3D Organization Research Articles

Targeting Noncoding cis-Regulatory Elements for Cancer Therapy in the Context of the 3D Genome.

Significant efforts have been made to identify and validate oncoproteins and ncRNAs as therapeutic targets for cancer therapy; however, emerging observations suggest that noncoding cis-regulatory elements, which orchestrate the 3D organization of the genome and thus the transcriptional landscape, are potential therapeutic targets as well. In this commentary, we envisage that further efforts to decipher the noncoding cis-regulatory code and performing systematic surveys of functional noncoding cis-regulatory elements and recurrent 3D genome alterations in both cancerous and nonmalignant cells within tumor tissues will pave the way to the development of novel therapeutic strategies.

Mutations of PDS5 genes enhance TAD-like domain formation Arabidopsis thaliana

In eukaryotes, topologically associating domains (TADs) organize the genome into functional compartments. While TAD-like structures are common in mammals and many plants, they are challenging to detect in Arabidopsis thaliana. Here, we demonstrate that Arabidopsis PDS5 proteins play a negative role in TAD-like domain formation. Through Hi-C analysis, we show that mutations in PDS5 genes lead to the widespread emergence of enhanced TAD-like domains throughout the Arabidopsis genome, excluding pericentromeric regions. These domains exhibit increased chromatin insulation and enhanced chromatin interactions, without significant changes in gene expression or histone modifications. Our results suggest that PDS5 proteins are key regulators of genome architecture, influencing 3D chromatin organization independently of transcriptional activity. This study provides insights into the unique chromatin structure of Arabidopsis and the broader mechanisms governing plant genome folding.

An integrated view of the structure and function of the human 4D nucleome.

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.

Targeting the 3D genome by anthracyclines for chemotherapeutic effects.

The chromatins are folded into three-dimensional (3D) structures inside cells, which coordinates the regulation of gene transcription by the non-coding regulatory elements. Aberrant chromatin 3D folding has been shown in many diseases, such as acute myeloid leukemia (AML), and may contribute to tumorigenesis. The anthracycline topoisomerase II inhibitors can induce histone eviction and DNA damage. We performed genome-wide high-resolution mapping of the chemotherapeutic effects of various clinically used anthracycline drugs. ATAC-seq was used to profile the histone eviction effects of different anthracyclines. TOP2A ChIP-seq was used to profile the potential DNA damage regions. Integrated analyses show that different anthracyclines have distinct target selectivity on epigenomic regions, based on their respective ATAC-seq and ChIP-seq profiles. We identified the underlying molecular mechanism that unique anthracycline variants selectively target chromatin looping anchors via disrupting CTCF binding, suggesting an additional potential therapeutic effect on the 3D genome. We further performed Hi-C experiments, and data from K562 cells treated with the selective anthracycline drugs indicate that the 3D chromatin organization is disrupted. Furthermore, AML patients receiving anthracycline drugs showed altered chromatin structures around potential looping anchors, which linked to distinct clinical outcomes. Our data indicate that anthracyclines are potent and selective epigenomic targeting drugs and can target the 3D genome for anticancer therapy, which could be used for personalized medicine to treat tumors with aberrant 3D chromatin structures.

A guide to studying 3D genome structure and dynamics in the kidney.

The human genome is tightly packed into the 3D environment of the cell nucleus. Rapidly evolving and sophisticated methods of mapping 3D genome architecture have shed light on fundamental principles of genome organization and gene regulation. The genome is physically organized on different scales, from individual genes to entire chromosomes. Nuclear landmarks such as the nuclear envelope and nucleoli have important roles in compartmentalizing the genome within the nucleus. Genome activity (for example, gene transcription) is also functionally partitioned within this 3D organization. Rather than being static, the 3D organization of the genome is tightly regulated over various time scales. These dynamic changes in genome structure over time represent the fourth dimension of the genome. Innovative methods have been used to map the dynamic regulation of genome structure during important cellular processes including organism development, responses to stimuli, cell division and senescence. Furthermore, disruptions to the 4D genome have been linked to various diseases, including of the kidney. As tools and approaches to studying the 4D genome become more readily available, future studies that apply these methods to study kidney biology will provide insights into kidney function in health and disease.

