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Features of higher-order chromatin organization—such as A/B compartments, topologically associating domains and chromatin loops—are temporarily disrupted during mitosis 1 , 2 . Because these structures are thought to influence gene regulation, it is important to understand how they are re-established after mitosis. Here we examine the dynamics of chromosome reorganization by Hi-C after mitosis in highly purified, synchronous mouse erythroid cell populations. We observed rapid establishment of A/B compartments, followed by their gradual intensification and expansion. Contact domains form from the ‘bottom up’—smaller subTADs are formed initially, followed by convergence into multi-domain TAD structures. CTCF is partially retained on mitotic chromosomes and immediately resumes full binding in ana/telophase. By contrast, cohesin is completely evicted from mitotic chromosomes and regains focal binding at a slower rate. The formation of CTCF/cohesin co-anchored structural loops follows the kinetics of cohesin positioning. Stripe-shaped contact patterns—anchored by CTCF—grow in length, which is consistent with a loop-extrusion process after mitosis. Interactions between cis -regulatory elements can form rapidly, with rates exceeding those of CTCF/cohesin-anchored contacts. Notably, we identified a group of rapidly emerging transient contacts between cis -regulatory elements in ana/telophase that are dissolved upon G1 entry, co-incident with the establishment of inner boundaries or nearby interfering chromatin loops. We also describe the relationship between transcription reactivation and architectural features. Our findings indicate that distinct but mutually influential forces drive post-mitotic chromatin reconfiguration. Analysis of the dynamics of chromosome reorganization after exit from mitosis reveals the distinct but mutually influential forces that drive chromatin reconfiguration.

Advanced liver preservation strategies could revolutionize liver transplantation by extending preservation time, thereby allowing for broader availability and better matching of transplants. However, developing new cryopreservation protocols requires exploration of a complex design space, further complicated by the scarcity of real human livers to experiment upon. We aim to create computational models of the liver to aid in the development of new cryopreservation protocols. Towards this goal, we present an approach for generating 3D models of the liver vasculature by building upon the space colonization algorithm. Additionally, we introduce the concept of a super lobule which enables a computational abstraction of biological liver lobules. User-tunable parameters allow for vasculatures of varying depth and topology to be generated. In each model, we solve for a common lumped resistance value assigned to the super lobules, allowing the overall physiological blood pressure and flow rate through the liver to be preserved. We demonstrate our approach’s ability to maintain consistency between models of varying depth. Finally, we simulate steady state machine perfusion of the generated models and demonstrate how they can be used to quickly test the effect of different boundary conditions when designing organ preservation protocols.

DNA replication occurs through an intricately regulated series of molecular events and is fundamental for genome stability 1 , 2 . At present, it is unknown how the locations of replication origins are determined in the human genome. Here we dissect the role of topologically associating domains (TADs) 3 – 6 , subTADs 7 and loops 8 in the positioning of replication initiation zones (IZs). We stratify TADs and subTADs by the presence of corner-dots indicative of loops and the orientation of CTCF motifs. We find that high-efficiency, early replicating IZs localize to boundaries between adjacent corner-dot TADs anchored by high-density arrays of divergently and convergently oriented CTCF motifs. By contrast, low-efficiency IZs localize to weaker dotless boundaries. Following ablation of cohesin-mediated loop extrusion during G1, high-efficiency IZs become diffuse and delocalized at boundaries with complex CTCF motif orientations. Moreover, G1 knockdown of the cohesin unloading factor WAPL results in gained long-range loops and narrowed localization of IZs at the same boundaries. Finally, targeted deletion or insertion of specific boundaries causes local replication timing shifts consistent with IZ loss or gain, respectively. Our data support a model in which cohesin-mediated loop extrusion and stalling at a subset of genetically encoded TAD and subTAD boundaries is an essential determinant of the locations of replication origins in human S phase. A study shows that the three-dimensional conformation of the human genome influences the positioning of DNA replication initiation zones, highlighting cohesin-mediated loop anchors as essential determinants of their precise location.

Mammalian genomes are folded in a hierarchy of compartments, topologically associating domains (TADs), subTADs and looping interactions. Here, we describe 3DNetMod, a graph theory-based method for sensitive and accurate detection of chromatin domains across length scales in Hi-C data. We identify nested, partially overlapping TADs and subTADs genome wide by optimizing network modularity and varying a single resolution parameter. 3DNetMod can be applied broadly to understand genome reconfiguration in development and disease.

