Ubiquitous nucleosome unwrapping in the yeast genome

Nucleosome core particle is a dynamic structure -- DNA may transiently peel off the histone octamer surface due to thermal fluctuations or the action of chromatin remodeling enzymes. Partial DNA unwrapping enables easier access of DNA-binding factors to their target sites and thus may provide a dominant pathway for effecting rapid and robust access to DNA packaged into chromatin. Indeed, a recent high-resolution map of distances between neighboring nucleosome dyads in \emph{S.cerevisiae} shows that at least 38.7\% of all nucleosomes are partially unwrapped. The extent of unwrapping follows a stereotypical pattern in the vicinity of genes, reminiscent of the canonical pattern of nucleosome occupancy in which nucleosomes are depleted over promoters and well-positioned over coding regions. To explain these observations, we developed a biophysical model which employs a 10-11 base pair periodic nucleosome energy profile. The profile, based on the pattern of histone-DNA contacts in nucleosome crystal structures and the idea of linker length discretization, accounts for both nucleosome unwrapping and higher-order chromatin structure. Our model reproduces the observed genome-wide distribution of inter-dyad distances, and accounts for patterns of nucleosome occupancy and unwrapping around coding regions. At the same time, our approach explains \emph{in vitro} measurements of accessibility of nucleosome-covered binding sites, and of nucleosome-induced cooperativity between DNA-binding factors. We are able to rule out several alternative scenarios of nucleosome unwrapping as inconsistent with the genomic data.Comment: 49 pages; 15 figure

DNA-encoded nucleosome occupancy is associated with transcription levels in the human malaria parasite Plasmodium falciparum.

BackgroundIn eukaryotic organisms, packaging of DNA into nucleosomes controls gene expression by regulating access of the promoter to transcription factors. The human malaria parasite Plasmodium falciparum encodes relatively few transcription factors, while extensive nucleosome remodeling occurs during its replicative cycle in red blood cells. These observations point towards an important role of the nucleosome landscape in regulating gene expression. However, the relation between nucleosome positioning and transcriptional activity has thus far not been explored in detail in the parasite.ResultsHere, we analyzed nucleosome positioning in the asexual and sexual stages of the parasite's erythrocytic cycle using chromatin immunoprecipitation of MNase-digested chromatin, followed by next-generation sequencing. We observed a relatively open chromatin structure at the trophozoite and gametocyte stages, consistent with high levels of transcriptional activity in these stages. Nucleosome occupancy of genes and promoter regions were subsequently compared to steady-state mRNA expression levels. Transcript abundance showed a strong inverse correlation with nucleosome occupancy levels in promoter regions. In addition, AT-repeat sequences were strongly unfavorable for nucleosome binding in P. falciparum, and were overrepresented in promoters of highly expressed genes.ConclusionsThe connection between chromatin structure and gene expression in P. falciparum shares similarities with other eukaryotes. However, the remarkable nucleosome dynamics during the erythrocytic stages and the absence of a large variety of transcription factors may indicate that nucleosome binding and remodeling are critical regulators of transcript levels. Moreover, the strong dependency between chromatin structure and DNA sequence suggests that the P. falciparum genome may have been shaped by nucleosome binding preferences. Nucleosome remodeling mechanisms in this deadly parasite could thus provide potent novel anti-malarial targets

Theory of Nucleosome Corkscrew Sliding in the Presence of Synthetic DNA Ligands

Histone octamers show a heat-induced mobility along DNA. Recent theoretical studies have established two mechanisms that are qualitatively and quantitatively compatible with in vitro experiments on nucleosome sliding: Octamer repositiong through one-basepair twist defects and through ten-basepair bulge defects. A recent experiment demonstrated that the repositioning is strongly suppressed in the presence of minor-groove binding DNA ligands. In the present study we give a quantitative theory for nucleosome repositioning in the presence of such ligands. We show that the experimentally observed octamer mobilities are consistent with the picture of bound ligands blocking the passage of twist defects through the nucleosome. This strongly supports the model of twist defects inducing a corkscrew motion of the nucleosome as the underlying mechanism of nucleosome sliding. We provide a theoretical estimate of the nucleosomal mobility without adjustable parameters, as a function of ligand concentration, binding affinity, binding site orientiation, temperature and DNA anisotropy. Having this mobility at hand we speculate about the interaction between a nucleosome and a transcribing RNA polymerase and suggest a novel mechanism that might account for polymerase induced nucleosome repositioning.Comment: 23 pages, 4 figures, submitted to J. Mol. Bio

