Understanding genome organization requires integration of DNA sequence and 3D spatial context, however, existing genome-wide methods lack either base-pair sequence resolution or direct spatial localization. Here, we describe in situ genome sequencing (IGS), a method for simultaneously sequencing and imaging genomes within intact biological samples. We applied IGS to human fibroblasts and early mouse embryos, spatially localizing thousands of genomic loci in individual nuclei. Using these data, we characterized parent-specific changes in genome structure across...

Understanding genome organization requires integration of DNA sequence and 3D spatial context, however, existing genome-wide methods lack either base-pair sequence resolution or direct spatial localization. Here, we describe in situ genome sequencing (IGS), a method for simultaneously sequencing and imaging genomes within intact biological samples. We applied IGS to human fibroblasts and early mouse embryos, spatially localizing thousands of genomic loci in individual nuclei. Using these data, we characterized parent-specific changes in genome structure across...

MarrowQuant, a digital and quantitative pathology tool was used to quantify the different compartments within the bone marrow at different conditions, mainly aplasia and aging conditions. By using our tool, we can compare the skeletal map induced by lethal irradiation(aplasia), with longitudinal quantification of the “red-to-yellow-to-red” transition over 2 months (aging) in C57BL/6 mouse femurs and tibiae.

Recent evidence suggests that groups of cells are more likely to form clinically dangerous metastatic tumors, emphasizing the importance of understanding mechanisms underlying collective behavior. The emergent collective behavior of migrating cell sheets in vitro has been shown to be disrupted in tumorigenic cells but the connection between this behavior and in vivo tumorigenicity remains unclear. We use particle image velocimetry to measure a multidimensional migration phenotype for genetically defined human breast epithelial cell lines...

Coordination of RNA abundance and transcription rates with cell size has been observed in diverse organisms and isogenic cell populations. However, how cells achieve such ‘size-scaling’ of transcription is unknown. Here we describe a genome-wide siRNA screen to find novel regulators of global RNA production in human cells. We quantify single-cell RNA production rates using metabolic pulse-labelling of RNA and subsequent high-content imaging. Our quantitative, single-cell measurements of DNA, nascent RNA, proliferating cell nuclear antigen...

Deep learning has emerged as the technique of choice for identifying hidden patterns in cell imaging data, but is often criticized as ‘black-box’. Here, we demonstrate that a generative neural network captures subtle details of cell appearance, which permit the prediction of the metastatic efficiency of patient-derived melanoma xenografts with known clinical outcomes. To probe the predictor, we used the network to generate “in-silico” cell images that amplified the critical predictive cellular features. These images...

The pairing of homologous chromosomes in meiosis I is essential for sexual reproduction and is mediated, in part, by the formation and repair of Spo11-induced DNA double strand breaks (DSBs). In budding yeast, each cell receives 150-200 DSBs, yet only a fraction go on to form crossover products. How and why the cell initially coordinates so many interactions along each chromosome is not well understood. Using a fluorescent reporter-operator system (FROS), we measure the kinetics...

The incidence of cutaneous squamous cell carcinoma (SCC) has increased in recent years, while it remains as an understudied subject. Current methods for the diagnose of skin cancer are either invasive, present a low resolution, or images that are difficult to interpret. Full-field optical coherence tomography (FF-OCT) has proven its potential as a non-invasive imaging technique of epithelial tissue. The sub-micrometer resolution offered by FF-OCT is able to show fine features such as cell nuclei,...

The incidence of cutaneous squamous cell carcinoma (SCC) has increased in recent years, while it remains as an understudied subject. Current methods for the diagnose of skin cancer are either invasive, present a low resolution, or images that are difficult to interpret. Full-field optical coherence tomography (FF-OCT) has proven its potential as a non-invasive imaging technique of epithelial tissue. The sub-micrometer resolution offered by FF-OCT is able to show fine features such as cell nuclei,...

There has been resurgence in determining the role of host metabolism in viral infection yet deciphering how the metabolic state of single cells affects viral entry and fusion remains unknown. Here, we have developed a novel assay multiplexing genetically-encoded biosensors with single virus tracking (SVT) to evaluate the influence of global metabolic processes on the success rate of virus entry in single cells. We found that cells with a lower ATP:ADP ratio prior to virus...

Genetic and chemical perturbations impact diverse cellular phenotypes, including multiple indicators of cell health. These readouts reveal toxicity and antitumorigenic effects relevant to drug discovery and personalized medicine. We present an approach to predict multiple cell health phenotypes using Cell Painting, an inexpensive and scalable image-based morphology assay. In matched CRISPR perturbations of three cancer cell lines, we collected both Cell Painting data and targeted assay readouts from seven reagents that measure specific cell health...

Tissue clearing is one of the most powerful strategies for a comprehensive analysis of disease progression. Here, we established an integrated pipeline that combines tissue clearing, 3D imaging, and machine learning and applied to a mouse tumour model of experimental lung metastasis using human lung adenocarcinoma A549 cells. This pipeline provided the spatial information of the tumour microenvironment. We further explored the role of transforming growth factor-beta (TGF-beta) in cancer metastasis. TGF-beta-stimulated cancer cells enhanced...