7690 Acute hormonal signaling-induced 3D genome organization controls transcriptional dynamics during adipocyte thermogenesis

Abstract Disclosure: Y. Zhang: None. R. Zheng: None. T. Tsuji: None. Y. Xing: None. C. Wang: None. J. Darcy: None. K. Chen: None. Y. Tseng: None. Brown adipose tissue (BAT) maintains body temperature by dissipating energy as heat, presenting a promising target against obesity and its associated metabolic disorders. When exposed to cold, the β3-adrenergic receptor (β3-AR) in brown adipocytes is stimulated, initiating a sequential signaling cascade and activating a thermogenic gene program essential for heat generation. Despite remarkable progress has been made towards understanding the engagement of transcriptional and epigenetic regulators during thermogenesis in response to β3-AR stimulation, the precise impact of three-dimensional (3D) genome organization, an increasingly important modulator of gene expression, on the activation of thermogenic gene program remains elusive. In this study, we employed high-resolution Micro-C in murine brown adipocytes to interrogate the rapidity of 3D genome reorganization in response to β3-AR hormonal signaling and its impact on transcriptional dynamics during thermogenesis. Our findings unveiled substantial rewiring of chromatin loops within 4 hours of β3-AR stimulation. These dynamic changes in chromatin loops, particularly enhancer-promoter (E-P) interactions, were coupled with gene activation during thermogenesis. Furthermore, we uncovered the mechanisms by which β3-AR hormonal signaling mediates 3D chromatin reorganization to regulate the thermogenic gene program. We found that β3-AR hormonal signaling induced the phosphorylation of p18Hamlet, a subunit of the SRCAP complex, facilitating the deposition of the histone variant H2A.Z into regulatory DNA regions. This nucleosome incorporation of H2A.Z further mediated nucleosome rearrangement, favoring the formation of nucleosome-depleted regions (NDRs) and enhancing the accessibility of regulatory DNA regions. As a result, H2A.Z chromatin incorporation facilitated interactions between cis-regulatory regions and gene promoters, such as E-P interactions, thereby activating thermogenic gene expression in response to β3-AR stimulation. Loss of H2A.Z disrupted loop dynamics, impeding gene activation and BAT thermogenic capacity. The conserved role of H2A.Z in human thermogenesis was validated, and we identified adiposity-related SNPs in human homologous loop anchors that could potentially be targeted for treating obesity. Taken together, our results unveil a previously underappreciated aspect of acute hormonal signaling-induced chromatin dynamics in orchestrating the thermogenic program in brown adipocytes. These findings not only enhance our understanding of molecular regulation of thermogenesis but also shed light on the role of rapid and dynamic chromatin reconfiguration in regulating the cellular response to external stimuli. Our study bridges a critical gap between 3D genome organization and gene activation in the thermogenic program mediated by acute β3-AR hormonal signaling. Presentation: 6/2/2024

Predicting p53-dependent cell transitions from thermodynamic models.

A cell's fate involves transitions among its various states, each defined by a distinct gene expression profile governed by the topology of gene regulatory networks, which are affected by 3D genome organization. Here, we develop thermodynamic models to determine the fate of a malignant cell as governed by the tumor suppressor p53 signaling network, taking into account long-range chromatin interactions in the mean-field approximation. The tumor suppressor p53 responds to stress by selectively triggering one of the potential transcription programs that influence many layers of cell signaling. These range from p53 phosphorylation to modulation of its DNA binding affinity, phase separation phenomena, and internal connectivity among cell fate genes. We use the minimum free energy of the system as a fundamental property of biological networks that influences the connection between the gene network topology and the state of the cell. We constructed models based on network topology and equilibrium thermodynamics. Our modeling shows that the binding of phosphorylated p53 to promoters of target genes can have properties of a first order phase transition. We apply our model to cancer cell lines ranging from breast cancer (MCF-7), colon cancer (HCT116), and leukemia (K562), with each one characterized by a specific network topology that determines the cell fate. Our results clarify the biological relevance of these mechanisms and suggest that they represent flexible network designs for switching between developmental decisions.

