Biology subjects

Rice, G.

Publications and source records attributed to Rice, G..

15 recordsLinked to original sources

bioRxiv2026

X-SPEAR: An Extensible Framework for Spatial Panel Evaluation and Refinement

Targeted spatial transcriptomics platforms such as Xenium require a predefined gene panel, and the genes in that panel are detected less efficiently, and to gene-specific degrees, in the spatial assay than in the single-cell reference used to choose them. We present X-SPEAR, a framework that couples DART marker selection with probe-level off-target assessment. DART learns short, interpretable Boolean gene rules and assembles several gene-disjoint rules per cell type. Its detection-aware training thins the reference to per-gene detection rates measured in a different, already completed experiment, so no Xenium data from the target tissue are needed before the panel is ordered. On held-out Xenium data from ovarian cancer, mouse brain and cervical cancer, DART substantially outperformed existing marker-selection methods at matched panel size, by 0.04-0.11 macro-F1 over Wilcoxon rank-sum, 0.04-0.08 over COSG and 0.10-0.18 over NS-Forest, and it reached Wilcoxon's best accuracy with 42-69% fewer genes. Ablations show that most of this gain comes from detection-aware training, which also improves other rank-based selectors and requires a detection profile measured with the same assay chemistry. At similar accuracy, DART's rule structure adds rules that faithfully summarise the downstream classifier, robustness to single-probe loss across 45 retinal ganglion cell subtypes, and better resolution of closely related cell states. We are applying X-SPEAR to design a Xenium panel for mouse retinal ganglion cell subtypes, and its in situ evaluation is in progress. X-SPEAR is available as an open-source Python package.

genomics↗

bioRxiv2025

Comprehensive profiling of transcription factors for reprogramming human astrocytes to neuronal cells through endogenous CRISPR-based gene activation

Neuronal loss is a hallmark of neurodegeneration and brain injury. Direct reprogramming of astrocytes into neurons has emerged as a promising approach to restore lost neurons. Comprehensive mapping and characterization of candidate astrocyte-to-neuron reprogramming factors is an essential step to realizing the potential of this strategy. Here, we established a CRISPR activation (CRISPRa)-based approach for neuronal reprogramming of primary human astrocytes. We conducted high-throughput CRISPRa screens of all human genes encoding transcription factors (TFs) to identify novel and efficient reprogramming factors. scRNA-seq characterization of top hits revealed that single TFs reprogram primary human astrocytes into multiple neuronal subtypes with distinct cell type-specific gene signatures. We demonstrate that INSM1 reprograms astrocytes to a glutamatergic neuron-like state and has broad neurogenic activity across different cell types and across human and mouse contexts. Finally, we conduct paired CRISPRa screens to identify cofactors that cooperate with INSM1 to enhance neuronal reprogramming and subtype specification, and elucidate genomic mechanisms of interaction and downstream regulators.

cell biology↗

bioRxiv2025

Streamlined Montage Cryo-Electron Tomography for Exploring the Ultrastructure of Cells and Tissues

Cryo-electron tomography reveals the architecture of biological specimens at molecular resolution, albeit with a limited field of view. Montage tomography overcomes this limitation, but existing strategies depend on complex dose distribution imaging schemes or hardware modifications which prevent widespread adoption of the method. To lower the barrier to entry, we developed a straightforward workflow that deviates minimally from conventional image acquisition and leverages our software MontageMaker to merge overlapping tilt series. The resulting montage tomograms are produced with high throughput, retain nanoscale detail despite the uneven accumulation of dose, and are easily acquired on a wide variety of biological samples. Most importantly, this approach can be implemented immediately by any lab with access to automated cryo-ET data collection.

cell biology↗

bioRxiv2025

Parallels in the Regulatory Landscape of Dimorphic Female and Male Genital Structures in Drosophila melanogaster

Understanding how morphological structures evolve via changes to their development is an ongoing pursuit in biology. Comparative approaches examine changes in the expression or function of key developmental molecules (e.g. transcription factors, signaling molecules or cellular effectors) within homologous structures, and correlate these changes with structural divergence across species, populations, the sexes, or even between different body parts within individuals. The female and male genitalia of Drosophila offer an excellent opportunity to investigate homology and trait evolution, as fruit fly genital structures are developmentally tractable and evolve rapidly. While previous work has characterized gene regulatory networks operating in the development and evolution of male genital structures in Drosophila, female pupal genitalia are comparatively understudied. Here, we traced the development of female pupal genitalia to determine when and how individual structures form. We then measured the expression patterns of 29 transcription factors in both female and male genital structures at high resolution using hybridization chain reaction and confocal microscopy. We found that these transcription factors are highly patterned in both sexes, and some serve as marker genes for distinct genital structures in females. Our results suggest that the same transcription factors may control developmental processes in female and male genitalia, and this data enables future studies that interrogate how developmental gene regulatory networks specialize and evolve in both sexes.

