Nicolas Clauvelin

Internucleosomal Interactions Mediated by Histone Tails Allow Distant Communication in Chromatin

Background: Gene expression is regulated by DNA elements that often lie far apart along genomic sequences. Results: Novel computations and experiments provide new structural insights into long-range communication on chromatin. Conclusion: Efficient long-range association of transcriptional elements requires intact tails on the core histones. Significance: The understanding of action-at-a-distance in three dimensions helps to connect nucleosome structure/positioning to chromatin dynamics and gene regulation. Action across long distances on chromatin is a hallmark of eukaryotic transcriptional regulation. Although chromatin structure per se can support long-range interactions, the mechanisms of efficient communication between widely spaced DNA modules in chromatin remain a mystery. The molecular simulations described herein suggest that transient binary internucleosomal interactions can mediate distant communication in chromatin. Electrostatic interactions between the N-terminal tails of the core histones and DNA enhance the computed probability of juxtaposition of sites that lie far apart along the DNA sequence. Experimental analysis of the rates of communication in chromatin constructs confirms that long-distance communication occurs efficiently and independently of distance on tail-containing, but not on tailless, chromatin. Taken together, our data suggest that internucleosomal interactions involving the histone tails are essential for highly efficient, long-range communication between regulatory elements and their targets in eukaryotic genomes.

Insights into Gene Expression and Packaging from Computer Simulations

Within the nucleus of each cell lies DNA—an unfathomably long, twisted, and intricately coiled molecule—segments of which make up the genes that provide the instructions that a cell needs to operate. As we near the 60th anniversary of the discovery of the DNA double helix, crucial questions remain about how the physical arrangement of the DNA in cells affects how genes work. For example, how a cell stores the genetic information inside the nucleus is complicated by the necessity of maintaining accessibility to DNA for genetic processing. In order to gain insight into the roles played by various proteins in reading and compacting the genome, we have developed new methodologies to simulate the dynamic, three-dimensional structures of long, fluctuating, protein-decorated strands of DNA. Our a priori approach to the problem allows us to determine the effects of individual proteins and their chemical modifications on overall DNA structure and function. Here, we present our recent treatment of the communication between regulatory proteins attached to precisely constructed stretches of chromatin. Our simulations account for the enhancement in communication detected experimentally on chromatin compared to protein-free DNA of the same chain length, as well as the critical roles played by the cationic ‘tails’ of the histone proteins in this signaling. The states of chromatin captured in the simulations offer new insights into the ways that the DNA, histones, and regulatory proteins contribute to long-range communication along the genome.

Contributions of Sequence to the Higher-Order Structures of DNA

One of the critical unanswered questions in genome biophysics is how the primary sequence of DNA bases influences the global properties of very-long-chain molecules. The local sequence-dependent features of DNA found in high-resolution structures introduce irregularities in the disposition of adjacent residues that facilitate the specific binding of proteins and modulate the global folding and interactions of double helices with hundreds of basepairs. These features also determine the positions of nucleosomes on DNA and the lengths of the interspersed DNA linkers. Like the patterns of basepair association within DNA, the arrangements of nucleosomes in chromatin modulate the properties of longer polymers. The intrachromosomal loops detected in genomic studies contain hundreds of nucleosomes, and given that the simulated configurations of chromatin depend on the lengths of linker DNA, the formation of these loops may reflect sequence-dependent information encoded within the positioning of the nucleosomes. With knowledge of the positions of nucleosomes on a given genome, methods are now at hand to estimate the looping propensities of chromatin in terms of the spacing of nucleosomes and to make a direct connection between the DNA base sequence and larger-scale chromatin folding.