Comparing continuum and direct fiber models of soft tissues: An ocular biomechanics example reveals that continuum models may artificially disrupt the strains at both the tissue and fiber levels

Collagen fibers are the main load-bearing component of soft tissues but difficult to incorporate into models. Whilst simplified homogenization models suffice for some applications, a thorough mechanistic understanding requires accurate prediction of fiber behavior, including both detailed fiber-level strains and long-distance transmission. Our goal was to compare the performance of a continuum model of the optic nerve head (ONH) built using conventional techniques with a fiber model we recently introduced which explicitly incorporates the complex 3D organization and interaction of collagen fiber bundles [1]. To ensure a fair comparison, we constructed the continuum model with identical geometrical, structural, and boundary specifications as for the fiber model. We found that: 1) although both models accurately matched the intraocular pressure (IOP)-induced globally averaged displacement responses observed in experiments, they diverged significantly in their ability to replicate specific 3D tissue-level strain patterns. Notably, the fiber model faithfully replicated the experimentally observed depth-dependent variability of radial strain, the ring-like pattern of meridional strain, and the radial pattern of circumferential strain, whereas the continuum model failed to do so; 2) the continuum model disrupted the strain transmission along each fiber, a feature captured well by the fiber model. These results demonstrate limitations of the conventional continuum models that rely on homogenization and affine deformation assumptions, which render them incapable of capturing some complex tissue-level and fiber-level deformations. Our results show that the strengths of explicit fiber modeling help capture intricate ONH biomechanics. They potentially also help modeling other fibrous tissues. Statement of SignificanceUnderstanding the mechanics of fibrous tissues is crucial for advancing knowledge of various diseases. This study uses the ONH as a test case to compare conventional continuum models with fiber models that explicitly account for the complex fiber structure. We found that the fiber model captured better the biomechanical behaviors at both the tissue level and the fiber level. The insights gained from this study demonstrate the significant potential of fiber models to advance our understanding of not only glaucoma pathophysiology but also other conditions involving fibrous soft tissues. This can contribute to the development of therapeutic strategies across a wide range of applications.

Peculiar k-mer Spectra Are Correlated with 3D Contact Frequencies and Breakpoint Regions in the Human Genome.

It is widely accepted that the 3D chromatin organization in human cell nuclei is not random and recent investigations point towards an interactive relation of epigenetic functioning and chromatin (re-)organization. Although chromatin organization seems to be the result of self-organization of the entirety of all molecules available in the cell nucleus, a general question remains open as to what extent chromatin organization might additionally be predetermined by the DNA sequence and, if so, if there are characteristic differences that distinguish typical regions involved in dysfunction-related aberrations from normal ones, since typical DNA breakpoint regions involved in disease-related chromosome aberrations are not randomly distributed along the DNA sequence. Highly conserved k-mer patterns in intronic and intergenic regions have been reported in eukaryotic genomes. In this article, we search and analyze regions deviating from average spectra (ReDFAS) of k-mer word frequencies in the human genome. This includes all assembled regions, e.g., telomeric, centromeric, genic as well as intergenic regions. A positive correlation between k-mer spectra and 3D contact frequencies, obtained exemplarily from given Hi-C datasets, has been found indicating a relation of ReDFAS to chromatin organization and interactions. We also searched and found correlations of known functional annotations, e.g., genes correlating with ReDFAS. Selected regions known to contain typical breakpoints on chromosomes 9 and 5 that are involved in cancer-related chromosomal aberrations appear to be enriched in ReDFAS. Since transposable elements like ALUs are often assigned as major players in 3D genome organization, we also studied their impact on our examples but could not find a correlation between ALU regions and breakpoints comparable to ReDFAS. Our findings might show that ReDFAS are associated with instable regions of the genome and regions with many chromatin contacts which is in line with current research indicating that chromatin loop anchor points lead to genomic instability.

