The nucleosome is the fundamental building block of eukaryotic chromosomes. Access to genetic information encoded in chromosomes is dependent on the position of nucleosomes along the DNA. Alternative locations just a few nucleotides apart can have profound effects on gene expression. Yet the nucleosomal context in which chromosomal and gene regulatory elements reside remains ill-defined on a genomic scale. Here we sequence the DNA of 322,000 individual Saccharomyces cerevisiae nucleosomes, containing the histone variant H2A.Z, to provide a comprehensive map of H2A.Z nucleosomes in functionally important regions. With a median 4-base-pair resolution, we identify new and established signatures of nucleosome positioning. A single predominant rotational setting and multiple translational settings are evident. Chromosomal elements, ranging from telomeres to centromeres and transcriptional units, are found to possess characteristic nucleosomal architecture that may be important for their function. Promoter regulatory elements, including transcription factor binding sites and transcriptional start sites, show topological relationships with nucleosomes, such that transcription factor binding sites tend to be rotationally exposed on the nucleosome surface near its border. Transcriptional start sites tended to reside about one helical turn inside the nucleosome border. These findings reveal an intimate relationship between chromatin architecture and the underlying DNA sequence it regulates.

Despite being one of the first eukaryotic transcriptional regulatory elements identified, the sequence of a native TATA box and its significance remain elusive. Applying criteria associated with TATA boxes we queried several Saccharomyces genomes and arrived at the consensus TATA(A/T)A(A/T)(A/G). Approximately 20% of yeast genes contain a TATA box. Strikingly, TATA box-containing genes are associated with responses to stress, are highly regulated, and preferentially utilize SAGA rather than TFIID when compared to TATA-less promoters. Transcriptional regulation in yeast appears to be mechanistically bipolar, possibly reflecting a need to balance inducible stress-related responses with constitutive housekeeping functions.

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 ...

DNA sequence has long been recognized as an important contributor to nucleosome positioning, which has the potential to regulate access to genes. The extent to which the nucleosomal architecture at promoters is delineated by the underlying sequence is now being worked out. Here we use comparative genomics to report a genome-wide map of nucleosome positioning sequences (NPSs) located in the vicinity of all Saccharomyces cerevisiae genes. We find that the underlying DNA sequence provides a very good predictor of nucleosome locations that have been experimentally mapped to a small fraction of the genome. Notably, distinct classes of genes possess characteristic arrangements of NPSs that may be important for their regulation. In particular, genes that have a relatively compact NPS arrangement over the promoter region tend to have a TATA box buried in an NPS and tend to be highly regulated by chromatin modifying and remodeling factors.

Eukaryotic genes are controlled by sequence-specific DNA-binding proteins, chromatin regulators, general transcription factors, and elongation factors. Here we examine the genome-wide location of representative members of these groups and their redistribution when the Saccharomyces cerevisiae genome is reprogrammed by heat shock. As expected, assembly of active transcription complexes is coupled to eviction of H2A.Z nucleosomes, and disassembly is coupled to the return of nucleosomes. Remarkably, a large number of promoters assemble into partial preinitiation complexes (partial PICs), containing TFIIA, TFIID (and/or SAGA), TFIIB, TFIIE, and TFIIF. However, RNA polymerase II and TFIIH are generally not recruited, and nucleosomes are not displaced. These promoters may be preparing for additional stress that naturally accompany heat stress. For example, we find that oxidative stress, which often occurs with prolonged exposure of cells to high temperature, converts partial PICs into full PICs. Partial PICs therefore represent novel regulated intermediates that assemble at promoters in the midst of chromatin.[Keywords: ChIP-chip; GTF; preinitiation complex; microarrays; Saccharomyces; H2A.Z] Supplemental material is available at http://www.genesdev.org.