccessible genomic compartments. We furthermore establish H1 as a bona fide tumor suppressor, whose mutation drives malignant transformation primarily through three-dimensional genome reorganization, followed by epigenetic reprogramming and derepression of developmentally silenced genes.

Protein Arg methyltransferases function as coactivators of the tumor suppressor p53 to regulate gene expression. Peptidylarginine deiminase 4 (PAD4/PADI4) counteracts the functions of protein Arg methyltransferases in gene regulation by deimination and demethylimination. Here we show that the expression of a tumor suppressor gene, OKL38, is activated by the inhibition of PAD4 or the activation of p53 following DNA damage. Chromatin immunoprecipitation assays showed a dynamic change of p53 and PAD4 occupancy and histone Arg modifications at the OKL38 promoter during DNA damage, suggesting a direct role of PAD4 and p53 in the expression of OKL38. Furthermore, we found that OKL38 induces apoptosis through localization to mitochondria and induction of cytochrome c release. Together, our studies identify OKL38 as a novel p53 target gene that is regulated by PAD4 and plays a role in apoptosis.