Substantial Histone Reduction Modulates Genomewide Nucleosomal Occupancy and Global Transcriptional Output

Substantial Histone Reduction Modulates Genomewide Nucleosomal Occupancy and Global Transcriptional Output: "

by Barbara Celona, Assaf Weiner, Francesca Di Felice, Francesco M. Mancuso, Elisa Cesarini, Riccardo L. Rossi, Lorna Gregory, Dilair Baban, Grazisa Rossetti, Paolo Grianti, Massimiliano Pagani, Tiziana Bonaldi, Jiannis Ragoussis, Nir Friedman, Giorgio Camilloni, Marco E. Bianchi, Alessandra Agresti

The basic unit of genome packaging is the nucleosome, and nucleosomes have long been proposed to restrict DNA accessibility both to damage and to transcription. Nucleosome number in cells was considered fixed, but recently aging yeast and mammalian cells were shown to contain fewer nucleosomes. We show here that mammalian cells lacking High Mobility Group Box 1 protein (HMGB1) contain a reduced amount of core, linker, and variant histones, and a correspondingly reduced number of nucleosomes, possibly because HMGB1 facilitates nucleosome assembly. Yeast nhp6 mutants lacking Nhp6a and -b proteins, which are related to HMGB1, also have a reduced amount of histones and fewer nucleosomes. Nucleosome limitation in both mammalian and yeast cells increases the sensitivity of DNA to damage, increases transcription globally, and affects the relative expression of about 10% of genes. In yeast nhp6 cells the loss of more than one nucleosome in four does not affect the location of nucleosomes and their spacing, but nucleosomal occupancy. The decrease in nucleosomal occupancy is non-uniform and can be modelled assuming that different nucleosomal sites compete for available histones. Sites with a high propensity to occupation are almost always packaged into nucleosomes both in wild type and nucleosome-depleted cells; nucleosomes on sites with low propensity to occupation are disproportionately lost in nucleosome-depleted cells. We suggest that variation in nucleosome number, by affecting nucleosomal occupancy both genomewide and gene-specifically, constitutes a novel layer of epigenetic regulation."

Reshuffling genomic landscapes to study the regulatory evolution of Hox gene clusters [Genetics]

Reshuffling genomic landscapes to study the regulatory evolution of Hox gene clusters [Genetics]: "The emergence of Vertebrata was accompanied by two rounds of whole-genome duplications. This enabled paralogous genes to acquire novel functions with high evolutionary potential, a process suggested to occur mostly by changes in gene regulation, rather than in protein sequences. In the case of Hox gene clusters, such duplications favored the appearance of distinct global regulations. To assess the impact of such “regulatory evolution” upon neo-functionalization, we developed PANTHERE (PAN-genomic Translocation for Heterologous Enhancer RE-shuffling) to bring the entire megabase-scale HoxD regulatory landscape in front of the HoxC gene cluster via a targeted translocation in vivo. At this chimeric locus, Hoxc genes could both interpret this foreign regulation and functionally substitute for their Hoxd counterparts. Our results emphasize the importance of evolving regulatory modules rather than their target genes in the process of neo-functionalization and offer a genetic tool to study the complexity of the vertebrate regulatory genome."

p53 binding to nucleosomes within the p21 promoter in vivo leads to nucleosome loss and transcriptional activation [Biochemistry]

p53 binding to nucleosomes within the p21 promoter in vivo leads to nucleosome loss and transcriptional activation [Biochemistry]: "It is well established that p53 contacts DNA in a sequence-dependent manner in order to transactivate its myriad target genes. Yet little is known about how p53 interacts with its binding site/response element (RE) within such genes in vivo in the context of nucleosomal DNA. In this study we demonstrate that both distal (5′) and proximal (3′) p53 REs within the promoter of the p21 gene in unstressed HCT116 colon carcinoma cells are localized within a region of relatively high nucleosome occupancy. In the absence of cellular stress, p53 is prebound to both p21 REs within nucleosomal DNA in these cells. Treatment of cells with the DNA-damaging drug doxorubicin or the p53 stabilizing agent Nutlin-3, however, is accompanied by p53-dependent subsequent loss of nucleosomes associated with such p53 REs. We show that in vitro p53 can bind to mononucleosomal DNA containing the distal p21 RE, provided the binding site is not close to the diad center of the nucleosome. In line with this, our data indicate that the p53 distal RE within the p21 gene is located close to the end of the nucleosome. Thus, low- and high-resolution mapping of nucleosome boundaries around p53 REs within the p21 promoter have provided insight into the mechanism of p53 binding to its sites in cells and the consequent changes in nucleosome occupancy at such sites."

