bluuebunny/crossref_metadata_2025 huggingface dataset:
inDrops-2: a flexible, versatile and cost-efficient droplet microfluidics approach for high-throughput scRNA-seq of fresh and preserved clinical samples
Authors: Juzenas Simonas , Kiseliovas Vaidotas , Goda Karolis , Zvirblyte Justina , Quintinal-Villalonga Alvaro , Nainys Juozas , Mazutis Linas
Published: September 2023

The development of a large variety of single-cell analytical techniques has empowered researchers to explore diverse biological questions at the level of individual cells. Among these, droplet-based single-cell RNA sequencing (scRNA-seq) methods have been particularly prevalent owing to their high-throughput capabilities and reduced reaction volumes. While commercial systems have contributed to the widespread adoption of droplet-based scRNA-seq, the relatively high cost impose limitations for profiling large numbers of samples. Moreover, as the scope and scale of single cell sequencing methods keeps expanding, the possibility to accommodate diverse molecular biology workflows and inexpensively profile multiple biospecimens simultaneously becomes highly relevant. Herein, we present inDrops-2: an open-source scRNA-seq platform designed to profile fresh or preserved clinical samples with a sensitivity matching that of state-of-the-art commercial systems, yet at a few folds lower cost. Using inDrops-2, we conducted a comparative analysis of two prominent scRNA-seq protocols – those based on exponential and linear amplification of cDNA – and provide useful insights about the pros and cons inherited to each approach. We showcase the utility of inDrops-2 by simultaneously profiling 18 human lung carcinoma samples, all in one run, following cell preservation, long-term storage and multiplexing, to obtain a multiregional cellular profile of tumor microenvironment. The scalability, experimental flexibility and cost-efficiency offered by inDrops-2 should make it appealing for various single-cell transcriptomic studies.
Phased assembly of neo-sex chromosomes reveals extensive Y degeneration and rapid genome evolution inRumex hastatulus
Authors: Sacchi Bianca , Humphries Zoë , Kružlicová Jana , Bodláková Markéta , Pyne Cassandre , Choudhury Baharul , Gong Yunchen , Bačovský Václav , Hobza Roman , Barrett Spencer C.H. , Wright Stephen I.
Published: September 2023

Y chromosomes are thought to undergo progressive degeneration due to stepwise loss of recombination and subsequent reduction in selection efficiency. However, the timescales and evolutionary forces driving degeneration remain unclear. To investigate the evolution of sex chromosomes on multiple timescales, we generated a high-quality phased genome assembly of the massive older (<10MYA) and neo (<200,000 years) sex chromosomes in the XYY cytotype of the dioecious plantRumex hastatulusand a hermaphroditic outgroupR. salicifolius. Our assemblies, supported by fluorescence in situ hybridization, confirmed the neo-sex chromosomes were formed by two key events: an X-autosome fusion and a reciprocal translocation between the homologous autosome and the Y chromosome. The enormous sex-linked regions of the X (296 MB) and two Y chromosomes (503 MB) both evolved from large repeat-rich genomic regions with low recombination; however, the complete loss of recombination on the Y still led to over 30% gene loss and major rearrangements. In the older sex-linked region, there has been a significant increase in transposable element abundance, even into and near genes. In the neo sex-linked regions, we observed evidence of extensive rearrangements without gene degeneration and loss. Overall, we inferred significant degeneration during the first 10 million years of Y chromosome evolution but not on very short timescales. Our results indicate that even when sex chromosomes emerge from repetitive regions of already-low recombination, the complete loss of recombination on the Y chromosome still leads to a substantial increase in repetitive element content and gene degeneration.
Threshold public goods game in well mixed populations: Cooperation in density independent and density dependent cases
Authors: Károlyi Adél , Scheuring István
Published: September 2023

The threshold public goods game is one of the best-known models of nonlinear public goods dilemmas. Cooperators and defectors typically coexist in this game when the population is assumed to follow the so-called structured deme model. In this paper we develop a dynamical model of a general N-player game in which there is no deme structure: individuals interact with randomly chosen neighbours and selection occurs between randomly chosen pairs of individuals. We show that in the deterministic limit the dynamics in this model leads to the same replicator dynamics as in the structured deme model, i.e. coexistence of cooperators and defectors is typical in threshold public goods game even when the population is completely well-mixed. We extend the model to study the effect of density dependence and density fluctuation on the dynamics. We show analytically and numerically that decreasing population density increases the equilibrium frequency of cooperators till the fixation of this strategy, but below a critical density cooperators abruptly disappear from the population. Our numerical investigations show that weak density fluctuations enhance cooperation, while strong fluctuations suppress it.