Weak operator binding enhances simulated lac repressor‐mediated DNA looping

The 50th anniversary of Biopolymers coincides closely with the like celebration of the discovery of the Escherichia coli (lac) lactose operon, a classic genetic system long used to illustrate the influence of biomolecular structure on function. The looping of DNA induced by the binding of the Lac repressor protein to sequentially distant operator sites on DNA continues to serve as a paradigm for understanding long‐range genomic communication. Advances in analyses of DNA structures and in incorporation of proteins in computer simulations of DNA looping allow us to address long‐standing questions about the role of protein‐mediated DNA loop formation in transcriptional control. Here we report insights gained from studies of the sequence‐dependent contributions of the natural lac operators to Lac repressor‐mediated DNA looping. Novel superposition of the ensembles of protein‐bound operator structures derived from NMR measurements reveals variations in DNA folding missed in conventional structural alignments. The changes in folding affect the predicted ease with which the repressor induces loop formation and the ways that DNA closes between the protein headpieces. The peeling of the auxiliary operators away from the repressor enhances the formation of loops with the 92‐bp wildtype spacing and hints of a structural reason behind their weak binding.

Nucleosome-free DNA regions differentially affect distant communication in chromatin

Abstract Communication between distantly spaced genomic regions is one of the key features of gene regulation in eukaryotes. Chromatin per se can stimulate efficient enhancer-promoter communication (EPC); however, the role of chromatin structure and dynamics in this process remains poorly understood. Here we show that nucleosome spacing and the presence of nucleosome-free DNA regions can modulate chromatin structure/dynamics and, in turn, affect the rate of EPC in vitro and in silico. Increasing the length of internucleosomal linker DNA from 25 to 60 bp results in more efficient EPC. The presence of longer nucleosome-free DNA regions can positively or negatively affect the rate of EPC, depending upon the length and location of the DNA region within the chromatin fiber. Thus the presence of histone-free DNA regions can differentially affect the efficiency of EPC, suggesting that gene regulation over a distance could be modulated by changes in the length of internucleosomal DNA spacers.

What Controls DNA Looping

The looping of DNA provides a means of communication between sequentially distant genomic sites that operate in tandem to express, copy, and repair the information encoded in the DNA base sequence. The short loops implicated in the expression of bacterial genes suggest that molecular factors other than the naturally stiff double helix are involved in bringing the interacting sites into close spatial proximity. New computational techniques that take direct account of the three-dimensional structures and fluctuations of protein and DNA allow us to examine the likely means of enhancing such communication. Here, we describe the application of these approaches to the looping of a 92 base-pair DNA segment between the headpieces of the tetrameric Escherichia coli Lac repressor protein. The distortions of the double helix induced by a second protein--the nonspecific nucleoid protein HU--increase the computed likelihood of looping by several orders of magnitude over that of DNA alone. Large-scale deformations of the repressor, sequence-dependent features in the DNA loop, and deformability of the DNA operators also enhance looping, although to lesser degrees. The correspondence between the predicted looping propensities and the ease of looping derived from gene-expression and single-molecule measurements lends credence to the derived structural picture.

Parallels between DNA and collagen – comparing elastic models of the double and triple helix

Multi-stranded helices are widespread in nature. The interplay of polymeric properties with biological function is seldom discussed. This study probes analogies between structural and mechanical properties of collagen and DNA. We modeled collagen with Eulerian rotational and translational parameters of adjacent rungs in the triple-helix ladder and developed statistical potentials by extracting the dispersion of the parameters from a database of atomic-resolution structures. The resulting elastic model provides a common quantitative way to describe collagen deformations upon interacting with integrins or matrix metalloproteinase and DNA deformations upon protein binding. On a larger scale, deformations in Type I collagen vary with a periodicity consistent with the D-periodic banding of higher-order fibers assemblies. This indicates that morphologies of natural higher-order collagen packing might be rooted in the characteristic deformation patterns.