Single-tube linear DNA amplification (LinDA) for robust ChIP-seq

Single-tube linear DNA amplification (LinDA) for robust ChIP-seq: "

Single-tube linear DNA amplification (LinDA) for robust ChIP-seq

Nature Methods 8, 565 (2011).
doi:10.1038/nmeth.1626

Authors: Pattabhiraman Shankaranarayanan, Marco-Antonio Mendoza-Parra, Mannu Walia, Li Wang, Ning Li, Luisa M Trindade & Hinrich Gronemeyer

Genome-wide profiling of transcription factors based on massive parallel sequencing of immunoprecipitated chromatin (ChIP-seq) requires nanogram amounts of DNA. Here we describe a high-fidelity, single-tube linear DNA amplification method (LinDA) for ChIP-seq and reChIP-seq with picogram DNA amounts obtained from a few thousand cells. This amplification technology will facilitate global analyses of transcription-factor binding and chromatin with very small cell populations, such as stem or cancer-initiating cells.

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A gene regulatory network controlling the embryonic specification ofendoderm

A gene regulatory network controlling the embryonic specification of<br>endoderm: "

A gene regulatory network controlling the embryonic specification of
endoderm

Nature 474, 7353 (2011). doi:10.1038/nature10100

Authors: Isabelle S. Peter & Eric H. Davidson

Specification of endoderm is the prerequisite for gut formation in the
embryogenesis of bilaterian organisms. Modern lineage labelling studies have shown that in the sea urchin embryo model system, descendants
of the veg1 and veg2 cell lineages produce the endoderm, and that the veg2
lineage also gives rise to mesodermal cell types. It is known that Wnt/β-catenin
signalling is required for endoderm specification and
Delta/Notch signalling is required for mesoderm specification.
Some direct cis-regulatory targets of these signals have been found and various phenomenological patterns of gene expression have
been observed in the pre-gastrular endomesoderm. However, no comprehensive,
causal explanation of endoderm specification has been conceived for sea urchins,
nor for any other deuterostome. Here we propose a model, on the basis of the
underlying genomic control system, that provides such an explanation, built
at several levels of biological organization. The hardwired core of the control
system consists of the cis-regulatory apparatus of endodermal regulatory
genes, which determine the relationship between the inputs to which these
genes are exposed and their outputs. The architecture of the network circuitry
controlling the dynamic process of endoderm specification then explains, at
the system level, a sequence of developmental logic operations, which generate
the biological process. The control system initiates non-interacting endodermal
and mesodermal gene regulatory networks in veg2-derived cells and extinguishes
the endodermal gene regulatory network in mesodermal precursors. It also generates
a cross-regulatory network that specifies future anterior endoderm in veg2
descendants and institutes a distinct network specifying posterior endoderm
in veg1-derived cells. The network model provides an explanatory framework
that relates endoderm specification to the genomic regulatory code.

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Determinants of nucleosome organization in primary human cells

Determinants of nucleosome organization in primary human cells: "

Determinants of nucleosome organization in primary human cells

Nature 474, 7352 (2011). doi:10.1038/nature10002

Authors: Anton Valouev, Steven M. Johnson, Scott D. Boyd, Cheryl L. Smith, Andrew Z. Fire & Arend Sidow

Nucleosomes are the basic packaging units of chromatin, modulating accessibility of regulatory proteins to DNA and thus influencing eukaryotic gene regulation. Elaborate chromatin remodelling mechanisms have evolved that govern nucleosome organization at promoters, regulatory elements, and other functional regions in the genome. Analyses of chromatin landscape have uncovered a variety of mechanisms, including DNA sequence preferences, that can influence nucleosome positions. To identify major determinants of nucleosome organization in the human genome, we used deep sequencing to map nucleosome positions in three primary human cell types and in vitro. A majority of the genome showed substantial flexibility of nucleosome positions, whereas a small fraction showed reproducibly positioned nucleosomes. Certain sites that position in vitro can anchor the formation of nucleosomal arrays that have cell type-specific spacing in vivo. Our results unveil an interplay of sequence-based nucleosome preferences and non-nucleosomal factors in determining nucleosome organization within mammalian cells.

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Non-adaptive origins of interactome complexity

Non-adaptive origins of interactome complexity: "

Non-adaptive origins of interactome complexity

Nature 474, 7352 (2011). doi:10.1038/nature09992

Authors: Ariel Fernández & Michael Lynch

The boundaries between prokaryotes, unicellular eukaryotes and multicellular eukaryotes are accompanied by orders-of-magnitude reductions in effective population size, with concurrent amplifications of the effects of random genetic drift and mutation. The resultant decline in the efficiency of selection seems to be sufficient to influence a wide range of attributes at the genomic level in a non-adaptive manner. A key remaining question concerns the extent to which variation in the power of random genetic drift is capable of influencing phylogenetic diversity at the subcellular and cellular levels. Should this be the case, population size would have to be considered as a potential determinant of the mechanistic pathways underlying long-term phenotypic evolution. Here we demonstrate a phylogenetically broad inverse relation between the power of drift and the structural integrity of protein subunits. This leads to the hypothesis that the accumulation of mildly deleterious mutations in populations of small size induces secondary selection for protein–protein interactions that stabilize key gene functions. By this means, the complex protein architectures and interactions essential to the genesis of phenotypic diversity may initially emerge by non-adaptive mechanisms.

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