Highlights

The dynamics of threshold dilemma game in a well-mixed population.

Fluctuation of density can help or suppress cooperation.

Cooperation is more pronounced at lower density.

Cooperation abruptly disappears below a critical density level.
Cell and tissue-specific glycosylation pathways informed by single-cell transcriptomics
Authors: Chrysinas Panagiotis , Venkatesan Shriramprasad , Ang Isaac , Ghosh Vishnu , Chen Changyou , Neelamegham Sriram , Gunawan Rudiyanto
Published: September 2023

While single cell studies have made significant impacts in various subfields of biology, they lag in the Glycosciences. To address this gap, we analyzed single-cell glycogene expressions in the Tabula Sapiens dataset of human tissues and cell types using a recent glycosylation-specific gene ontology (GlycoEnzOnto). At the median sequencing (count) depth, ∼40-50 out of 400 glycogenes were detected in individual cells. Upon increasing the sequencing depth, the number of detectable glycogenes saturates at ∼200 glycogenes, suggesting that the average human cell expresses about half of the glycogene repertoire. Hierarchies in glycogene and glycopathway expressions emerged from our analysis: nucleotide-sugar synthesis and transport exhibited the highest gene expressions, followed by genes for core enzymes, glycan modification and extensions, and finally terminal modifications. Interestingly, the same cell types showed variable glycopathway expressions based on their organ or tissue origin, suggesting nuanced cell- and tissue-specific glycosylation patterns. Probing deeper into the transcription factors (TFs) of glycogenes, we identified distinct groupings of TFs controlling different aspects of glycosylation: core biosynthesis, terminal modifications, etc. We present webtools to explore the interconnections across glycogenes, glycopathways, and TFs regulating glycosylation in human cell/tissue types. Overall, the study presents an overview of glycosylation across multiple human organ systems.
Cardiac Specific Overexpression of Adenylyl Cyclase 8 Reprograms the Mouse Sinoatrial Node Transcriptome
Authors: Qu Jia-Hua , Tarasov Kirill V. , Chakir Khalid , Lakatta Edward G.
Published: September 2023

A coupled-clock system intrinsic to sinoatrial node (SAN) pacemaker cells that regulates the rate and rhythm of spontaneous action potential firing, is activated by Ca2+/calmodulin-stimulated Adenylyl Cyclase (AC) types 8 and 1. Our previous work in mice with cardiac specific overexpression of human AC8 gene (TGAC8) discovered that compared to its wild-type (WT) littermates, the heart rate (HR) of TGAC8 is elevated (by about 30%, 24 hours a day, 7days a week), and that the TGAC8 heart rhythm is markedly coherent, i.e., the HR Variability (HRV) lacks complexity, similar to that associated with aging or cardiac pathology. Reprogramming of molecular mechanisms, particularly SAN transcriptomic regulation that underlies the remarkable chronic shift in HR and HRV, however, has not been delineated. We conducted deep RNA sequencing (RNA-seq) in TGAC8 and WT SANs, using Mm10plus with human ADCY8 DNA sequence as the reference genome. Utilizing multiple bioinformatic techniques, we not only profiled the expression of marker genes related to SAN functions and AC-cAMP-PKA signaling, but also discovered negatively enriched hub pathways that differed in TGAC8 vs. WT, the top three being OXPHOS, ribosome, and cardiac muscle contraction. In contrast, signaling pathways related to inositol phosphate and its metabolism were positively enriched in TGAC8. Further, we identified two transcription regulators, KDM5A and PPARGC1A, that mediate effects of TGAC8 on ribosome and mitochondria. In summary, reprogrammed transcriptional regulation increases the HR of TGAC8 at the cost of impairment of other SAN cell functions, i.e., altered ribosome and inositol phosphate signaling and its crosstalk with mitochondria. Because these cell signaling alterations are associated with cardiac aging and age-associated CVDs, the TGAC8 mouse appears to be an ideal model in which to probe for potential therapeutic targets for cardiac aging and age-associated CVDs.
PaGeSearch: A Tool for Identifying Genes within Pathways in Unannotated Genomes
Authors: Won Sohyoung , Yu Jaewoong , Kim Heebal
Published: September 2023

