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Glioblastoma multiforme (GBM) encompasses brain malignancies marked by phenotypic and transcriptional heterogeneity thought to render these tumors aggressive, resistant to therapy, and inevitably recurrent. However, little is known about how the spatial organization of GBM genomes underlies this heterogeneity and its effects. Here, we compiled a cohort of 28 patient-derived glioblastoma stem cell-like lines (GSCs) known to reflect the properties of their tumor-of-origin; six of these were primary-relapse tumor pairs from the same patient. We generated and analyzed kbp-resolution chromosome conformation capture (Hi-C) data from all GSCs to systematically map >3,100 standalone and complex structural variants (SVs) and the >6,300 neoloops arising as a result. By combining Hi-C, histone modification, and gene expression data with chromatin folding simulations, we explain how the pervasive, uneven, and idiosyncratic occurrence of neoloops sustains tumor-specific transcriptional programs via the formation of new enhancer-promoter contacts. We also show how even moderately recurrent neoloops can help us infer patient-specific vulnerabilities. Together, our data provide a resource for dissecting GBM biology and heterogeneity, as well as for informing therapeutic approaches.

Python scripts to implement Loop Extrusion mechanism and RNAPolII-driven transcription condensates formation.The framework relies on the API of EspressoMD to perform Molecular Dynamics simulations.

SUMMARYThe spatiotemporal configuration of genes with distal regulatory elements, and the impact of chromatin mobility on transcription, remain unclear. Loop extrusion is an attractive model for bringing genetic elements together, but how this functionally interacts with transcription is also largely unknown. We combine live tracking of genomic loci and nascent transcripts with molecular dynamics simulations to assess the 4D arrangement of theSox2gene and its enhancer, in response to a battery of perturbations. We find that alterations in chromatin mobility, not promoter-enhancer distance, is more informative about transcriptional status. Active elements display more constrained mobility, consistent with confinement within specialized nuclear sites, and alterations in enhancer mobility distinguish poised from transcribing alleles. Strikingly, we find that whereas loop extrusion and transcription factor-mediated clustering contribute to promoter-enhancer proximity, they have antagonistic effects on chromatin dynamics. This provides an experimental framework for the underappreciated role of chromatin dynamics in genome regulation.

The combination of modelling and experimental advances can provide deep insights for understanding chromatin 3D organization and ultimately its underlying mechanisms. In particular, models of polymer physics can help comprehend the complexity of genomic contact maps, as those emerging from technologies such as Hi-C, GAM or SPRITE. Here we discuss a method to reconstruct 3D structures from Genome Architecture Mapping (GAM) data, based on PRISMR, a computational approach introduced to find the minimal polymer model best describing Hi-C input data from only polymer physics. After recapitulating the PRISMR procedure, we describe how we extended it for treating GAM data. We successfully test the method on a 6 Mb region around the Sox9 gene and, at a lower resolution, on the whole chromosome 7 in mouse embryonic stem cells. The PRISMR derived 3D structures from GAM co-segregation data are finally validated against independent Hi-C contact maps. The method results to be versatile and robust, hinting that it can be similarly applied to different experimental data, such as SPRITE or microscopy distance data.

By simulating the use of single photon detector for coherent laser detection we performed a feasibility study for earth-satellite Dual Comb Spectroscopy (DCS) through numerical simulation. We conceived a scheme to use a single optical frequency comb source to perform DCS experiments and then we extended this scheme for application of DCS on earth-satellite path using the Doppler induced frequency shift to simulate the second comb source. We simulated the acquisition using single photon detector and an event timer to assign the time of photon detection. Then we calculated the probability density function through time-interval between two successive photoelectrons to extract the DCS signal. Finally we tested the conceived scheme on real high repetition rate satellite laser ranging data obtaining promising results.

Frequency modulated continuous wave (FMCW) laser detection and ranging is a technique for absolute distance measurements with high performances in terms of resolution, non-ambiguity range, accuracy and fast detection. It is based on a simple experimental setup, thus resulting in cost restraint with potential wide spread, not only limited to research institutions. The technique has been widely studied and improved both in terms of experimental setup by absolute reference or active stabilization and in terms of data analysis. Very recently a multi-wavelength approach has been exploited, demonstrating high precision and non ambiguity range. The variability of refractive index along the path was not taken into account with consequent degradation of range accuracy. In this work we developed a simple model able to take into account refractive index effect in multi-wavelength FMCW measurement. We performed a numerical simulation in different atmospheric conditions of temperature, pressure, humidity and CO2 concentration showing a net improvement of range accuracy when refractive index modeling is used.

