3D Hollow Xerogels with Ordered Cellulose Nanocrystals for Tailored Mechanical Properties.

Advanced materials with aligned cellulose nanocrystals (CNCs) have attracted much attention due to their remarkable mechanical and optical properties, but most of them still focus on 1D or 2D architectures. Herein, complex 3D architectures as pseudo catenoid hollow xerogels with aligned CNCs are prepared from dynamic hydrogels by mechanical stretching and air-drying process. Aligned CNCs endow the pseudo catenoids with distinct birefringence in addition to reinforcement. The mechanical properties of pseudo catenoid architecture are revealed for the first time to be controlled at two stages on diverse length scales. Both the aligned CNCs on the nanoscale and the geometry of the xerogels affect the mechanical properties. The inwardly curved surface of the pseudo catenoid xerogel makes the structure conducive to energy dissipation. These both stages of controls on the mechanical properties can be adjusted by changing the morphology of the initial hydrogels and the mechanical stretching ratios. These results will provide a new perspective for the design and manufacture advanced materials with tailored mechanical properties and functions.

Spatiotemporal Interaction Residual Networks with Pseudo3D for Video Action Recognition.

Action recognition is a significant and challenging topic in the field of sensor and computer vision. Two-stream convolutional neural networks (CNNs) and 3D CNNs are two mainstream deep learning architectures for video action recognition. To combine them into one framework to further improve performance, we proposed a novel deep network, named the spatiotemporal interaction residual network with pseudo3D (STINP). The STINP possesses three advantages. First, the STINP consists of two branches constructed based on residual networks (ResNets) to simultaneously learn the spatial and temporal information of the video. Second, the STINP integrates the pseudo3D block into residual units for building the spatial branch, which ensures that the spatial branch can not only learn the appearance feature of the objects and scene in the video, but also capture the potential interaction information among the consecutive frames. Finally, the STINP adopts a simple but effective multiplication operation to fuse the spatial branch and temporal branch, which guarantees that the learned spatial and temporal representation can interact with each other during the entire process of training the STINP. Experiments were implemented on two classic action recognition datasets, UCF101 and HMDB51. The experimental results show that our proposed STINP can provide better performance for video recognition than other state-of-the-art algorithms.

Characterizing 3D inflorescence architecture in grapevine using X-ray imaging and advanced morphometrics: implications for understanding cluster density.

Inflorescence architecture provides the scaffold on which flowers and fruits develop, and consequently is a primary trait under investigation in many crop systems. Yet the challenge remains to analyse these complex 3D branching structures with appropriate tools. High information content datasets are required to represent the actual structure and facilitate full analysis of both the geometric and the topological features relevant to phenotypic variation in order to clarify evolutionary and developmental inflorescence patterns. We combined advanced imaging (X-ray tomography) and computational approaches (topological and geometric data analysis and structural simulations) to comprehensively characterize grapevine inflorescence architecture (the rachis and all branches without berries) among 10 wild Vitis species. Clustering and correlation analyses revealed unexpected relationships, for example pedicel branch angles were largely independent of other traits. We identified multivariate traits that typified species, which allowed us to classify species with 78.3% accuracy, versus 10% by chance. Twelve traits had strong signals across phylogenetic clades, providing insight into the evolution of inflorescence architecture. We provide an advanced framework to quantify 3D inflorescence and other branched plant structures that can be used to tease apart subtle, heritable features for a better understanding of genetic and environmental effects on plant phenotypes.

Three-dimensional Architecture Enabled by Strained Two-dimensional Material Heterojunction.