developmental biology↗

bioRxiv2024

Muscle growth by sarcomere divisions

The sarcomere is the elementary contractile unit of muscles. Adult muscle cells chain thousands of sarcomeres into long periodic myofibrils that attach to the skeleton. How new sarcomeres are added during muscle growth is unknown. By live imaging and high-throughput image analysis, we have now tracked sarcomeric components during Drosophila muscle development and discovered that individual sarcomeres divide along the myofibril tension axis into daughter sarcomeres. This way, new sarcomeres can be inserted into contractile and mechanically intact myofibrils. We propose that sarcomere division is triggered by tension and local sarcomere damage originating from skeletal growth and muscle contractions. Sarcomere divisions repair damaged sarcomeres, ensure their mechanical integrity and synchronise sarcomere addition with skeletal growth during animal development.

developmental biology↗

bioRxiv2024

Sox9 marks limbal stem cells and is required for asymmetric cell fate switch in the corneal epithelium

Adult tissues with high cellular turnover require a balance between stem cell renewal and differentiation, yet the mechanisms underlying this equilibrium are unclear. The cornea exhibits a polarized lateral flow of progenitors from the peripheral stem cell niche to the center; attributed to differences in cellular fate. To identify genes that are critical for regulating the asymmetric fates of limbal stem cells and their transient amplified progeny in the central cornea, we utilized an in vivo cell cycle reporter to isolate proliferating basal cells across the anterior ocular surface epithelium and performed single-cell transcriptional analysis. This strategy greatly increased the resolution and revealed distinct basal cell identities with unique expression profiles of structural genes and transcription factors. We focused on Sox9; a transcription factor implicated in stem cell regulation across various organs. Sox9 was found to be differentially expressed between limbal stem cells and their progeny in the central corneal. Lineage tracing analysis confirmed that Sox9 marks long-lived limbal stem cells and conditional deletion led to abnormal differentiation and squamous metaplasia in the central cornea. These data suggest a requirement for Sox9 for the switch to asymmetric fate and commitment toward differentiation, as transient cells exit the limbal niche. By inhibiting terminal differentiation of corneal progenitors and forcing them into perpetual symmetric divisions, we replicated the Sox9 loss-of-function phenotype. Our findings reveal an essential role for Sox9 for the spatial regulation of asymmetric fate in the corneal epithelium that is required to sustain tissue homeostasis.

developmental biology↗

bioRxiv2023

Using random forests to uncover the predictive power of distance-varying cell interactions in tumor microenvironments

Tumor microenvironments (TMEs) contain vast amounts of information on patients cancer through their cellular composition and the spatial distribution of tumor cells and immune cell populations. Exploring variations in TMEs among patients and cancer types, as well as determining the extent to which this information can predict variables such as patient survival or treatment success with emerging immunotherapies, is of great interest. Moreover, in the face of a large number of potential spatial cell interactions to consider, we often wish to identify specific interactions that are useful in making such predictions. We present an approach to achieve these goals based on summarizing spatial relationships in the TME using spatial K functions, and then applying functional data analysis and random forest models to both predict outcomes of interest and identify important spatial relationships. This approach is shown to be effective in simulation experiments. We further used the proposed approach to interrogate two real data sets of Multiplexed Ion Beam Images of TMEs in triple negative breast cancer and lung cancer patients. The methods proposed are publicly available in a companion R package funkycells. Author summarySpatial data on the tumor microenvironment (TME) are becoming more prevalent. Existing methods to interrogate such data often have several deficiencies: (1) they rely on estimating the spatial relationships among cells by examining simple counts of cells within a single radius, (2) they do not come with ways to evaluate the statistical significance of any findings, or (3) they consider multiple individual interactions resulting in overly optimistic estimates of interaction importances. Our approach, which leverages techniques in spatial statistics and uses a benchmark ensemble machine learning method addresses (1), since the K functions used encode the relative densities of cells over all radii up to a user-selected maximum radius, and (2) we have developed a custom approach based on permutation and cross-validation to evaluate the statistical significance of any findings of significant spatial interactions in the TME, (3) over potentially multiple interactions. Our approach is also freely available with an R implementation called funkycells. In the analysis of two real data sets, we have seen that the method performs well, and gives the expected results. We think this will be a robust tool to add to the toolbox for researchers looking to interrogate, what can be sometimes unwieldy, TME data.

bioinformatics↗

bioRxiv2022

Stress vesicles are induced by acute mechanical force and precede the commitment of epidermal stem cells to terminal differentiation