Cohesin organizes 3D DNA contacts surrounding active enhancers in C. elegans.

In mammals, cohesin and CTCF organize the 3D genome into topologically associated domains (TADs) to regulate communication between cis-regulatory elements. Many organisms, including S. cerevisiae, C. elegans, and A. thaliana contain cohesin but lack CTCF. Here, we used C. elegans to investigate the function of cohesin in 3D genome organization in the absence of CTCF. Using Hi-C data, we observe cohesin-dependent features called "fountains", which are also reported in zebrafish and mice. These are population average reflections of DNA loops originating from distinct genomic regions and are ~20-40 kb in C. elegans. Hi-C analysis upon cohesin and WAPL depletion support the idea that cohesin is preferentially loaded at NIPBL occupied sites and loop extrudes in an effectively two-sided manner. ChIP-seq analyses show that cohesin translocation along the fountain trajectory depends on a fully intact complex and is extended upon WAPL-1 depletion. Hi-C contact patterns at individual fountains suggest that cohesin processivity is unequal on each side, possibly due to collision with cohesin loaded from surrounding sites. The putative cohesin loading sites are closest to active enhancers and fountain strength is associated with transcription. Compared to mammals, average processivity of C. elegans cohesin is ~10-fold shorter and NIPBL binding does not depend on cohesin. We propose that preferential loading and loop extrusion by cohesin is an evolutionarily conserved mechanism that regulates the 3D interactions of enhancers in animal genomes.

Nucleosome placement and polymer mechanics explain genomic contacts on 100kbp scales.

The 3d organization of the genome - in particular, which two regions of DNA are in contact with each other - plays a role in regulating gene expression. Several factors influence genome 3d organization. Nucleosomes (where ~ 100 basepairs of DNA wrap around histone proteins) also bend, twist and compactify chromosomal DNA, altering its polymer mechanics. How much does the positioning of nucleosomes between gene loci influence contacts between those gene loci? And, to what extent is polymer mechanics responsible for this? To address this question, we combine a stochastic polymer mechanics model of chromosomal DNA including twists and wrapping induced by nucleosomes with two data-driven pipelines. The first estimates nucleosome positioning from ATACseq data in regions of high accessibility. Most of the genome is low-accessibility, so we combine this with a novel image analysis method that estimates the distribution of nucleosome spacing from electron microscopy data. There are no free parameters in the biophysical model. We apply this method to IL6, IL15, CXCL9, and CXCL10, inflammatory marker genes in macrophages, before and after immune stimulation, and compare the predictions with contacts measured by conformation capture experiments (4C-seq). We find that within a 500 kilo-basepairs genomic region, polymer mechanics with nucleosomes can explain 71% of close contacts. These results suggest that, while genome contacts on 100kbp-scales are multifactorial, they may be amenable to mechanistic, physical explanation. Our work also highlights the role of nucleosomes, not just at the loci of interest, but between them, and not just the total number of nucleosomes, but their specific placement. The method generalizes to other genes, and can be used to address whether a contact is under active regulation by the cell (e.g., a macrophage during inflammatory stimulation). Importantly, our findings suggest that gene function may have evolved through selective pressures that co-opted contact-mediated regulatory mechanisms reliant largely on polymer mechanics.

Virus Infection Induces Immune Gene Activation with CTCF Anchored Enhancers and Chromatin Interactions in Pig Genome.