Engineering the structure of materials endows them with novel physical properties across a wide range of length scales. With high in-plane stiffness and strength, but low flexural rigidity, two-dimensional (2D) materials are excellent building blocks for nanostructure engineering. They can be easily bent and folded to build three-dimensional (3D) architectures. Taking advantage of the large lattice mismatch between the constituents, we demonstrate a 3D heterogeneous architecture combining a basal Bi2Se3 nanoplate and wavelike Bi2Te3 edges buckling up and down forming periodic ripples. Unlike 2D heterostructures directly grown on substrates, the solution-based synthesis allows the heterostructures to be free from substrate influence during the formation process. The balance between bending and in-plane strain energies gives rise to controllable rippling of the material. Our experimental results show clear evidence that the wavelengths and amplitudes of the ripples are dependent on both the widths and thicknesses of the rippled material, matching well with continuum mechanics analysis. The rippled Bi2Se3/Bi2Te3 heterojunction broadens the horizon for the application of 2D materials heterojunction and the design and fabrication of 3D architectures based on them, which could provide a platform to enable nanoscale structure generation and associated photonic/electronic properties manipulation for optoelectronic and electromechanic applications.

Revealing the three dimensional architecture of focal adhesion components to explain Ca2+-mediated turnover of focal adhesions.

BACKGROUND: Focal adhesions (FAs) are large, dynamic protein complexes located close to the plasma membrane, which serve as the mechanical linkages and a biochemical signaling hub of cells. The coordinated and dynamic regulation of focal adhesion is required for cell migration. Degradation, or turnover, of FAs is a major event at the trailing edge of a migratory cell, and is mediated by Ca2+/calpain-dependent proteolysis and disassembly. Here, we investigated how Ca2+ influx induces cascades of FA turnover in living cells. METHODS: Images obtained with a total internal reflection fluorescence microscope (TIRFM) showed that Ca2+ ions induce different processes in the FA molecules focal adhesion kinase (FAK), paxillin, vinculin, and talin. Three mutated calpain-resistant FA molecules, FAK-V744G, paxillin-S95G, and talin-L432G, were used to clarify the role of each FA molecule in FA turnover. RESULTS: Vinculin was resistant to degradation and was not significantly affected by the presence of mutated calpain-resistant FA molecules. In contrast, talin was more sensitive to calpain-mediated turnover than the other molecules. Three-dimensional (3D) fluorescence imaging and immunoblotting demonstrated that outer FA molecules were more sensitive to calpain-mediated proteolysis than internal FA molecules. Furthermore, cell contraction is not involved in degradation of FA. CONCLUSIONS: These results suggest that Ca2+-mediated degradation of FAs was mediated by both proteolysis and disassembly. The 3D architecture of FAs is related to the different dynamics of FA molecule degradation during Ca2+-mediated FA turnover. GENERAL SIGNIFICANCE: This study will help us to clearly understand the underlying mechanism of focal adhesion turnover by Ca2+.

A random effect model for reconstruction of spatial chromatin structure.

A gene may be controlled by distal enhancers and repressors, not merely by regulatory elements in its promoter. Spatial organization of chromosomes is the mechanism that brings genes and their distal regulatory elements into close proximity. Recent molecular techniques, coupled with Next Generation Sequencing (NGS) technology, enable genome-wide detection of physical contacts between distant genomic loci. In particular, Hi-C is an NGS-aided assay for the study of genome-wide spatial interactions. The availability of such data makes it possible to reconstruct the underlying three-dimensional (3D) spatial chromatin structure. In this article, we present the Poisson Random effect Architecture Model (PRAM) for such an inference. The main feature of PRAM that separates it from previous methods is that it addresses the issue of over-dispersion and takes correlations among contact counts into consideration, thereby achieving greater consistency with observed data. PRAM was applied to Hi-C data to illustrate its performance and to compare the predicted distances with those measured by a Fluorescence In Situ Hybridization (FISH) validation experiment. Further, PRAM was compared to other methods in the literature based on both real and simulated data.

Nanoscale Channel Length MoS2 Vertical Field-Effect Transistor Arrays with Side-Wall Source/Drain Electrodes.

