Bulletin of the American Physical Society
APS March Meeting 2015
Volume 60, Number 1
Monday–Friday, March 2–6, 2015; San Antonio, Texas
Session Z47: Physics of Genome Organization |
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Sponsoring Units: DBIO Chair: Alexandre Morozov, Rutgers University Room: 217B |
Friday, March 6, 2015 11:15AM - 11:27AM |
Z47.00001: The Stochastic Signature of Mixed Promoter States Leonardo Sepulveda, Ido Golding Gene promoters typically contain multiple binding sites for transcription factors. This allows for distinct transcription-factor binding configurations, each characterized by a different transcriptional activity of the regulated gene. However, at a given transcription-factor concentration, the promoter is not expected to exhibit a single configuration, but, instead, a ``mixed state'' with fractional probabilities for the different configurations. What is the nature of these mixed promoter states at the single-cell level? We investigate this question by measuring simultaneously, in individual cells, the concentration of the CI transcription factor and the transcriptional output of the regulated promoter, P$_{RM}$, in \textit{E. coli}. We use the mRNA copy-number statistics to reconstruct the stochastic kinetics of the different promoter configurations and calculate their probabilities at each CI concentration. We find that the mRNA distribution for cells in a mixed state can be described as a convolution of the pure-state distributions, indicating rapid switching between the pure promoter states. Thus, mixed promoter states do not result in different cell populations but instead appear as a new, well-defined promoter activity. [Preview Abstract] |
Friday, March 6, 2015 11:27AM - 11:39AM |
Z47.00002: Direct Quantification of Transcriptional Regulation at an Endogenous Gene Locus Heng Xu, Anna Sokac, Ido Golding The stochastic kinetics of gene activity in individual cells has been well characterized, but how this kinetics is modulated by the transcription factors that regulate expression remains largely unknown. We address this question using the Bicoid (Bcd) transcription factor and \textit{hunchback} (\textit{hb}) gene in early \textit{Drosophila} embryos. We measure, simultaneously, the number of nascent \textit{hb} mRNAs, nuclear Bcd concentration, and number of bound Bcd proteins, at individual gene loci. Using stochastic theoretical analysis, we find that Bcd modulates the probability of \textit{hb} switching to an active transcriptional state, while not affecting the probabilities of transcription initiation or gene inactivation. Gene activation is achieved through the cooperative binding of ~6 Bcd copies. Our data also reveals additional Bcd binding states of unknown function. In contrast to Bcd, binding of the Hunchback transcription factor represses \textit{hb} transcription. Our approach can be used to elucidate the combinatorial activity of multiple transcription factors without the need for genetic perturbation. [Preview Abstract] |
Friday, March 6, 2015 11:39AM - 11:51AM |
Z47.00003: Fast Chromatin Assembly facilitated by Nucleosome Breathing and Replication-Guided Packing Johannes Nuebler, Brendan Osberg, Philipp Korber, Ulrich Gerland The condensation of eukaryotic DNA into chromatin entails the formation of nucleosome arrays with high density at species-dependent nucleosome spacing. These arrays are frequently destroyed by transcription and replication, such that reassembly is required. Due to a well-known jamming effect in the random adsorption of mutually exclusive objects (aka the ``car parking problem''), the question was raised how in vivo nucleosome densities, and patterns, can be reached in the biologically relevant timescale of minutes [1]. We show that the ``softness'' of nucleosomes alleviates this kinetic challenge [2]. Nucleosome softness arises due to transient DNA unwrapping (breathing) and stepwise nucleosome assembly. From a physics perspective, the ``soft car parking problem'' differs fundamentally from its hard counterpart by exhibiting non-monotonic density and rapid equilibration. We also discuss scenarios how the progression of the replication fork can promote rapid reassembly in its wake. For example, tight packing arises naturally if the fork progresses slowly compared to the reassembly rate.\\[4pt] [1] R. Padinhateeri, J.F. Marko, PNAS 108, 7799 (2011).\newline [2] B. Osberg, J. Nuebler, P. Korber, U. Gerland, Nucl. Acids Res., in press. [Preview Abstract] |
Friday, March 6, 2015 11:51AM - 12:03PM |
Z47.00004: Topology, structure and energy landscape of human chromosomes Bin Zhang, Peter Wolynes The genomes' three-dimensional (3D) organization is crucial in regulating many biological processes, including gene regulation, DNA replication, and cell differentiation. We develop a statistically rigorous approach based on maximum entropy principle to determine a least-biased potential energy landscape that reproduces experimentally determined Hi-C contact frequency between genome pairs. The resulting energy landscape supports a knotless chromosome conformation, which has been highly anticipated since complex knotted conformations prohibit the access of gene information for transcription and hinder DNA replication. We further show that the topologically associating domain signal alone also enforces a chromosome structure free of knots. Our results highlight the importance of local interactions in determining the global topology of the chromosome structure. Finally, the derived landscapes for multiple chromosomes support the formation of territories that have long been observed in microscopy experiments. Together with Hi-C experiments, our approach provides a coherent picture of the 3D architecture of the genomes that is consistent with many the available experimental data. [Preview Abstract] |
