High-resolution nucleosome mapping reveals transcription-dependent promoter packaging

  1. Assaf Weiner1,
  2. Amanda Hughes2,
  3. Moran Yassour1,3,
  4. Oliver J. Rando2,5 and
  5. Nir Friedman1,4,5
  1. 1 School of Computer Science and Engineering, The Hebrew University, Jerusalem 91904, Israel;
  2. 2 Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA;
  3. 3 The Broad Institute of Harvard and MIT, Cambridge, Massachusetts 02142, USA;
  4. 4 Institute of Life Sciences, The Hebrew University, Jerusalem 91904, Israel

    Abstract

    Genome-wide mapping of nucleosomes has revealed a great deal about the relationships between chromatin structure and control of gene expression, and has led to mechanistic hypotheses regarding the rules by which chromatin structure is established. High-throughput sequencing has recently become the technology of choice for chromatin mapping studies, yet analysis of these experiments is still in its infancy. Here, we introduce a pipeline for analyzing deep sequencing maps of chromatin structure and apply it to data from S. cerevisiae. We analyze a digestion series where nucleosomes are isolated from under- and overdigested chromatin. We find that certain classes of nucleosomes are unusually susceptible or resistant to overdigestion, with promoter nucleosomes easily digested and mid-coding region nucleosomes being quite stable. We find evidence for highly sensitive nucleosomes located within “nucleosome-free regions,” suggesting that these regions are not always completely naked but instead are likely associated with easily digested nucleosomes. Finally, since RNA polymerase is the dominant energy-consuming machine that operates on the chromatin template, we analyze changes in chromatin structure when RNA polymerase is inactivated via a temperature-sensitive mutation. We find evidence that RNA polymerase plays a role in nucleosome eviction at promoters and is also responsible for retrograde shifts in nucleosomes during transcription. Loss of RNA polymerase results in a relaxation of chromatin structure to more closely match in vitro nucleosome positioning preferences. Together, these results provide analytical tools and experimental guidance for nucleosome mapping experiments, and help disentangle the interlinked processes of transcription and chromatin packaging.

    Footnotes

    • 5 Corresponding authors.

      E-mail nir{at}cs.huji.ac.il; fax +972-2-6585727.

      E-mail oliver.rando{at}umassmed.edu; fax (508) 856-6464.

    • [Supplemental material is available online at http://www.genome.org. The microarray data and the sequencing data from this study have been submitted to the NCBI Gene Expression Omnibus (http://www.ncbi.nlm.nih.gov/geo) under accession nos. GSE18629 and GSE18530.]

    • Article published online before print. Article and publication date are at http://www.genome.org/cgi/doi/10.1101/gr.098509.109.

      • Received July 16, 2009.
      • Accepted October 15, 2009.
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