Biology subjects

O'Connor, N. J.

Publications and source records attributed to O'Connor, N. J..

6 recordsLinked to original sources

bioRxiv2025

Hybrid Solid-Liquid Optics Enable Scalable, High-Resolution, Multi-Immersion Light-Sheet Microscopy

Modern biology increasingly depends on data-driven discovery, requiring scalable and affordable high-content 3D imaging across molecular to organ scales. Although tissue clearing, expansion microscopy, and light-sheet microscopy (LSM) enable subcellular-resolution imaging of intact specimens, their scalability remains fundamentally limited by detection optics: immersion objectives deliver high-resolution, aberration-free imaging but with short working distances, high cost, and multi-immersion incompatibility, while air objectives offer long working distances and portability at lower cost but suffer from severe aberrations and reduced photon collection when imaging immersed samples. We introduce the Hybrid Solid-Liquid Immersion Lens (HySIL) framework, which pairs an off-the-shelf solid optical component with a refractive index-matched liquid to precompensate aberrations and enhance resolution. Building on HySIL, we developed SCOPE and Super-SCOPE, objective-agnostic imaging devices achieving submicron lateral resolution (<0.75 {micro}m) across centimeter-scale samples using inexpensive air objectives with >30 mm working distances. Integration with a low-cost LSM platform yielded a compact, scalable system demonstrated for multi-immersion, multi-color, subcellular-resolution mapping of cleared or expanded mouse, salamander, and cavefish brains, human iPSC-derived organoids, and 3D histopathology of breast tissue. HySIL and SCOPE establish an accessible foundation for scalable, high-resolution volumetric imaging, advancing data-driven biological discovery.

bioengineering↗

bioRxiv2024

ClearScope: a fully integrated light sheet theta microscope for sub-cellular resolution imaging without lateral size constraints

Three-dimensional (3D) ex vivo imaging of cleared intact brains of animal models and large human and non-human primate postmortem brain specimens is important for understanding the physiological neural network connectivity patterns and the pathological alterations underlying neuropsychiatric and neurological disorders. Light-sheet microscopy has emerged as a highly effective imaging modality for rapid high-resolution imaging of large cleared samples. However, the orthogonal arrangements of illumination and detection optics in light sheet microscopy limits the size of specimen that can be imaged. Recently developed light sheet theta microscopy (LSTM) technology addressed this by utilizing a unique arrangement of two illumination light paths oblique to the detection light path, while allowing perpendicular arrangement of the detection light path relative to the specimen surface. Here, we report development of a next-generation, fully integrated, and user-friendly LSTM system for rapid sub-cellular resolution imaging uniformly throughout a large specimen without constraining the lateral (XY) size. In addition, we provide a seamlessly integrated workflow for image acquisition, data storage, pre- and post-processing, enhancement, and quantitative analysis. We demonstrate the system performance by high-resolution 3D imaging of intact mouse brains and human brain samples, and complete data analysis including digital neuron tracing, vessel reconstruction and design-based stereological analysis in 3D. This technically enhanced and user-friendly LSTM implementation will enable rapid quantitative mapping of molecular and cellular features of interests in diverse types of very large samples.

neuroscience↗

bioRxiv2023

Symmetry in frontal but not motor and somatosensory corticocortical and corticostriatal circuitry

Neocortex and striatum are topographically organized for sensory and motor functions. While sensory and motor areas are lateralized for touch and motor control, respectively, frontal areas are involved in decision making, where lateralization of function may be less important. This study contrasted the topographic precision of cell type-specific ipsilateral and contralateral cortical projections while varying the injection site location in transgenic mice of both sexes. While sensory cortical areas had strongly topographic outputs to ipsilateral cortex and striatum, they were weaker and not as topographically precise to contralateral targets. Motor cortex had somewhat stronger projections, but still relatively weak contralateral topography. In contrast, frontal cortical areas had high degrees of topographic similarity for both ipsilateral and contralateral projections to cortex and striatum. Corticothalamic organization is mainly ipsilateral, with weaker, more medial contralateral projections. Corticostriatal computations might integrate input outside closed basal ganglia loops using contralateral projections, enabling the two hemispheres to act as a unit to converge on one result in motor planning and decision making. Significance StatementEach cerebral hemisphere is responsible for sensation and movement of the opposite side of the body. Many axonal projections cross the midline to target contralateral areas. Crossed corticocortical, corticostriatal, and corticothalamic projections originate from much of neocortex, but how these projections vary across cortical regions and cell types is unknown. We quantify differences in the strength and targeting of ipsilateral and contralateral projections from frontal, motor, and somatosensory areas. The contralateral corticocortical and corticostriatal projections are proposed to play a larger role in frontal areas than in sensory or motor ones as a circuit basis for unifying computation across hemispheres in motor planning, while contralateral connectivity plays a smaller role in sensory and motor processing.