Friday, March 6, 2015 12:03PM - 12:15PM |
Z47.00005: Expediting the Sequencing Process: Using Soft Lithography to Fragment Aligned DNA Molecules Meena Jagadeesan, Adina Singer, NaHyun Cho, Ke Zhu, Julia Budassi, Jonathan Sokolov Current sequencing technologies output short read lengths on the order of 10 kb, requiring DNA to be fragmented into small pieces, analyzed separately and recombined to obtain the full sequence. The accuracy and cost efficiency of sequencing has been limited by the computer algorithms required to assemble the random subsequesnces. The fragmentation method in this study can significantly simplify the assembly process with a novel lithographic cutting procedure that retains the position and order of DNA fragments. DNA was stretched linearly on a polymer coated silicon wafer. A polydimethylsiloxane (PDMS) lithographic stamp was coated in DNase I (DNA cutting enzyme) solution. The stamp was placed in contact with the surface aligned DNA, producing 3.5 micron (10 kbp) DNA fragments. Fluorescence imaging of dye labeled DNA was used to monitor the cutting effectiveness. The improved enzyme application procedure presented in this study enabled uniform, ordered cutting of the surface aligned DNA over approximately 80{\%} of the 2 cm x 3 cm samples. Means to extract the cut DNA from the polymer surface were also explored. The effectiveness of solution methods, electric field desorption, and microfluidics will be discussed. [Preview Abstract] |
Friday, March 6, 2015 12:15PM - 12:27PM |
Z47.00006: New insights into nucleosome positioning Razvan Chereji, Josefina Ocampo, Tara Burke, David Clark A human body contains enough DNA to circle the Earth's Equator more than 2.5 million times. Nevertheless, the entire genetic material is packed inside the tiny nuclei of our cells. The basic units of DNA packaging are called nucleosomes. Their locations on the chromosomes play an essential role in gene regulation. We study nucleosome positioning in yeast, fly and mouse, both in vivo and in vitro, and build biophysical models in order to explain the genome-wide nucleosome organization. We show that DNA sequence is not the major cause of the phased arrays of nucleosomes observed in vivo near the transcription start sites. We discuss simple models which can account for the formation of nucleosome depleted regions and nucleosome phasing at the gene promoters. We analyze the effects of different factors which influence the chromatin organization in living cells: existence of potential barriers and wells, sequence-dependent nucleosome affinity, nucleosome unwrapping, competition between different DNA-binding proteins, action of ATP-dependent remodelers, among others. [Preview Abstract] |
Friday, March 6, 2015 12:27PM - 1:03PM |
Z47.00007: Statistical mechanics of nucleosome assembly and chromatin packaging Invited Speaker: Alexandre Morozov Eukaryotic genomes are organized into arrays of nucleosomes. Each fully wrapped nucleosome consists of 147 base pairs of genomic DNA bent around a histone octamer core. The resulting complex of DNA with histones forms a multi-scale structure called chromatin. At the most fundamental level of chromatin organization, arrays of nucleosomes form 10-nm fibers which resemble beads on a string; these in turn fold into higher-order structures. Depending on the organism and cell type, 75-90\% of genomic DNA is packaged into nucleosomes. The question of how cellular functions are carried out on this chromatin template is one of the outstanding puzzles in biology. Nucleosomal DNA may transiently peel off the histone octamer surface due to thermal fluctuations or interactions with chromatin remodelers. Thus neighboring nucleosomes can invade each other's territories through DNA unwrapping and translocation, or through initial assembly in partially wrapped states. A recent high-resolution map of distances between neighboring nucleosomes in baker's yeast [1] has revealed that at least 25\% of all nucleosomes overlap with DNA territories of their neighbors. To explain this observation, we have developed a statistical mechanics model of nucleosome assembly and unwrapping [2]. Our model is in agreement with genome-wide nucleosome positioning data and \textit{in vitro} measurements of accessibility of nucleosome-covered target sites. Furthermore, it explains nucleosome-induced cooperativity between DNA-binding factors. The observed extent of nucleosome crowding in the yeast genome strongly suggests that its treatment should be included in all future models of chromatin structure and energetics.\\[4pt] [1] Brogaard et al. A map of nucleosome positions in yeast at base-pair resolution. Nature (2012), 486:496-501. \newline [2] Chereji and Morozov. Ubiquitous nucleosome crowding in the yeast genome. Proc Natl Acad Sci USA (2014), 111:5236-41. [Preview Abstract] |