The organization of the genome in the nucleus and the interactions of genes with their regulatory elements are key features of transcriptional control and their disruption can cause disease. Here we report a genome-wide method, genome architecture mapping (GAM), for measuring chromatin contacts and other features of three-dimensional chromatin topology on the basis of sequencing DNA from a large collection of thin nuclear sections. We apply GAM to mouse embryonic stem cells and identify enrichment for specific interactions between active genes and enhancers across very large genomic distances using a mathematical model termed SLICE (statistical inference of co-segregation). GAM also reveals an abundance of three-way contacts across the genome, especially between regions that are highly transcribed or contain super-enhancers, providing a level of insight into genome architecture that, owing to the technical limitations of current technologies, has previously remained unattainable. Furthermore, GAM highlights a role for gene-expression-specific contacts in organizing the genome in mammalian nuclei.

Gene expression states influence the three-dimensional conformation of the genome through poorly understood mechanisms. Here, we investigate the conformation of the murineHoxBlocus, a gene-dense genomic region containing closely spaced genes with distinct activation states in mouse embryonic stem (ES) cells. To predict possible folding scenarios, we performed computer simulations of polymer models informed with different chromatin occupancy features, which define promoter activation states or CTCF binding sites. Single cell imaging of the locus folding was performed to test model predictions. While CTCF occupancy alone fails to predict the in vivo folding at genomic length scale of 10 kb, we found that homotypic interactions between active and Polycomb-repressed promoters co-occurring in the same DNA fibre fully explain the HoxB folding patterns imaged in single cells. We identify state-dependent promoter interactions as major drivers of chromatin folding in gene-dense regions.

Gene expression states influence the 3D conformation of the genome through poorly understood mechanisms. Here, we investigate the conformation of the murine HoxB locus, a gene-dense genomic region containing closely spaced genes with distinct activation states in mouse embryonic stem (ES) cells. To predict possible folding scenarios, we performed computer simulations of polymer models informed with different chromatin occupancy features that define promoter activation states or binding sites for the transcription factor CTCF. Single-cell imaging of the locus folding was performed to test model predictions. While CTCF occupancy alone fails to predict the in vivo folding at genomic length scale of 10 kb, we found that homotypic interactions between active and Polycomb-repressed promoters co-occurring in the same DNA fiber fully explain the HoxB folding patterns imaged in single cells. We identify state-dependent promoter interactions as major drivers of chromatin folding in gene-dense regions.

Chromatin has a complex spatial organization in the cell nucleus that serves vital functional purposes. A variety of chromatin folding conformations has been detected by single-cell imaging and chromosome conformation capture-based approaches. However, a unified quantitative framework describing spatial chromatin organization is still lacking. Here, we explore the “strings and binders switch” model to explain the origin and variety of chromatin behaviors that coexist and dynamically change within living cells. This simple polymer model recapitulates the scaling properties of chromatin folding reported experimentally in different cellular systems, the fractal state of chromatin, the processes of domain formation, and looping out. Additionally, the strings and binders switch model reproduces the recently proposed “fractal–globule” model, but only as one of many possible transient conformations.

X Chromosome Inactivation (XCI) is the process in mammal female cells whereby one of the X chromosomes is silenced to compensate dosage with respect to males. It is still myste- rious how precisely one X chromosome is randomly chosen for inactivation. We discuss here a mean-field theory of the Symmetry Breaking (SB) model of XCI, a Statistical Mechanics model introduced to explain that process. The SB model poses that a single regulatory factor, an aggregate of molecules, is produced which acts to preserve from inactivation one of the X’s. The model illustrates a physical mechanism, originating from a thermodynamic phase transition, for the self-assembling of such a single super-molecular aggregate which can spontaneously break the binding symmetry of equivalent targets. This results in a sharp, yet stochastic, regulatory mechanism of XCI. In particular, we focus here on how the model can predict the effects of genetic deletions.

In the cell nucleus, chromosomes have a complex spatial organization, spanning several length scales, which serves vital functional purposes. It is unknown, however, how their three-dimensional architecture is orchestrated. In the present article, we review the application of a model based on classical polymer physics, the strings and binders switch model, to explain the molecular mechanisms of chromatin self-organization. We explore the scenario where chromatin architecture is shaped and regulated by the interactions of chromosomes with diffusing DNA-binding factors via thermodynamics mechanisms and compare it with available experimental data.