Two-dimensional transition metal dichalcogenides (TMDCs) have natural advantages in overcoming the short-channel effect in field-effect transistors (FETs) and in fabricating three-dimensional FETs, which benefit in increasing device density. However, so far, most reported works related to MoS2 FETs with a sub-100 nm channel employ mechanically exfoliated materials and all of the works involve electron beam lithography (EBL), which may limit their application in fabricating wafer-scale device arrays as demanded in integrated circuits (ICs). In this work, MoS2 FET arrays with a side-wall source and drain electrodes vertically distributed are designed and fabricated. The channel length of the as-fabricated FET is basically determined by the thickness of an insulating layer between the source and drain electrodes. The vertically distributed source and drain electrodes enable to reduce the electrode-occupied area and increase in the device density. The as-fabricated vertical FETs exhibit on/off ratios comparable to those of mechanically exfoliated MoS2 FETs with a nanoscale channel length under identical VDS. In addition, the as-fabricated FETs can work at a VDS as low as 10 mV with a desirable on/off ratio (1.9 × 107), which benefits in developing low-power devices. Moreover, the fabrication process is free from EBL and can be applied to wafer-scale device arrays. The statistical results show that the fabricated FET arrays have a device yield of 87.5% and an average on/off ratio of about 1.7 × 106 at a VDS of 10 mV, with the lowest and highest ones to be about 1.3 × 104 and 1.9 × 107, respectively, demonstrating the good reliability of our fabrication process. Our work promises a bright future for TMDCs in realizing high-density and low-power nanoelectronic devices in ICs.

Automated Layer Analysis (ALAn): An Image Analysis Tool for the Unbiased Characterization of Mammalian Epithelial Architecture in Culture.

Cultured mammalian cells are a common model system for the study of epithelial biology and mechanics. Epithelia are often considered as pseudo-two dimensional and thus imaged and analyzed with respect to the apical tissue surface. We found that the three-dimensional architecture of epithelial monolayers can vary widely even within small culture wells, and that layers that appear organized in the plane of the tissue can show gross disorganization in the apical-basal plane. Epithelial cell shapes should be analyzed in 3D to understand the architecture and maturity of the cultured tissue to accurately compare between experiments. Here, we present a detailed protocol for the use of our image analysis pipeline, Automated Layer Analysis (ALAn), developed to quantitatively characterize the architecture of cultured epithelial layers. ALAn is based on a set of rules that are applied to the spatial distributions of DNA and actin signals in the apical-basal (depth) dimension of cultured layers obtained from imaging cultured cell layers using a confocal microscope. ALAn facilitates reproducibility across experiments, investigations, and labs, providing users with quantitative, unbiased characterization of epithelial architecture and maturity. Key features • This protocol was developed to spatially analyze epithelial monolayers in an automated and unbiased fashion. • ALAn requires two inputs: the spatial distributions of nuclei and actin in cultured cells obtained using confocal fluorescence microscopy. • ALAn code is written in Python3 using the Jupyter Notebook interactive format. • Optimized for use in Marbin-Darby Canine Kidney (MDCK) cells and successfully applied to characterize human MCF-7 mammary gland-derived and Caco-2 colon carcinoma cells. • This protocol utilizes Imaris software to segment nuclei but may be adapted for an alternative method. ALAn requires the centroid coordinates and volume of nuclei.

One-Step Synthesis of 3D Graphene Aerogel Supported Pt Nanoparticles as Highly Active Electrocatalysts for Methanol Oxidation Reaction.

Nowadays, two of the biggest obstacles restricting the further development of methanol fuel cells are excessive cost and insufficient catalytic activity of platinum-based catalysts. Herein, platinum nanoparticle supported graphene aerogel (Pt/3DGA) was successfully synthesized by a one-step hydrothermal self-assembly method. The loose three-dimensional structure of the aerogel is stabilized by a simple one-step method, which not only reduces cost compared to the freeze-drying technology, but also optimizes the loading method of nanoparticles. The prepared Pt/3DGA catalyst has a three-dimensional porous structure with a highly cross-linked, large specific surface area, even dispersion of Pt NPs and good electrical conductivity. It is worth noting that its catalytic activity is 438.4 mA/mg with long-term stability, which is consistent with the projected benefits of anodic catalytic systems in methanol fuel cells.. Our study provides an applicable method for synthesizing nano metal particles/graphene-based composites.