Quantitative test of the barrier nucleosome model for statistical positioning of nucleosomes up- and downstream of transcription start sites

The positions of nucleosomes in eukaryotic genomes determine which parts of the DNA sequence are readily accessible for regulatory proteins and which are not. Genome-wide maps of nucleosome positions have revealed a salient pattern around transcription start sites, involving a nucleosome-free region (NFR) flanked by a pronounced periodic pattern in the average nucleosome density. While the periodic pattern clearly reflects well-positioned nucleosomes, the positioning mechanism is less clear. A recent experimental study by Mavrich et al. argued that the pattern observed in S. cerevisiae is qualitatively consistent with a `barrier nucleosome model', in which the oscillatory pattern is created by the statistical positioning mechanism of Kornberg and Stryer. On the other hand, there is clear evidence for intrinsic sequence preferences of nucleosomes, and it is unclear to what extent these sequence preferences affect the observed pattern. To test the barrier nucleosome model, we quantitatively analyze yeast nucleosome positioning data both up- and downstream from NFRs. Our analysis is based on the Tonks model of statistical physics which quantifies the interplay between the excluded-volume interaction of nucleosomes and their positional entropy. We find that although the typical patterns on the two sides of the NFR are different, they are both quantitatively described by the same physical model, with the same parameters, but different boundary conditions. The inferred boundary conditions suggest that the first nucleosome downstream from the NFR (the +1 nucleosome) is typically directly positioned while the first nucleosome upstream is statistically positioned via a nucleosome-repelling DNA region. These boundary conditions, which can be locally encoded into the genome sequence, significantly shape the statistical distribution of nucleosomes over a range of up to ~1000 bp to each side.Comment: includes supporting materia

The chromatin remodeller ACF acts as a dimeric motor to space nucleosomes.

Evenly spaced nucleosomes directly correlate with condensed chromatin and gene silencing. The ATP-dependent chromatin assembly factor (ACF) forms such structures in vitro and is required for silencing in vivo. ACF generates and maintains nucleosome spacing by constantly moving a nucleosome towards the longer flanking DNA faster than the shorter flanking DNA. How the enzyme rapidly moves back and forth between both sides of a nucleosome to accomplish bidirectional movement is unknown. Here we show that nucleosome movement depends cooperatively on two ACF molecules, indicating that ACF functions as a dimer of ATPases. Further, the nucleotide state determines whether the dimer closely engages one or both sides of the nucleosome. Three-dimensional reconstruction by single-particle electron microscopy of the ATPase-nucleosome complex in an activated ATP state reveals a dimer architecture in which the two ATPases face each other. Our results indicate a model in which the two ATPases work in a coordinated manner, taking turns to engage either side of a nucleosome, thereby allowing processive bidirectional movement. This novel dimeric motor mechanism differs from that of dimeric motors such as kinesin and dimeric helicases that processively translocate unidirectionally and reflects the unique challenges faced by motors that move nucleosomes

PuFFIN--a parameter-free method to build nucleosome maps from paired-end reads.

BackgroundWe introduce a novel method, called PuFFIN, that takes advantage of paired-end short reads to build genome-wide nucleosome maps with larger numbers of detected nucleosomes and higher accuracy than existing tools. In contrast to other approaches that require users to optimize several parameters according to their data (e.g., the maximum allowed nucleosome overlap or legal ranges for the fragment sizes) our algorithm can accurately determine a genome-wide set of non-overlapping nucleosomes without any user-defined parameter. This feature makes PuFFIN significantly easier to use and prevents users from choosing the "wrong" parameters and obtain sub-optimal nucleosome maps.ResultsPuFFIN builds genome-wide nucleosome maps using a multi-scale (or multi-resolution) approach. Our algorithm relies on a set of nucleosome "landscape" functions at different resolution levels: each function represents the likelihood of each genomic location to be occupied by a nucleosome for a particular value of the smoothing parameter. After a set of candidate nucleosomes is computed for each function, PuFFIN produces a consensus set that satisfies non-overlapping constraints and maximizes the number of nucleosomes.ConclusionsWe report comprehensive experimental results that compares PuFFIN with recently published tools (NOrMAL, TEMPLATE FILTERING, and NucPosSimulator) on several synthetic datasets as well as real data for S. cerevisiae and P. falciparum. Experimental results show that our approach produces more accurate nucleosome maps with a higher number of non-overlapping nucleosomes than other tools