Understanding the mechanisms that control the organization of chromosomes in the space of the nucleus of cells, and its contribution to gene regulation, is a key open issue in molecular biology. New technologies have shown that chromosomes have a complex 3D organization, which dynamically changes across organisms and cell types. To understand such complex behaviors, quantitative models from polymer physics have been developed, to find the principles of chromosome folding, their origin and function. Here, we provide a short review of recent progress in such an important research field where Physical and Life Sciences meet.

The underlying global organization of chromatin within the cell nucleus has been the focus of intense recent research. Hi-C methods have allowed for the detection of genome-wide chromatin interactions, revealing a complex large-scale organization where chromosomes tend to partition into megabase-sized “topological domains” of local chromatin interactions and intra-chromosomal contacts extends over much longer scales, in a cell-type and chromosome specific manner. Until recently, the distinct chromatin folding properties observed experimentally have been difficult to explain in a single conceptual framework. We reported that a simple polymer-physics model of chromatin, the strings and binders switch (SBS) model, succeeds in describing the full range of chromatin configurations observed in vivo. The SBS model simulates the interactions between randomly diffusing binding molecules and binding sites on a polymer chain. It explains how polymer architectural patterns can be established, how different stable conformations can be produced and how conformational changes can be reliably regulated by simple strategies, such as protein upregulation or epigenetic modifications, via fundamental thermodynamics mechanisms.

The exploration of the spatial organiza-tion of chromosomes in the cell nucleushas been greatly enhanced by genome-scale technologies such as Hi-C methods.Polymermodels arehelpingto understandthe new emerging complex scenarios andhere we review some recent developments.In the cell nucleus of eukaryotes, chro-mosomes have a complex spatial organi-zation serving vital functional purposes,with structural disruptions being linkedto disease (Fraser and Bickmore, 2007;Lanctot et al., 2007; Misteli, 2007; Pomboand Branco, 2007). The development oftechnologies such as Hi-C ( Lieberman-Aiden et al., 2009) has opened the wayto mapping chromatin interactions ata genomic scale. It is emerging thatchromosomes tend to form 1Mb sizeddomains with increased levels of intra-interactions (known, e.g., as TopologicalDomains, TDs) (Dixon et al., 2012; Noraet al., 2012), but contacts extend acrossentire chromosomes (Branco and Pombo,2006; Shopland et al., 2006; Fraser andBickmore, 2007; Kalhor et al., 2011;Sexton et al., 2012), as highlighted bythe average contact probability of twosites

We review some of the recent progresses made by the application of polymer physics to the understanding of the spatial organization of chromosomes in the cell nucleus. In particular, we focus on some scaling properties of the contact probability of pairs of sites, discovered by recent genome-wide experiments exploiting new technologies such as Hi-C methods.

X-Chromosome Inactivation (XCI) is the process whereby one, randomly chosen X becomes transcriptionally silenced in female cells. XCI is governed by the Xic, a locus on the X encompassing an array of genes which interact with each other and with key molecular factors. The mechanism, though, establishing the fate of the X's, and the corresponding alternative modifications of the Xic architecture, is still mysterious. In this study, by use of computer simulations, we explore the scenario where chromatin conformations emerge from its interaction with diffusing molecular factors. Our aim is to understand the physical mechanisms whereby stable, non-random conformations are established on the Xic's, how complex architectural changes are reliably regulated, and how they lead to opposite structures on the two alleles. In particular, comparison against current experimental data indicates that a few key cis-regulatory regions orchestrate the organization of the Xic, and that two major molecular regulators are involved., Author Summary In mammal female cells X-Chromosome Inactivation (XCI) is the vital process whereby one X, randomly chosen, is silenced to compensate dosage of X products with respect to males. XCI is governed by a region on the X, the X Inactivation Centre (Xic), which undergoes a sequence of conformational modifications during the process. The two Xic are exposed, though, to the same environment, and it is obscure how they attain different architectures. By use of computer simulations of a molecular model, here we individuate general physical mechanisms whereby random Brownian molecules can assemble chromatin stable architectures, reliably regulate conformational changes, and establish opposite transformations on identical alleles. In the case-study of the murine Xic, our analysis highlights the existence of a few key regulatory regions and molecular factors. It also predicts, e.g., the effects of genetic modifications in the locus, which are compared with current deletion/insertion experiments. The physical mechanisms we describe are rooted in thermodynamics and could be relevant well beyond XCI.