Remodeling of the 3D chromatin architecture in the marine microalga Nannochloropsis oceanica during lipid accumulation.

BACKGROUND: Genomic three-dimensional (3D) spatial organization plays a key role in shaping gene expression and associated chromatin modification, and it is highly sensitive to environmental stress conditions. In microalgae, exposure to nitrogen stress can drive lipid accumulation, yet the associated functional alterations in the spatial organization of the microalgal genome have yet to be effectively characterized. RESULTS: Accordingly, the present study employed RNA-seq, Hi-C, and ChIP-seq approaches to explore the relationship between 3D chromosomal architecture and gene expression during lipid accumulation in the marine microalga Nannochloropsis oceanica in response to nitrogen deprivation (ND). These analyses revealed that ND resulted in various changes in chromosomal organization, including A/B compartment transitions, topologically associating domain (TAD) shifts, and the disruption of short-range interactions. Significantly higher levels of gene expression were evident in A compartments and TAD boundary regions relative to B compartments and TAD interior regions, consistent with observed histone modification enrichment in these areas. ND-induced differentially expressed genes (DEGs) were notably enriched in altered TAD-associated regions and regions exhibiting differential genomic contact. These DEGs were subjected to Gene Ontology (GO) term analyses that indicated they were enriched in the 'fatty acid metabolism', 'response to stress', 'carbon fixation' and 'photosynthesis' functional categories, in line with the ND treatment conditions used to conduct this study. These data indicate that Nannochloropsis cells exhibit a clear association between chromatin organization and transcriptional activity under nitrogen stress conditions. Pronounced and extensive histone modifications were evident in response to ND. Observed changes in chromatin architecture were linked to shifts in histone modifications and gene expression. CONCLUSIONS: Overall, the reprogramming of many lipid metabolism-associated genes was evident under nitrogen stress conditions with respect to both histone modifications and chromosomal organization. Together these results revealed that higher-order chromatin architecture represents a new layer that can guide efforts to understand the transcriptional regulation of lipid metabolism in nitrogen-deprived microalgae.

Tailored Sky-Parking Architectures of 3D Graphene Oxide Towards Highly-Efficient Water Purification.

The widespread use of chemicals has brought serious water pollution threatening human health and environment, which requires green, fast, and low-cost purification urgently. Here, we build up a novel material family of sky-parking-like 3D structured graphene oxides (SP-GOs) with adjustable interlayer-space of 0.8-1.7 nm via the insertion of different sized diamine compounds as support pillars between GO layers. The assembled 3D SP-GOs exhibit superior adsorption capacity and short removal time for various aromatic organic compounds in water, achieving record-breaking maximum adsorption capacity of 535.79 mg g-1 toward the most common water-pollutant bisphenol A (BPA) at ambient conditions as well as significantly improved removal of other organic pollutants including sulfapyridine, carbamazepine, ketoprofen and 2-naphthol. The construction of SP-GO provides a simple approach for evolving the GO material from 2D to 3D and a new avenue for the decontamination of pollutants in environmental remediation.

Controlled Desiccation of Preprinted Hydrogel Scaffolds Toward Complex 3D Microarchitectures.

Additive manufacturing (AM) is the key to creating a wide variety of 3D structures with unique and programmable functionalities. Direct ink writing is one of the widely used AM technologies with numerous printable materials. However, the extrude-based method is limited by low fabrication resolution, which is confined to printing macrostructures. Herein, a new AM strategy is reported, using a low-cost extrusion 3D printer, to create 3D microarchitectures at the macroscopic level through controlled desiccation of preprinted hydrogel scaffolds followed by infilling objective components. A printable hydrogel with a high-water content ensures maximum shrinkage (≈99.5% in volume) of the printed scaffolds to achieve high resolution. Stable covalent cross-linking and a suitable drying rate enable uniform shrinkage of the scaffolds to retain their original architectures. Particularly, this method can be adapted to produce liquid-metal-based 3D circuits and nanocomposite-based microrobots, indicating its capability to fabricate functional and complex 3D architectures with micron-level resolution from different material systems.

