Biology subjects

Regan, M.

Publications and source records attributed to Regan, M..

5 recordsLinked to original sources

bioRxiv2025

Host-microbe mutualism drives urea carbon salvage and acetogenesis during hibernation

2.Hibernation is a seasonal survival strategy employed by certain mammals that, through torpor use, reduces overall energy expenditure and permits long-term fasting. Although fasting solves the challenge of winter food scarcity, it also removes dietary carbon, a critical biomolecular building block. Here, we demonstrate a process of urea carbon salvage (UCS) in hibernating 13-lined ground squirrels, whereby urea carbon is reclaimed through gut microbial ureolysis and used in reductive acetogenesis to produce acetate, a short-chain fatty acid (SCFA) of major value to the host and its gut microbiota. We find that urea carbon incorporation into acetate is more efficient during hibernation than the summer active season, and that while both host and gut microbes oxidize acetate for energy supply throughout the year, the hosts ability to absorb and oxidize acetate is highest during hibernation. Metagenomic analysis of the gut microbiome indicates that genes involved in the degradation of gut mucins, an abundant endogenous nutrient, are retained during hibernation. The hydrogen disposal associated with reductive acetogenesis from urea carbon helps facilitate this mucin degradation by providing a luminal environment that sustains fermentation, thereby generating SCFAs and other metabolites usable by both the host and its gut microbes. Our findings introduce UCS as a mechanism that enables hibernating squirrels and their gut microbes to exploit two key endogenous nutrient sources - urea and mucins - in the resource-limited hibernation season. 3. SIGNIFICANCE STATEMENTWhen food becomes scarce during winter, hibernating mammals induce torpor to minimize energy demands and enable monthslong fasting. However, fasting eliminates the intake of essential nutrients such as carbon. We identified a two-step microbial-host interaction in ground squirrels - urea carbon salvage (UCS) - which counters carbon limitation by salvaging carbon from waste urea. Through activities of ureolytic and acetogenic bacteria, urea-derived CO2 is reduced by free hydrogen to form acetate, whose oxidation provides energy for gut microbes and the host. This process also helps maintain a permissive environment for fermentation of other host-derived, energy-dense compounds such as mucins. UCS broadens our understanding of host-microbe mutualism under extreme nutritional constraints and may represent a widespread adaptation among fasting mammals.

physiology↗

bioRxiv2022

Single Cell Spatial Analysis Reveals the Topology of Immunomodulatory Purinergic Signaling in Glioblastoma

Glioblastoma develops an immunosuppressive microenvironment that fosters tumorigenesis and resistance to current therapeutic strategies. Here we use multiplexed tissue imaging and single-cell RNA-sequencing to characterize the composition, spatial organization, and clinical significance of extracellular purinergic signaling in glioblastoma. We show that glioblastoma exhibit strong expression of CD39 and CD73 ectoenzymes, correlating with increased adenosine levels. Microglia are the predominant source of CD39, while CD73 is principally expressed by tumor cells, particularly in tumors with amplification of EGFR and astrocyte-like differentiation. Spatially-resolved single-cell analyses demonstrate strong spatial correlation between tumor CD73 and microglial CD39, and that their spatial proximity is associated with poor clinical outcomes. Together, this data reveals that tumor CD73 expression correlates with tumor genotype, lineage differentiation, and functional states, and that core purine regulatory enzymes expressed by neoplastic and tumor-associated myeloid cells interact to promote a distinctive adenosine-rich signaling niche and immunosuppressive microenvironment potentially amenable to therapeutic targeting.

cancer biology↗

bioRxiv2021

massNet: integrated processing and classification of spatially resolved mass spectrometry data using deep learning for rapid tumor delineation

MotivationMass spectrometry imaging (MSI) provides rich biochemical information in a label-free manner and therefore holds promise to substantially impact current practice in disease diagnosis. However, the complex nature of MSI data poses computational challenges in its analysis. The complexity of the data arises from its large size, high dimensionality, and spectral non-linearity. Preprocessing, including peak picking, has been used to reduce raw data complexity, however peak picking is sensitive to parameter selection that, perhaps prematurely, shapes the downstream analysis for tissue classification and ensuing biological interpretation. ResultsWe propose a deep learning model, massNet, that provides the desired qualities of scalability, non-linearity, and speed in MSI data analysis. This deep learning model was used, without prior preprocessing and peak picking, to classify MSI data from a mouse brain harboring a patient-derived tumor. The massNet architecture established automatically learning of predictive features, and automated methods were incorporated to identify peaks with potential for tumor delineation. The models performance was assessed using cross-validation, and the results demonstrate higher accuracy and a 174-fold gain in speed compared to the established classical machine learning method, support vector machine. Availability and ImplementationThe code is publicly available on GitHub.

bioinformatics↗

bioRxiv2020

Sequential Activation of Guide RNAs for Algorithmic Multiplexing of Cas9 Activities

SUMMARYGenetic manipulation of mammalian cells is instrumental to modern biomedical research but is currently limited by poor capabilities of sequentially controlling multiple manipulations in cells. Currently, either highly multiplexed manipulations can be delivered to populations of cells all at one time, or gene regulatory sequences can be engineered to conditionally activate a few manipulations within individual cells. Here, we provide proof-of-principle for a new system enabling multiple genetic manipulations to be executed as a preprogrammed cascade of events. The system leverages the programmability of the S. pyogenes Cas9 RNA-guided nuclease and is based on flexible arrangements of individual modules of activity. The basic module consists of an inactive single guide RNA (sgRNA) - like component that is converted to an active state through the effects of another sgRNA. Modules can be arranged to bring about an algorithmic program of genetic manipulations without the need for engineering cell type specific promoters or gene regulatory sequences. With the expanding diversity of available tools that utilize spCas9 to edit, repress or activate genes, this sgRNA-based system provides multiple levels for interfacing with host cell biology. In addition, ability of the system to progress through multiple modules from episomal plasmid DNA makes it suitable for applications sensitive to the presence of heterologous genomic DNA sequences and broadly applicable to biomedical research and mammalian cell engineering.Competing Interest StatementThe following competing interests are declared for RC, ART, HP, MSM, MR, and BJM as shareholders in Cellgorithmics, Inc., RC, HP, MSM, and BJM as cofounders of Cellgorithmics, Inc., and RC, HP, MSM, and BJM inventors on patent application number PCT/US2018/052211.View Full Text

synthetic biology↗

bioRxiv2018

Enhanced bacterial immunity and mammalian genome editing via RNA polymerase-mediated dislodging of Cas9 from double strand DNA breaks.

The ability to target the Cas9 nuclease to DNA sequences via Watson-Crick base pairing with a single guide RNA (sgRNA) has provided a dynamic tool for genome editing and an essential component of adaptive immune systems in bacteria. After generating a double strand break (DSB), Cas9 remains stably bound to it. Here we show persistent Cas9 binding blocks access to DSB by repair enzymes, reducing genome editing efficiency. Cas9 can be dislodged by translocating RNA polymerases, but only if the polymerase approaches one direction towards the Cas9-DSB complex. By exploiting these RNA polymerase-Cas9 interactions, Cas9 can be conditionally converted into a multi-turnover nuclease, mediating increased mutagenesis frequencies in mammalian cells and enhancing bacterial immunity to bacteriophages. These consequences of a stable Cas9-DSB complex provide insights into the evolution of PAM sequences and a simple method of improving selection of highly active sgRNA for genome editing.

molecular biology↗