The skin has a pronounced ability to adapt to physical changes in the environment by exhibiting plasticity at the cellular level. Transient mechanical deformations applied to the skin are accommodated without permanent changes to tissue structure. However, sustained physical stress induces long-lasting alterations in the skin, which are mediated by shifts in the fates of epidermal stem cells. To investigate this phenomenon, we implemented two-photon intravital imaging to capture the responses of epidermal cells when an acute mechanical force is applied to the live skin. We show that mechanical stress induces the formation of intracellular vesicles in epidermal stem cells, which are filled with extracellular fluid and gradually enlarge, causing the deformation of the cell nucleus. By lineage tracing analysis we demonstrate that the degree of nuclear deformation is linked to cell fate. Utilizing a fluorescent in vivo reporter, to capture intracellular calcium dynamics, we show that mechanical force induces a sustained increase in intracellular calcium within basal epidermal stem cells. Conditional deletion of Piezo1, a mechanosensitive ion channel, alters intracellular calcium dynamics and increases the number of stress vesicles in epidermal stem cells. Using a human skin xenograft model, we show that stress vesicles are a conserved phenomenon in mammalian skin. This study uncovers stress vesicles as key manifestations of the mechanism that regulates the fate of epidermal stem cells under conditions of mechanical stress, in which Piezo1 and calcium dynamics are also involved.

cell biology↗

bioRxiv2022

TomoTwin: Generalized 3D Localization of Macromolecules in Cryo-electron Tomograms with Structural Data Mining

Cryoelectron tomography enables the visualization of cellular environments in extreme detail through the lens of a benign observer; what remains lacking however are tools to analyze the full amount of information contained within these densely packed volumes. Detailed analysis of macromolecules through subtomogram averaging requires particles to first be localized within the tomogram volume, a task complicated by several factors including a low signal to noise ratio and crowding of the cellular space. Available methods for this task suffer either from being error prone or requiring manual annotation of training data. To assist in this crucial particle picking step, we present TomoTwin: a robust, first in class general picking model for cryo-electron tomograms based on deep metric learning. By embedding tomograms in an information-rich, high-dimensional space which separates macromolecules according to their 3-dimensional structure, TomoTwin allows users to identify proteins in tomograms de novo without manually creating training data or retraining the network each time a new protein is to be located. TomoTwin is open source and available at https://github.com/MPI-Dortmund/tomotwin-cryoet.

biophysics↗

bioRxiv2021

Resolving Between Novelty and Homology in the Rapidly Evolving Phallus of Drosophila

The genitalia present some of the most rapidly evolving anatomical structures in the animal kingdom, possessing a variety of parts that can distinguish recently diverged species. In the Drosophila melanogaster group, the phallus is adorned with several processes, pointed outgrowths, that are similar in size and shape between species. However, the complex three-dimensional nature of the phallus can obscure the exact connection points of each process. Previous descriptions based upon adult morphology have primarily assigned phallic processes by their approximate positions in the phallus and have remained largely agnostic regarding their homology relationships. In the absence of clearly identified homology, it can be challenging to model when each structure first evolved. Here, we employ a comparative developmental analysis of these processes in eight members of the melanogaster species group to precisely identify the tissue from which each process forms. Our results indicate that adult phallic processes arise from three pupal primordia in all species. We found that in some cases the same primordia generate homologous structures whereas in other cases, different primordia produce phenotypically similar but remarkably non-homologous structures. This suggests that the same gene regulatory network may have been redeployed to different primordia to induce phenotypically similar traits. Our results highlight how traits diversify and can be redeployed, even at short evolutionary scales. Research HighlightBy incorporating developmental analysis, we find that genital structures previously identified as homologs are novel structures. This highlights how developmental analysis can help resolve contentious claims of homology.

evolutionary biology↗

bioRxiv2021

Differential levels of IFNα subtypes in autoimmunity and viral infection

Type I interferons are essential for host response to viral infections, while dysregulation of their response can result in autoinflammation or autoimmunity. Among IFN (alpha) responses, 13 subtypes exist that signal through the same receptor, but have been reported to have different effector functions. However, the lack of available tools for discriminating these closely related subtypes, in particular at the protein level, has restricted the study of their differential roles in disease. We developed a digital ELISA with specificity and high sensitivity for the IFN2 subtype. Application of this assay, in parallel with our previously described pan-IFN assay, allowed us to study different IFN protein responses following cellular stimulation and in diverse patient cohorts. We observed different ratios of IFN protein responses between viral infection and autoimmune patients. This analysis also revealed a small percentage of autoimmune patients with high IFN2 protein measurements but low pan-IFN measurements. Correlation with an ISG score and functional activity showed that in this small sub group of patients, IFN2 protein measurements did not reflect its biological activity. This unusual phenotype was partly explained by the presence of anti-IFN auto-antibodies in a subset of autoimmune patients. This study reports ultrasensitive assays for the study of IFN proteins in patient samples and highlights the insights that can be obtained from the use of multiple phenotypic readouts in translational and clinical studies.