59 The synergy between DNA and nucleosomes in chromatin

Although it is now common to look at the information carried by the genome as a linear sequence of nucleotides, such representation does not say much about the organization of the genetic information within the cell nucleus. How the genome accommodates the tight packing needed to fit in the cell nucleus and at the same time maintains the accessibility necessary for specific expression is one of the open questions in modern biology. In eukaryotic cells, DNA is wrapped and packaged into chromatin through the binding of histones assembled into nucleosomes. In addition to bundling DNA, the nucleosomes also facilitate communication between distant genomic sites, such as enhancers and promoters found at the ends of protein-mediated loops. In order to understand the physical and chemical basis of such processes, we have begun to investigate chromatin organization and looping. We have developed a mesoscale model of chromatin at a resolution of a single base pair and used Monte Carlo numerical strategies to understand how the presence of nucleosomes on DNA can influence and possibly control chromatin looping. We have validated this model by successfully reproducing experimental measurements of gene expression on nucleosomal arrays (Kulaeva et al., 2012). Our results show a wide variety of chromatin organization depending on the way nucleosomes are positioned on DNA (i.e. on the spacing between successive nucleosomes as illustrated in the pictures below) and also on chemical details at the histone level, such as modifications of the N-terminal tails. This diversity in chromatin organization, which extends beyond the conventional solenoid and zigzag models, comes along with very different physical and mechanical properties and looping propensities. Furthermore, our simulations reveal some surprising properties of chromatin: for example, we found that tightly packed chromatin fragments are highly flexible, a remarkable feature for a material that needs to fit inside a cell nucleus. In conclusion, our work uncovers parts of a rich and dynamic picture of chromatin where the DNA sequence influences the positioning of nucleosomes, which in turn shape the genomic material and have an impact on the accessibility and expression of the genetic message.

Dynamics of the Nucleosome Core Particle Revealed from a New Database of High-Resolution X-Ray Crystallographic and Simulated Structures

The nucleosome core particle is a highly conserved structure which can play diverse roles depending on the organism, cell, or part of chromatin in which it resides. The Protein Data Bank currently contains approximately 90 nucleosome core particle structures, most of which were determined in the last five years. The recent emergence of the field of epigenetics, and the increase in data available from experiments, warrants a need to develop new approaches to compare features of interest across multiple structures.The growing ensemble of structural data garnered from in vitro and in silico experiments provides a unique platform to study the mechanochemical properties of the nucleosome. We have developed a database and new computational tools to allow researchers to analyze and compare the nucleosome core particle structures deposited in the Protein Data Bank. The features of the DNA-protein assembly can be examined in novel coordinate frames placed on the structure, allowing researchers to obtain a better understanding of the organization and subtleties of the macromolecular complexes. This comparison allows one to examine the ‘motion’ of specific residues of interest, including specific sites of post-translational histone modification. The database also includes DNA-histone contact points, DNA conformational parameters, and information about protein features such as the secondary structure in the globular histone core and the ‘motion’ of the histone tails. Along with these features, we also characterize the dynamics of the global structure of the nucleosome core particle, including changes in the superhelical path of the DNA, rearrangements of the histone tetramers, and nucleosome stacking inside crystals. This information allows us to understand and model the critical role of mono-nucleosome structural propensity in processes such as carcinogenic modifications of the DNA, and nucleosome remodeling.

The Synergy Between DNA and Nucleosomes in Chromatin

How the genome accommodates the tight packing needed to fit in the cell nucleus and at the same time maintains the accessibility necessary for specific expression is one of the open questions in modern biology. In eukaryotic cells DNA is wrapped and packaged into chromatin through the binding of histones and formation of nucleosomes. In addition to bundling DNA, the nucleosomes also facilitate communication between distant genomic sites, such as enhancers and promoters found at the ends of protein-mediated loops. In order to understand such processes, we have begun to investigate chromatin organization and looping. We have developed a mesoscale model of chromatin at a resolution of a single base pair and used Monte Carlo numerical strategies to unravel how the presence of nucleosomes on DNA can influence and possibly control chromatin looping. We have validated this model by successfully reproducing experimental measurements of gene expression on nucleosomal arrays. Our results show a wide variety of chromatin organization depending on the way nucleosomes are positioned on DNA and also on chemical details at the histone level, such as modifications of the N-terminal tails. This diversity in chromatin organization, which extends beyond the conventional solenoid and zigzag models, comes along with very different physical and mechanical properties and looping propensities. Furthermore, our simulations reveal some surprising properties of chromatin: for example, we found that tightly packed chromatin fragments are the most flexible, a remarkable feature for a material that needs to fit inside a cell nucleus. In conclusion, our work uncovers parts of a rich and dynamic picture of chromatin where the DNA sequence influences the positioning of nucleosomes, which in turn shapes the genomic material and has an impact on the accessibility and expression of the genetic message.