Mammalian genomes are folded into a hierarchy of compartments, topologically associating domains (TADs), subTADs, and long-range looping interactions. The higher-order folding patterns of chromatin contacts within TADs and how they localize to disease-associated single nucleotide variants (daSNVs) remains an open area of investigation. Here, we analyze high-resolution Hi-C data with graph theory to understand possible mesoscale network architecture within chromatin domains. We identify a subset of TADs exhibiting strong core-periphery mesoscale structure in embryonic stem cells, neural progenitor cells, and cortical neurons. Hyper-connected core nodes co-localize with genomic segments engaged in multiple looping interactions and enriched for occupancy of the architectural protein CCCTC binding protein (CTCF). CTCF knockdown and in silico deletion of CTCF-bound core nodes disrupts core-periphery structure, whereas in silico mutation of cell type-specific enhancer or gene nodes has a negligible effect. Importantly, neuropsychiatric daSNVs are significantly more likely to localize with TADs folded into core-periphery networks compared to domains devoid of such structure. Together, our results reveal that a subset of TADs encompasses looping interactions connected into a core-periphery mesoscale network. We hypothesize that daSNVs in the periphery of genome folding networks might preserve global nuclear architecture but cause local topological and functional disruptions contributing to human disease. By contrast, daSNVs co-localized with hyper-connected core nodes might cause severe topological and functional disruptions. Overall, these findings shed new light into the mesoscale network structure of fine scale genome folding within chromatin domains and its link to common genetic variants in human disease.

A method that allows a fast and accurate registration of digital tissue models obtained during preoperative, diagnostic imaging with those captured intraoperatively using lower-fidelity ultrasound imaging techniques is presented. Minimally invasive surgeries are often planned using preoperative, high-fidelity medical imaging techniques such as MRI and CT imaging. While these techniques allow clinicians to obtain detailed 3D models of the surgical region of interest (ROI), various factors such as physical changes to the tissue, changes in the body’s configuration, or apparatus used during the surgery may cause large, non-linear deformations of the ROI. Such deformations of the tissue can result in a severe mismatch between the preoperatively obtained 3D model and the real-time image data acquired during surgery, potentially compromising surgical success. To overcome this challenge, this work presents a new approach for predicting intraoperative soft tissue deformations. The approach works by simply tracking the displacements of a handful of fiducial markers or analogous biological features embedded in the tissue, and produces a 3D deformed version of the high-fidelity ROI model that registers accurately with the intraoperative data. In an offline setting, we use the finite element method to generate deformation fields given various boundary conditions that mimic the realistic environment of soft tissues during a surgery. To reduce the dimensionality of the 3D deformation field involving thousands of degrees of freedom, we use an autoencoder neural network to encode each computed deformation field into a short latent space representation, such that a neural network can accurately map the fiducial marker displacements to the latent space. Our computational tests on a head and neck tumor, a kidney, and an aorta model show prediction errors as small as 0.5 mm. Considering that the typical resolution of interventional ultrasound is around 1 mm and each prediction takes less than 0.5 s, the proposed approach has the potential to be clinically relevant for an accurate tracking of soft tissue deformations during image-guided surgeries.

This dissertation examines Two Streams, a cantata for choir and string orchestra by Daniel Knaggs, from analytical, textual, and performance-oriented perspectives, providing conductors with a clearer understanding of the work and practical guidance for its preparation and performance. Joining texts from Scripture, the Catholic Mass, and the Diary of Saint Faustina Kowalska (1905-1938), the cantata centers on the theme of Divine Mercy. The focal point of the “two streams”—the blood and water flowing from Christ's pierced side—serves as the primary metaphor through which the work explores the relationship between human cruelty and divine mercy. Each movement is examined individually through a combination of textual analysis, music–text relationships, musical style and texture, and conducting considerations. Hallmarks of Knaggs’s compositional language are highlighted, including his use of interval-driven harmony, open sonorities, non-functional harmonic relationships, and recurring motivic material, which contribute to the work’s large-scale unity. In addition to analytical insights, the dissertation provides practical rehearsal strategies and conducting considerations to assist conductors in navigating the score’s technical and interpretive challenges. Issues such as ensemble coordination, balance between choir and orchestra, tempo relationships, and tuning traps are addressed to facilitate efficient rehearsal and a compelling performance. By combining theological interpretation, musical analysis, and performance guidance, this study aims to increase accessibility to Two Streams for conductors and ensembles while contributing to the broader scholarship on sacred choral works.