Chromatin organization is important for gene transcription in pig genome. However, its three-dimensional (3D) structure and dynamics are much less investigated than those in human. Here we applied the long-reads chromatin interaction analysis by paired-end tag sequencing (ChIA-PET) method to map the whole-genome chromatin interactions mediated by CCCTC-binding factor (CTCF) and RNA polymerase Ⅱ (RNAPⅡ or POLⅡ) in porcine macrophage cells before and after polyinosinic-polycytidylic acid [Poly(I:C)] induction. Our results revealed that Poly(I:C) induction impacts the 3D genome organization in the 3D4/21 cells at the fine-scale chromatin loop level rather than at the large-scale domain level. Furthermore, our findings underscored the pivotal role of CTCF anchored chromatin interactions in reshaping chromatin architecture during immune responses. Knock-out of the CTCF locus further confirmed that the CTCF anchored enhancers are associated with the activation of immune genes via long-range interactions. Notably, ChIA-PET data also supported the spatial relationship between single nucleotide polymorphisms (SNPs) and the related gene transcription in 3D genome aspect. Our findings in this study provide new clues and potential targets to explore key elements related to diseases in swine and are also likely to shed light on elucidating chromatin organization and dynamics underlying the process of mammalian infectious diseases.

Molecular pathology, developmental changes and synaptic dysfunction in (pre-) symptomatic human C9ORF72-ALS/FTD cerebral organoids

A hexanucleotide repeat expansion (HRE) in C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Human brain imaging and experimental studies indicate early changes in brain structure and connectivity in C9-ALS/FTD, even before symptom onset. Because these early disease phenotypes remain incompletely understood, we generated iPSC-derived cerebral organoid models from C9-ALS/FTD patients, presymptomatic C9ORF72-HRE (C9-HRE) carriers, and controls. Our work revealed the presence of all three C9-HRE-related molecular pathologies and developmental stage-dependent size phenotypes in cerebral organoids from C9-ALS/FTD patients. In addition, single-cell RNA sequencing identified changes in cell type abundance and distribution in C9-ALS/FTD organoids, including a reduction in the number of deep layer cortical neurons and the distribution of neural progenitors. Further, molecular and cellular analyses and patch-clamp electrophysiology detected various changes in synapse structure and function. Intriguingly, organoids from all presymptomatic C9-HRE carriers displayed C9-HRE molecular pathology, whereas the extent to which more downstream cellular defects, as found in C9-ALS/FTD models, were detected varied for the different presymptomatic C9-HRE cases. Together, these results unveil early changes in 3D human brain tissue organization and synaptic connectivity in C9-ALS/FTD that likely constitute initial pathologies crucial for understanding disease onset and the design of therapeutic strategies.

Different Biomechanical Cell Behaviors in an Epithelium Drive Collective Epithelial Cell Extrusion.

In vertebrates, many organs, such as the kidney and the mammary gland form ductal structures based on the folding of epithelial sheets. The development of these organs relies on coordinated sorting of different cell lineages in both time and space, through mechanisms that remain largely unclear. Tissues are composed of several cell types with distinct biomechanical properties, particularly at cell-cell and cell-substrate boundaries. One hypothesis is that adjacent epithelial layers work in a coordinated manner to shape the tissue. Using in vitro experiments on model epithelial cells, differential expression of atypical Protein Kinase C iota (aPKCi), a key junctional polarity protein, is shown to reinforce cell epithelialization and trigger sorting by tuning cell mechanical properties at the tissue level. In a broader perspective, it is shown that in a heterogeneous epithelial monolayer, in which cell sorting occurs, forces arising from epithelial cell growth under confinement by surrounding cells with different biomechanical properties are sufficient to promote collective cell extrusion and generate emerging 3D organization related to spheroids and buds. Overall, this research sheds light on the role of aPKCi and the biomechanical interplay between distinct epithelial cell lineages in shaping tissue organization, providing insights into the understanding of tissue and organ development.

Step-by-Step Protocol to Generate Hi-C Contact Maps Using the rfy_hic2 Pipeline.