Chromosome Conformation Capture of Mitotic Chromosomes.

Despite more than a century of intensive study of mitotic chromosomes, their three-dimensional organization remains enigmatic. The last decade established Hi-C as a method of choice for study of spatial genome-wide interactions. Although its utilization has been focused mainly on studying genomic interactions in interphase nuclei, the method can be also successfully applied to study 3D architecture and genome folding in mitotic chromosomes. However, obtaining sufficient number of mitotic chromosomes as an input material and effective coupling with Hi-C method is challenging in plant species. An elegant way to overcome hindrances with obtaining a pure mitotic chromosome fraction is their isolation via flow cytometric sorting. This chapter presents a protocol describing plant sample preparation for chromosome conformation studies, for flow-sorting of plant mitotic metaphase chromosomes and for the Hi-C procedure.

The shape of our gut: Dissecting its impact on drug absorption in a 3D bioprinted intestinal model.

The small intestine is a complex organ with a characteristic architecture and a major site for drug and nutrient absorption. The three-dimensional (3D) topography organized in finger-like protrusions called villi increases surface area remarkably, granting a more efficient absorption process. The intestinal mucosa, where this process occurs, is a multilayered and multicell-type tissue barrier. In vitro intestinal models are routinely used to study different physiological and pathological processes in the gut, including compound absorption. Still, standard models are typically two-dimensional (2D) and represent only the epithelial barrier, lacking the cues offered by the 3D architecture and the stromal components present in vivo, often leading to inaccurate results. In this work, we studied the impact of the 3D architecture of the gut on drug transport using a bioprinted 3D model of the intestinal mucosa containing both the epithelial and the stromal compartments. Human intestinal fibroblasts were embedded in a previously optimized hydrogel bioink, and enterocytes and goblet cells were seeded on top to mimic the intestinal mucosa. The embedded fibroblasts thrived inside the hydrogel, remodeling the surrounding extracellular matrix. The epithelial cells fully covered the hydrogel scaffolds and formed a uniform cell layer with barrier properties close to in vivo. In particular, the villus-like model revealed overall increased permeability compared to a flat counterpart composed by the same hydrogel and cells. In addition, the efflux activity of the P-glycoprotein (P-gp) transporter was significantly reduced in the villus-like scaffold compared to a flat model, and the genetic expression of other drugs transporters was, in general, more relevant in the villus-like model. Globally, this study corroborates that the presence of the 3D architecture promotes a more physiological differentiation of the epithelial barrier, providing more accurate data on drug absorbance measurements.

Contributions of leaf distribution and leaf functions to photosynthesis and water-use efficiency from leaf to canopy in apple: A comparison of interstocks and cultivars.