neuroscience↗

bioRxiv2022

Safety and biodistribution of NanoligomersTM targeting SARS-CoV-2 genome for treatment of COVID-19

As the world braces to enter its third year in the coronavirus disease 2019 (COVID-19) pandemic, the need for accessible and effective antiviral therapeutics continues to be felt globally. The recent surge of Omicron variant cases has demonstrated that vaccination and prevention alone cannot quell the spread of highly transmissible variants. A safe and nontoxic therapeutic with an adaptable design to respond to the emergence of new variants is critical for transitioning to treatment of COVID-19 as an endemic disease. Here, we present a novel compound, called SBCoV202, that specifically and tightly binds the translation initiation site of RNA-dependent RNA polymerase within the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genome, inhibiting viral replication. SBCoV202 is a Nanoligomer, a molecule that includes peptide nucleic acid sequences capable of binding viral RNA with single-base-pair specificity to accurately target the viral genome. The compound has been shown to be safe and nontoxic in mice, with favorable biodistribution, and has shown efficacy against SARS-CoV-2 in vitro. Safety and biodistribution were assessed after three separate administration methods, namely intranasal, intravenous, and intraperitoneal. Safety studies showed the Nanoligomer caused no outward distress, immunogenicity, or organ tissue damage, measured through observation of behavior and body weight, serum levels of cytokines, and histopathology of fixed tissue, respectively. SBCoV202 was evenly biodistributed throughout the body, with most tissues measuring Nanoligomer concentrations well above the compound KD of 3.37 nM. In addition to favorable availability to organs such as the lungs, lymph nodes, liver, and spleen, the compound circulated through the blood and was rapidly cleared through the renal and urinary systems. The favorable biodistribution and lack of immunogenicity and toxicity set Nanoligomers apart from other antisense therapies, while the adaptability of the nucleic acid sequence of Nanoligomers provides a defense against future emergence of drug resistance, making these molecules an attractive potential treatment for COVID-19.

bioengineering↗

bioRxiv2020

Engineering Transcriptional Interference through RNA Polymerase Processivity Control

Antisense transcription is widespread in all kingdoms of life and has been shown to influence gene expression through transcriptional interference (TI), a phenomenon in which one transcriptional process negatively influences another in cis. The processivity, or uninterrupted transcription, of an RNA Polymerase (RNAP) is closely tied to levels of antisense transcription in bacterial genomes, but its influence on TI, while likely important, is not well-characterized. Here we show that TI can be tuned through processivity control via three distinct antitermination strategies: the antibiotic bicyclomycin, phage protein Psu, and ribosome-RNAP coupling. We apply these methods toward TI and tune ribosome-RNAP coupling to produce 38-fold gene repression due to RNAP collisions. We then couple protein roadblock and RNAP collisions to design minimal genetic NAND and NOR logic gates. Together these results show the importance of processivity control for strong TI and demonstrate the potential for TI to create sophisticated switching responses.

systems biology↗

bioRxiv2019

Construction of two-input logic gates using Transcriptional Interference

Transcriptional Interference (TI) has been shown to regulate gene expression at the DNA level via different molecular mechanisms. The obstacles present on the DNA that a transcribing RNA Polymerase might encounter, e.g. a DNA-bound protein or another RNA Polymerase, can result in TI causing termination of transcription, thus reducing gene expression. However, the potential of TI as a new strategy to engineer complex gene expression modules has not been fully explored yet. Here we created a series of two-input devices using the presence of a roadblocking protein using both experimental and mathematical modeling approaches. We explore how multiple characteristics affect the response of genetic devices engineered to act like either AND, OR, or Single Input logic gates. We show that the dissociation constant of the roadblocking protein, inducer activation of promoter and operator sites, and distance between tandem promoters tune gate behavior. This work highlights the potential of rationally creating different types of genetic responses using the same transcription factors in subtly different genetic architectures.

synthetic biology↗