The Hi-C method has emerged as an indispensable tool for analyzing the 3D organization of the genome, becoming increasingly accessible and frequently utilized in chromatin research. To effectively leverage 3D genomics data obtained through advanced technologies, it is crucial to understand what processes are undertaken and what aspects require special attention within the bioinformatics pipeline. This protocol aims to demystify the Hi-C data analysis process for field newcomers. In a step-by-step manner, we describe how to process Hi-C data, from the initial sequencing of the Hi-C library to the final visualization of Hi-C contact data as heatmaps. Each step of the analysis is clearly explained to ensure an understanding of the procedures and their objectives. By the end of this chapter, readers will be equipped with the knowledge to transform raw Hi-C reads into informative visual representations, facilitating a deeper comprehension of the spatial genomic structures critical to cellular functions.

Abstract A083: Genomic Analysis of Structural Variations in Pancreatic Cancer Using Long-Read Sequencing

Abstract Objective: Structural variations (SVs) constitute the greatest source of genomic alterations leading to the oncogenesis of many cancers, including the deadliest pancreatic cancer (PC). However, SVs in PC remain largely undefined due to technological limitations of the traditional short-read sequencing. This study aims to establish a comprehensive signature of genomic SVs in PC. Methods: PacBio whole genome long-read sequencing was employed to investigate the genomic landscape of SVs in eight tissues from four pancreatic cancer patients. A variety of SVs callers, including pbsv, Sniffles, cuteSV, and svim, were comprehensively adopted to improve the accuracy of SVs detection. Many undefined characteristics of germline and somatic SVs were revealed by comparing with multiple previously published databases. Besides, the impacts of structural variations on 3D chromatin organization were analyzed by integrating multi-omics data including Hi-C, RNA seq and ChIP seq. Results: A total of 26,844 and 27,028 non-redundant SVs were identified in human pancreatic cancer and matched para-cancerous tissues, respectively. Notably, among these SVs, 23.97% were novel and had not been previously reported. Germline SVs were further characterized and found enriched in 25 well-known susceptible genes, such as TP63, PVT1, and ABO. Additionally, we proposed a panel of germline SVs with predicted highest pathogenicity, involving STOX1, DSPP, and WRN. Totally, 616 somatic SVs were identified and presented high burdens in patients with lymph node metastasis. We observed that somatic DELs and INSs were significantly more prevalent in repetitive regions than in non-repetitive regions. Interestingly, DELs showed a decrease in the proportion occurring in simple repeat regions but an increase in the proportion occurring in LINE and SINE regions compared to INSs. Moreover, it was observed that deletions within topologically associated domains were associated with local enhanced interactions and disrupted chromatin loops, which might influence H3K27ac modification of histone contributing to enhancer or silencer hijacking. Conclusion: This study provides insights into the genomic signature of SVs in human pancreatic cancer. These findings might shed new light on the complexity of SVs and their pathogenesis by interplaying with chromatin organization. Citation Format: Yongxing Du, Xiaohao Zheng, Yunjie Duan, Chengfeng Wang. Genomic Analysis of Structural Variations in Pancreatic Cancer Using Long-Read Sequencing [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A083.

Nucleotide excision repair of aflatoxin-induced DNA damage within the 3D human genome organization.

Aflatoxin B1 (AFB1), a potent mycotoxin, is one of the environmental risk factors that cause liver cancer. In the liver, the bioactivated AFB1 intercalates into the DNA double helix to form a bulky DNA adduct which will lead to mutation if left unrepaired. Here, we adapted the tXR-seq method to measure the nucleotide excision repair of AFB1-induced DNA adducts at single-nucleotide resolution on a genome-wide scale, and compared it with repair data obtained from conventional UV-damage XR-seq. Our results showed that transcription-coupled repair plays a major role in the damage removal process. We further analyzed the distribution of nucleotide excision repair sites for AFB1-induced DNA adducts within the 3D human genome organization. Our analysis revealed a heterogeneous AFB1-dG repair across four different organization levels, including chromosome territories, A/B compartments, TADs, and chromatin loops. We found that chromosomes positioned closer to the nuclear center and regions within A compartments have higher levels of nucleotide excision repair. Notably, we observed high repair activity around both TAD boundaries and loop anchors. These findings provide insights into the complex interplay between AFB1-induced DNA damage repair, transcription, and 3D genome organization, shedding light on the mechanisms underlying AFB1-induced mutagenesis.