Oligodendrocyte precursor cells (OPCs) retain the capacity to remyelinate axons in the corpus callosum (CC) upon demyelination. However, the dynamics of OPC activation, mode of cell division, migration, and differentiation on a single cell level remain poorly understood due to the lack of longitudinal observations of individual cells within the injured brain. After inducing focal demyelination with lysophosphatidylcholin (LPC) in the CC of adult mice, we used 2-photon microscopy to follow for up to 2...

In most organisms, the number and distribution of crossovers that occur during meiosis are tightly controlled. All chromosomes must receive at least one “obligatory crossover” and crossovers are prevented from occurring near one another by “crossover interference”. However, the mechanistic basis of this phenomenon of crossover interference has remained mostly mysterious. Using quantitative super-resolution cytogenetics and mathematical modelling, we investigated crossover positioning in the Arabidopsis thaliana wild-type, an over-expressor of the conserved E3 ligase HEI10,...

During early Drosophila embryogenesis, a network of gene regulatory interactions orchestrates terminal patterning, playing a critical role in the subsequent formation of the gut. We utilized CRISPR gene editing at endogenous loci to create live reporters of transcription and light-sheet microscopy to monitor the individual components of posterior patterning network across 90 minutes prior to gastrulation. We developed a computational pipeline for fusing imaging datasets of the individual components into a common multivariable trajectory. This...

The aim of the study was to investigate relationships between ALS-related RNA-binding proteins and ALS, under different stress conditions.

We report the development of a semi-automated image analysis plug-in, MarrowQuant, which employs the open-source software QuPath, to systematically quantify multiple bone components in H&E sections in an unbiased manner. MarrowQuant discerns and quantifies the areas occupied by bone, adipocyte ghosts, hematopoietic cells and the interstitial/microvascular compartment. A separate feature, AdipoQuant, fragments adipocyte ghosts in H&E-stained sections of extramedullary adipose tissue sections to render adipocyte area and size distribution. Quantification of BM hematopoietic cellularity with...

The incidence of cutaneous squamous cell carcinoma (SCC) has increased in recent years, while it remains as an understudied subject. Current methods for the diagnose of skin cancer are either invasive, present a low resolution, or images that are difficult to interpret. Full-field optical coherence tomography (FF-OCT) has proven its potential as a non-invasive imaging technique of epithelial tissue. The sub-micrometer resolution offered by FF-OCT is able to show fine features such as cell nuclei,...

Drug resistance threatens the effective prevention and treatment of an ever-increasing range of human infections. This highlights an urgent need for new and improved drugs with novel mechanisms of action to avoid cross-resistance. Current cell-based drug screens are, however, restricted to binary live/dead readouts with no provision for mechanism of action prediction. Machine learning methods are increasingly being used to improve information extraction from imaging data. Such methods, however, work poorly with heterogeneous cellular phenotypes...

Libraries composed of 789,011 and 523,319 optically validated reference confocal micrographs of 17 and 20 EGFP fusion proteins localized at key cell organelles as landmarks in murine and human cells were generated for assignment of subcellular localization in mammalian cells. For each image of individual cells, 160 morphology and statistical features were used to train a random forests classifier to automatically assign the localization of proteins and dyes in both cell types and to analyze...

Libraries composed of 789,011 and 523,319 optically validated reference confocal micrographs of 17 and 20 EGFP fusion proteins localized at key cell organelles as landmarks in murine and human cells were generated for assignment of subcellular localization in mammalian cells. For each image of individual cells, 160 morphology and statistical features were used to train a random forests classifier to automatically assign the localization of proteins and dyes in both cell types and to analyze...

Libraries composed of 789,011 and 523,319 optically validated reference confocal micrographs of 17 and 20 EGFP fusion proteins localized at key cell organelles as landmarks in murine and human cells were generated for assignment of subcellular localization in mammalian cells. For each image of individual cells, 160 morphology and statistical features were used to train a random forests classifier to automatically assign the localization of proteins and dyes in both cell types and to analyze...

Recent evidence suggests that groups of cells are more likely to form clinically dangerous metastatic tumors, emphasizing the importance of understanding mechanisms underlying collective behavior. The emergent collective behavior of migrating cell sheets in vitro has been shown to be disrupted in tumorigenic cells but the connection between this behavior and in vivo tumorigenicity remains unclear. We use particle image velocimetry to measure a multidimensional migration phenotype for genetically defined human breast epithelial cell lines...

Recent evidence suggests that groups of cells are more likely to form clinically dangerous metastatic tumors, emphasizing the importance of understanding mechanisms underlying collective behavior. The emergent collective behavior of migrating cell sheets in vitro has been shown to be disrupted in tumorigenic cells but the connection between this behavior and in vivo tumorigenicity remains unclear. We use particle image velocimetry to measure a multidimensional migration phenotype for genetically defined human breast epithelial cell lines...

We explored the power of 3D super-resolution microscopy combined with computational classification and averaging to explore the 3D structure of the NPC in single human cells. We show that this approach can build the first integrated 3D structural map containing both central as well as peripheral NPC subunits with molecular specificity and nanoscale resolution. Our unbiased classification of over ten thousand individual NPCs indicates that the nuclear ring and the nuclear basket can adopt different...