immunology↗

bioRxiv2021

PDX1 directs a core developmentally and evolutionarily conserved gene program in the pancreatic islet

Pancreatic and duodenal homeobox 1 (PDX1) is crucial for pancreas organogenesis, yet the dynamic changes in PDX1 targets in mouse or human pancreas development have not been examined. We integrated the PDX1 cistrome with cell lineage-specific gene expression in both mouse and human developing pancreas. We identified a core set of developmentally and evolutionarily conserved PDX1 bound genes that reveal the broad multifaceted role of PDX1 in pancreas development. Despite the well-known, dramatic changes in PDX1 function and expression, we showed that PDX1 binding is largely stable from embryonic pancreas to adult islet. This may point towards a dual role of PDX1, activating or repressing the expression of its targets at different ages, dependent on other functionally-congruent or directly-interacting partners. Our work also suggests that PDX1 functions not only in initiating pancreas differentiation, but also as a potential keepsake of the progenitor program in the adult beta cells.

developmental biology↗

bioRxiv2020

Bi-compartmentalized stem cell organization of the corneal limbal niche

Stem cells exist in precise locations within tissues, yet how their organization supports tissue architecture and function is poorly understood. The limbus is the presumptive stem cell niche of the corneal epithelium. Here, we visualize the live limbus and track the activity of single stem cells in their native environment by 2-photon microscopy. We identify previously unknown niche compartments and show that long implicated slow-cycling cells form separate lineages in the outer limbus, with only local clonal dynamics. Instead, we find distinct stem cells in the pericorneal limbus to be required for corneal regeneration. Unbiased photolabeling captures their progeny exiting the niche, then moving centripetally in unison before undergoing terminal differentiation. This study demonstrates how a compartmentalized stem cell organization coordinates tissue regeneration.One Sentence Summary In vivo live imaging of the regenerating cornea reveals distinct stem cell activities in the limbal nicheCompeting Interest StatementThe authors have declared no competing interest.View Full Text

developmental biology↗

bioRxiv2019

The neural bases of resilient cognitive systems: Evidence of variable neuro-displacement in the semantic system.

The purpose of this study was to initiate exploration of an equally-important research goal: what are the neurocomputational mechanisms that make these cognitive systems \"well engineered\" and thus resilient across a range of performance demands and to mild levels of perturbation or even damage? We achieved this aim by investigating the neural dynamics of the semantic network with two task difficulty manipulations. We found that intrinsic resilience-related mechanisms were observed in both the domain-specific semantic representational system and the parallel executive control networks. Functional connectivity between these regions was also increased and these increases were related to better semantic task performance. Our results suggest that higher cognitive functions are made resilient by flexible, dynamic changes (variable neuro-displacement) across both domain-specific and multi-demand networks. Our findings provide strong evidence that the compensatory functional alterations in the impaired brain might reflect intrinsic mechanisms of a well-engineered neural system.

neuroscience↗

bioRxiv2019

Modular tissue-specific regulation of doublesex underpins sexually dimorphic development in Drosophila

The ability of a single genome to produce distinct and often dramatically different male and female forms is one of the wonders of animal development. In most animals, sex-specific phenotypes are shaped by interactions between a sex determination pathway and spatial patterning gene networks. In Drosophila melanogaster, most sexually dimorphic traits are controlled by sex-specific isoforms of the doublesex (dsx) transcription factor, and dsx expression is mostly limited to cells that give rise to sexually dimorphic traits. However, it is unknown how this mosaic of \"sex-naive\" and \"sex-aware\" tissues arises. Here, we characterize the cis-regulatory sequences that control dsx expression in the foreleg, which contains multiple types of sex-specific sensory organs. We find that separate modular enhancers are responsible for dsx expression in each sexually dimorphic organ. Expression of dsx in the sex comb is co-regulated by two enhancers with distinct spatial and temporal specificities that are separated by a genitalia-specific enhancer. Thus, the mosaic of sexually dimorphic and monomorphic organs depends on modular regulation of dsx transcription by dedicated cell type-specific enhancers.\n\nSummary StatementWe identify the modular cis-regulatory elements that direct expression of doublesex in sexually dimorphic structures in Drosophila legs and genitalia. This regulatory landscape provides insight into how cells obtain their sex-specific identity.

developmental biology↗