Comparing continuum and direct fiber models of soft tissues. An ocular biomechanics example reveals that continuum models may artificially disrupt the strains at both the tissue and fiber levels.

Collagen fibers are the main load-bearing component of soft tissues but difficult to incorporate into models. Whilst simplified homogenization models suffice for some applications, a thorough mechanistic understanding requires accurate prediction of fiber behavior, including both detailed fiber-level strains and long-distance transmission. Our goal was to compare the performance of a continuum model of the optic nerve head (ONH) built using conventional techniques with a fiber model we recently introduced which explicitly incorporates the complex 3D organization and interaction of collagen fiber bundles [1]. To ensure a fair comparison, we constructed the continuum model with identical geometrical, structural, and boundary specifications as for the fiber model. We found that: 1) although both models accurately matched the intraocular pressure (IOP)-induced globally averaged displacement responses observed in experiments, they diverged significantly in their ability to replicate specific 3D tissue-level strain patterns. Notably, the fiber model faithfully replicated the experimentally observed depth-dependent variability of radial strain, the ring-like pattern of meridional strain, and the radial pattern of circumferential strain, whereas the continuum model failed to do so; 2) the continuum model disrupted the strain transmission along each fiber, a feature captured well by the fiber model. These results demonstrate limitations of the conventional continuum models that rely on homogenization and affine deformation assumptions, which render them incapable of capturing some complex tissue-level and fiber-level deformations. Our results show that the strengths of explicit fiber modeling help capture intricate ONH biomechanics. They potentially also help modeling other fibrous tissues.

Electrophysiological features of cortical 3D networks are deeply modulated by scaffold properties.

Three-dimensionality (3D) was proven essential for developing reliable models for different anatomical compartments and many diseases. However, the neuronal compartment still poses a great challenge as we still do not understand precisely how the brain computes information and how the complex chain of neuronal events can generate conscious behavior. Therefore, a comprehensive model of neuronal tissue has not yet been found. The present work was conceived in this framework: we aimed to contribute to what must be a collective effort by filling in some information on possible 3D strategies to pursue. We compared directly different kinds of scaffolds (i.e., PDMS sponges, thermally crosslinked hydrogels, and glass microbeads) in their effect on neuronal network activity recorded using micro-electrode arrays. While the overall rate of spiking activity remained consistent, the type of scaffold had a notable impact on bursting dynamics. The frequency, density of bursts, and occurrence of random spikes were all affected. The examination of inter-burst intervals revealed distinct burst generation patterns unique to different scaffold types. Network burst propagation unveiled divergent trends among configurations. Notably, it showed the most differences, underlying that functional variations may arise from a different 3D spatial organization. This evidence suggests that not all 3D neuronal constructs can sustain the same level of richness of activity. Furthermore, we commented on the reproducibility, efficacy, and scalability of the methods, where the beads still offer superior performances. By comparing different 3D scaffolds, our results move toward understanding the best strategies to develop functional 3D neuronal units for reliable pre-clinical studies.

Deep Learning Sequence Models for Transcriptional Regulation.

Deciphering the regulatory code of gene expression and interpreting the transcriptional effects of genome variation are critical challenges in human genetics. Modern experimental technologies have resulted in an abundance of data, enabling the development of sequence-based deep learning models that link patterns embedded in DNA to the biochemical and regulatory properties contributing to transcriptional regulation, including modeling epigenetic marks, 3D genome organization, and gene expression, with tissue and cell-type specificity. Such methods can predict the functional consequences of any noncoding variant in the human genome, even rare or never-before-observed variants, and systematically characterize their consequences beyond what is tractable from experiments or quantitative genetics studies alone. Recently, the development and application of interpretability approaches have led to the identification of key sequence patterns contributing to the predicted tasks, providing insights into the underlying biological mechanisms learned and revealing opportunities for improvement in future models.