Friday, March 6, 2015 1:03PM - 1:15PM |
Z47.00008: Exploring telomeric DNA-protein-DNA interactions under nanoconfinement Maedeh Roushan, Parminder kaur, Jianguo Lin, Hong Wang, Robert Riehn Genomes are organized through DNA binding proteins. In particular, telomeres are organized into $\sim$ 10 kbp loops by a multi-protein complex. For understanding how proteins interact with DNA we have investigated the effect of different DNA-binding proteins on DNA configuration by injecting different proteins inside a nanofabricated channel system. DNA molecules stretch in nanochannels with a channel cross-section roughly about 100x100 nm2, so allowing analysis by observation of a fluorescent dye. The length and configuration of DNA can be directly observed, as well as the binding location of proteins. Here we show the binding patterns and molecular action of a set of telomere-associated proteins, namely TFR1, TRF2, RAP1, SA1, as well the model protein T4 DNA ligase. In particular, we demonstrate formation of stable loops, sliding, and general DNA condensation. [Preview Abstract] |
Friday, March 6, 2015 1:15PM - 1:27PM |
Z47.00009: Computational modeling of autocatalytic heteropolymer replication Hemachander Subramanian, Robert Gatenby We computationally study replication of a hypothetical autocatalytic heteropolymer using Kinetic Monte Carlo. When cooperativity is included in the model, we observe better replication characteristics and higher fitness in strands with certain symmetries broken. We correlate this symmetry-breaking with the observed broken mirror symmetry of extant heteropolymers. [Preview Abstract] |
Friday, March 6, 2015 1:27PM - 1:39PM |
Z47.00010: A unified description of biological effects caused by radiation exposure: Whack-a-mole (WAM) model Takahiro Wada, Yuichiro Manabe, Issei Nakamura, Masako Bando We present our novel rate equations to study DNA mutation in cells caused by artificial radiation exposure, accounting for the DNA damage and repair simultaneously. In our theory, the dependence of mutation frequencies on the dose rate is critically important to predict both the time course and the stationary effect of the DNA mutation in cell cycles. Experimentally, irradiation at high dose rates causes linear increases in the mutation frequency with total dose, whereas the saturation of the mutation frequency is observed at low dose rates. We demonstrate that this fact arises from counteracting effects among the DNA damage and mutation, the DNA repair, and the proliferation and apoptosis of cells. Our theory thus captures observed quantities at both high and low dose rates, marking a substantial difference from conventional theories based only on the total dose. Importantly, we have derived a scaling function from our rate equations that predicts a universal feature in the mutation frequency of living organisms. In this study, we have analyzed the experimental data of five species; mouse, drosophila, chrysanthemum, maize, and tradescantia. Despite the difference between animal and plant, all these data reasonably fall on a single line for our scaling function. [Preview Abstract] |
Friday, March 6, 2015 1:39PM - 2:15PM |
Z47.00011: Extracting free energies of interaction from chromosome conformation capture data Invited Speaker: Eldon Emberly A variety of DNA binding proteins are involved in regulating and shaping the packing of chromatin. They aid the formation of loops in the DNA that function to isolate different structural domains. A recent experimental technique, Hi-C, provides a method for determining the frequency of such looping between all distant parts of the genome. Given that the binding locations of many chromatin associated proteins have also been measured, it is possible to make estimates for their influence on the long-range interactions as measured by Hi-C. However, a challenge in this analysis is the predominance of non-specific contacts that has made making quantitative estimates for the strengths of interactions between chromatin factors difficult. In this talk I will show that transforming the Hi-C contact frequencies into free energies of interaction gives a natural method for separating out the distance dependent non-specific interactions. In particular, using Principal Component Analysis (PCA) on the transformed free energy matrix can identify the dominant modes of interaction within the genome. Some of these modes correspond to systematic biases that can then be subtracted out. I will then show that a pairwise interaction model can be fit to the corrected free energies to determine the couplings between known bound chromatin factors. By correcting for the systematic effects identified by PCA, a consistent set of predictions for the couplings among the various chromatin factors can be made. Many of the known interactions within the network of chromatin factors are found along with several novel predictions. Finally, I will present efforts to predict the local 3D structure of chromatin using the fitted interaction model and the locations of bound factors. [Preview Abstract] |
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