Grafting has been widely used in horticulture to induce dwarfing and avoid stress-derived limitations on plant growth and yield by affecting plant architecture and leaf functions. However, the respective effects on plant photosynthesis and water use efficiency (WUE) of leaf distribution and functions that depend on both rootstock and scion have not been fully elucidated. This study aimed to (i) clarify the scion × interstock impacts on the variability of leaf photosynthetic traits and WUE, and (ii) decipher the respective effects of leaf distribution and functions on canopy photosynthesis and WUE (WUEc). Leaf gas exchange over light gradients and responses to light, CO2, temperature, and vapor pressure deficit were measured in two apple cultivars, 'Liquan Fuji' ('Fuji') and 'Regal Gala' ('Gala'), grafted onto rootstocks combined with interstocks: a vigorous (VV, 'Qinguan'), or a dwarf one (VD, M26). The 3D architecture-based RATP model was parameterized to estimate the canopy photosynthesis rate (Ac ), transpiration rate (E c), and WUEc. Then, virtual scenarios were used to compare the relative contributions of cultivar and interstock to canopy A c, E c, and WUE c. These scenarios changed the leaf distribution and functions of either cultivar or interstock. At the leaf scale, VD trees had significantly higher leaf nitrogen per area but a lower maximum carboxylation rate and dark respiration in both cultivars. In parallel with higher leaf stomatal conductance (gs ) and transpiration in VD 'Fuji' and similar gs in VD 'Gala', VD trees showed significantly lower leaf photosynthesis rate and WUE than VV trees. However, lower leaf photosynthetic capacities in VD trees were compensated at the canopy scale, with A c and WUE c for 'Fuji' significantly improved in VD trees under both sunny and cloudy conditions, and for 'Gala' significantly improved in VD trees under cloudy conditions compared with VV trees. Switching scenarios highlighted that 'Gala' leaf functions and distribution and VD leaf distributions enhanced A c and WUE c simultaneously, irrespective of weather conditions. Up-scaling leaf gas exchange to the canopy scale by utilizing 3D architecture-based modeling and reliable measurements of tree architecture and leaf functional traits provides insights to explore the influence of genetic materials and tree management practices.

Three-dimensional Ti3C2Tx and MnS composites as anode materials for high performance alkalis (Li, Na, K) ion batteries.

Exploring capable and universal electrode materials could promote the development of alkalis (Li, Na, K) ion batteries. 2D MXene material is an ideal host for the alkalis (Li, Na, K) ion storage, but its electrochemical performance is limited by serious re-stacking and aggregation problems. Herein, we cleverly combined electrostatic self-assembly with gas-phase vulcanization method to successfully combine Ti3C2Tx-MXene with ultra-long recyclability and high conductivity with MnS, which presents high specific capacity but poor conductivity. The as-prepared 3D hierarchical Ti3C2Tx/MnS composites have an unique sandwich-like constituent units. The tiny MnS nanoparticles are restricted between the Ti3C2Tx layers and play a key role in expanding the Ti3C2Tx interlayer spacing. As a result, the 3D Ti3C2Tx/MnS composites as the anode of LIBs exhibits a superior capacities of 826 and 634 mAh/g after 1000 and 3000 cycles at 0.5 and 1.0 A/g, respectively. More importantly, we reveal the reaction mechanism that the specific capacity first increases and then gradually stabilizes with the increase of charge and discharge cycle times when the as-prepared 3D Ti3C2Tx/MnS was used as the anode of LIBs. In addition, we have also used this material in SIBs and PIBs and achieved remarkable electrochemical capability, with a specific capacity of 107 mAh/g after 2500 cycles at 0.5 A/g or 127 mAh/g after the 2000th cycle at 0.2 A/g, respectively.

CeO2-x quantum dots decorated nitrogen-doped hollow porous carbon for supercapacitors.

The pseudocapacitive properties of CeO2 are largely dependent on its surface Faradaic redox reaction kinetics; however, its electrochemical performance is still limited by the low utilization due to the inefficient diffusionfreeways and the limited active sites. Herein, we prepare a 0D/3D composite composed of oxygen-deficient CeO2 quantum dots (0D) anchored on a 3D hollow porous N-doped carbon framework (CeO2-x QD@PHC) via a facile template-confined strategy followed by a chemical co-precipitation. The refined QDs and hollow structure greatly shorten the ion diffusion paths and lower the internal strain during cycling. The integration of CeO2-x QDs with PHC structure endows enriched accessible active sites and enhances the electrical properties. As a result, the optimized CeO2-x QD@PHC exhibits an improved specific capacitance and good rate performance in comparison to those of the CeO2-x-free PHC. Moreover, a symmetric supercapacitor with CeO2-x QD@PHC as an electrode is constructed, delivering a high energy density of 3.874 Wh kg-1 at a power density of 149.98 W kg-1.

Bifunctional iron molybdate as highly effective heterogeneous electro-Fenton catalyst and Li-ion battery anode.