Liver transplantation is a breakthrough treatment option that has saved innumerable lives worldwide. However, there is a significant unmet need with extensive transplant waiting lists and a general lack of availability to this life-saving procedure. Advanced preservation methods have shown the ability to significantly extend the ex vivo life of the organ, allowing for improved availability, better donor matching, assessment of marginal grafts which would typically be discarded, and even restoration of function. These advanced preservation strategies are highly complex processes with an incredible number of free design variables. Furthermore, there is scarce availability of human livers to develop these experimental protocols on. Researchers often use animal models instead, but successes are not directly translational due to differences in structure and scale.To address these challenges, we propose the creation of computational models of the liver to conduct early phases of cryopreservation protocol development. While it is impossible to fully capture all the complexity in the human liver in a model, we see these computational models as a method to inform experimentalists to more efficiently conduct experiments with plausible protocols. These computational models will allow for broad exploration of a highly complex design space, which was not possible in the constrained experimental case.We start by adapting an algorithm to generate three-dimensional vascular models of the liver. We demonstrate the ability of this method to create models of varying depth and topology, as well as the ability to generate models that closely match morphological statistics and structure. We create fully connected models of the liver, such that flow can be simulated in and out of the vasculature. With these models, we create a linear system of equations to simulate fluid flow throughout the entire vasculature. We then simulate the loading of cryoprotectants throughout the vasculature and highlight how the model can be used to interrogate the effects of varying boundary conditions on wall shear stress and cryoprotectant concentration throughout the model. We view this type of investigation as the model’s primary benefit to experimentalists and clinicians when developing new cryopreservation strategies.We extend the liver vascular model to simulate heat transfer, and we couple the fluid and heat transfer models to account for heat transfer due to perfusion using the bioheat equation. The heat transfer model produces a temperature field that is used to update the viscosity of the perfusate in the fluid model. The fluid model then solves for vessel flow rates at every step, which are in turn used to determine the heat transfer due to perfusion. We step between the two models and can account for changes in the flow and heat transfer due to the freezing of specific vessels and regions in the liver. We demonstrate how the model can be used to investigate the effect of varied boundary conditions on the spatial and temporal behavior of temperature and flow throughout the model.Finally, we utilize machine learning and Bayesian optimization techniques in conjunction with high-throughput screening techniques to optimize cryoprotective agent cocktails. We iterate between experiments to determine the viability of various cocktails, and machine learning methods that intelligently select the most informative and optimal prospective cocktails to test next. We optimize for multiple objectives, namely low toxicity and high concentration, by making use of state of the the art hypervolume Bayesian optimization methods. Similar to the liver models, this project leverages computational methods to accelerate experimental discovery for the benefit of cryopreservation protocol development.

The three-dimensional organization of the genome plays a major role in modulating biological processes such as gene expression and DNA replication, which are crucial to establishment and maintenance of cell identity in human development. Recent technological advances have enabled the creation of high-resolution maps of chromatin architecture genome-wide from which to test new biological hypotheses. In this thesis, we first present a networks based algorithm, 3DNetMod, for identifying the prominent feature of Topological Associating Domains (TADs) and their inner subTADs in high resolution Chromosome-Conformation-Capture Hi-C data. We then apply our method to chromatin dynamics during mitosis, uncovering how TADs and subTADs evolve temporally. Afterwards, we classify TADs/subTADs by the presence of other genome folding features, compartments and loops, as well as cohesin and CTCF placement and correlate with replication initiation enrichment. We show that perturbing looping TAD boundaries both locally and globally affects initiation placement, thus, genome folding architecture plays a functional role in replication. Finally, as part of the 4D-Nucleome consortium analysis, we integrate genome folding with other features of nuclear spatial positioning (SPIN states) and transcription. We show a common unifying trend amongst these multimodal features. This work results in a better understanding of the dynamical aspects of chromatin folding and how it influences and coincides with other various biological mechanisms, including aspects of DNA replication timing.

The dynamic three-dimensional (3D) organization of the human genome (the \"4D Nucleome\") is closely linked to genome function. Here, we integrate a wide variety of genomic data generated by the 4D Nucleome Project to provide a detailed view of human 3D genome organization in widely used embryonic stem cells (H1-hESCs) and immortalized fibroblasts (HFFc6). We provide extensive benchmarking of 3D genome mapping assays and integrate these diverse datasets to annotate spatial genomic features across scales. The data reveal a rich complexity of chromatin domains and their sub-nuclear positions, and over one hundred thousand structural loops and promoter-enhancer interactions. We developed 3D models of population-based and individual cell-to-cell variation in genome structure, establishing connections between chromosome folding, nuclear organization, chromatin looping, gene transcription, and DNA replication. We demonstrate the use of computational methods to predict genome folding from DNA sequence, uncovering potential effects of genetic variants on genome structure and function. Together, this comprehensive analysis contributes insights into human genome organization and enhances our understanding of connections between the regulation of genome function and 3D genome organization in general.