Three-dimensional materials have attracted considerable interest in energy and environmental remediation fields. Iron molybdate (FMO) materials have prepared via a facile hydrothermal technique with glycerol assistance, and their structural and chemical composition confirmed using various physico-chemical techniques. The prepared bi-functional material is a strong candidate for energy storage and electrocatalytic degradation of Methylene blue and Congo red. Experimental results confirmed the synthesized FMO-10 catalyst was extremely efficient for methylene blue and Congo red breakdown, achieving 91 % and 96 % degradation in 36 h, respectively. This high catalytic activity was attributed to FMO significant visible light absorption, and reactive OH formation from H2O2 synergistically triggered by both Fe3+ and MoO42-. Prepared FMO samples demonstrated excellent potential as negative electrode material for lithium ion batteries. Electrode specific capacity initially dropped then rebounded to 1265 mAh g-1 after 100 cycles at 100 mA g-1 change rate between 0.01 and 3.0 V.

V(D)J Recombination: Recent Insights in Formation of the Recombinase Complex and Recruitment of DNA Repair Machinery.

V(D)J recombination is an essential mechanism of the adaptive immune system, producing a diverse set of antigen receptors in developing lymphocytes via regulated double strand DNA break and subsequent repair. DNA cleavage is initiated by the recombinase complex, consisting of lymphocyte specific proteins RAG1 and RAG2, while the repair phase is completed by classical non-homologous end joining (NHEJ). Many of the individual steps of this process have been well described and new research has increased the scale to understand the mechanisms of initiation and intermediate stages of the pathway. In this review we discuss 1) the regulatory functions of RAGs, 2) recruitment of RAGs to the site of recombination and formation of a paired complex, 3) the transition from a post-cleavage complex containing RAGs and cleaved DNA ends to the NHEJ repair phase, and 4) the potential redundant roles of certain factors in repairing the break. Regulatory (non-core) domains of RAGs are not necessary for catalytic activity, but likely influence recruitment and stabilization through interaction with modified histones and conformational changes. To form long range paired complexes, recent studies have found evidence in support of large scale chromosomal contraction through various factors to utilize diverse gene segments. Following the paired cleavage event, four broken DNA ends must now make a regulated transition to the repair phase, which can be controlled by dynamic conformational changes and post-translational modification of the factors involved. Additionally, we examine the overlapping roles of certain NHEJ factors which allows for prevention of genomic instability due to incomplete repair in the absence of one, but are lethal in combined knockouts. To conclude, we focus on the importance of understanding the detail of these processes in regards to off-target recombination or deficiency-mediated clinical manifestations.

Three-dimensionally visualized rhizoid system of moss, Physcomitrium patens, by refraction-contrast X-ray micro-computed tomography.

Land plants have two types of shoot-supporting systems, root system and rhizoid system, in vascular plants and bryophytes. However, since the evolutionary origin of the systems is different, how much they exploit common systems or distinct systems to architect their structures is largely unknown. To understand the regulatory mechanism of how bryophytes architect the rhizoid system responding to environmental factors, we have developed the methodology to visualize and quantitatively analyze the rhizoid system of the moss, Physcomitrium patens, in 3D. The rhizoids having a diameter of 21.3 µm on the average were visualized by refraction-contrast X-ray micro-computed tomography using coherent X-ray optics available at synchrotron radiation facility SPring-8. Three types of shape (ring-shape, line and black circle) observed in tomographic slices of specimens embedded in paraffin were confirmed to be the rhizoids by optical and electron microscopy. Comprehensive automatic segmentation of the rhizoids, which appeared in three different form types in tomograms, was tested by a method using a Canny edge detector or machine learning. The accuracy of output images was evaluated by comparing with the manually segmented ground truth images using measures such as F1 score and Intersection over Union, revealing that the automatic segmentation using machine learning was more effective than that using the Canny edge detector. Thus, machine learning-based skeletonized 3D model revealed quite dense distribution of rhizoids. We successfully visualized the moss rhizoid system in 3D for the first time.