In biological research, the identification and comparison of genes within specific pathways across the genomes of various species are invaluable. However, annotating the entire genome is resource-intensive and sequence similarity searches often yield results that are not actually genes. To address these limitations, we introduce PaGeSearch (Pathway Gene Search), a tool designed to identify genes from predefined lists, especially those in specific pathways, within genomes. The tool employs an initial sequence similarity search to identify relevant genomic regions, followed by targeted gene prediction and neural network-based result filtering. PaGeSearch suggests the regions that are most likely the orthologs of the genes in the query and is designed to be applicable for species within five classes: mammals, fish, birds, eudicotyledons, and liliopsida. When compared with GeMoMa, PaGeSearch consistently outperformed in terms of sensitivity and positive predictive value. Although its performance showed increased variability when applied to actual biological pathways, it nonetheless maintained an acceptable level of accuracy.
imply: improving cell-type deconvolution accuracy using personalized reference profiles
Authors: Meng Guanqun , Pan Yue , Tang Wen , Zhang Lijun , Cui Ying , Schumacher Fredrick R. , Wang Ming , Wang Rui , He Sijia , Krischer Jeffrey , Li Qian , Feng Hao
Published: September 2023

Real-world clinical samples are often admixtures of signal mosaics from multiple pure cell types. Using computational tools, bulk transcriptomics can be deconvoluted to solve for the abundance of constituent cell types. However, existing deconvolution methods are conditioned on the assumption that the whole study population is served by a single reference panel, which ignores person-to-person heterogeneity. Here we presentimply, a novel algorithm to deconvolute cell type proportions using personalized reference panels.implycan borrow information across repeatedly measured samples for each subject, and obtain precise cell type proportion estimations. Simulation studies demonstrate reduced bias in cell type abundance estimation compared with existing methods. Real data analyses on large longitudinal consortia show more realistic deconvolution results that align with biological facts. Our results suggest that disparities in cell type proportions are associated with several disease phenotypes in type 1 diabetes and Parkin-son’s disease. Our proposed toolimplyis available through the R/Bioconductor packageISLETathttps://bioconductor.org/packages/ISLET/.
Hierarchical VAEs provide a normative account of motion processing in the primate brain
Authors: Vafaii Hadi , Yates Jacob L. , Butts Daniel A.
Published: September 2023

The relationship between perception and inference, as postulated by Helmholtz in the 19th century, is paralleled in modern machine learning by generative models like Variational Autoencoders (VAEs) and their hierarchical variants. Here, we evaluate the role of hierarchical inference and its alignment with brain function in the domain of motion perception. We first introduce a novel synthetic data framework, Retinal Optic Flow Learning (ROFL), which enables control over motion statistics and their causes. We then present a new hierarchical VAE and test it against alternative models on two downstream tasks: (i) predicting ground truth causes of retinal optic flow (e.g., self-motion); and (ii) predicting the responses of neurons in the motion processing pathway of primates. We manipulate the model architectures (hierarchical versus non-hierarchical), loss functions, and the causal structure of the motion stimuli. We find that hierarchical latent structure in the model leads to several improvements. First, it improves the linear decodability of ground truth factors and does so in a sparse and disentangled manner. Second, our hierarchical VAE outperforms previous state-of-the-art models in predicting neuronal responses and exhibits sparse latent-to-neuron relationships. These results depend on the causal structure of the world, indicating that alignment between brains and artificial neural networks depends not only on architecture but also on matching ecologically relevant stimulus statistics. Taken together, our results suggest that hierarchical Bayesian inference underlines the brain’s understanding of the world, and hierarchical VAEs can effectively model this understanding.
Unifying evolutionary dynamics: a set theory exploration of symmetry and interaction
Authors: Berkemeier Francisco , Page Karen M.
Published: September 2023

Within the expansive landscape of evolutionary dynamics, symmetry features embedded in well-established models significantly influence the interpretation of individual interaction patterns. Such symmetries are determined through interaction kernel functions, which serve as mathematical models for characterizing the complexity of interactions between individuals, each with distinct phenotypes. By incorporating analytical tools from logic and set theory, we aim to provide a deeper understanding of these functions, relevant to mechanisms of evolution. We prove that the kernels introduced in Champagnat et al.’s unifying framework exist provided birth and death rates are symmetric with respect to non-focal traits. The kernels may nevertheless be highly challenging to construct, thereby indicating a complex underlying mathematical infrastructure within unified evolutionary dynamics. We show how interaction kernels for asymmetric frameworks arising in evolutionary graph theory can be derived by incorporating individuals’ graph labels into their phenotypes. These insights invite new avenues for research, providing a fresh understanding of the interactions between individuals in broader biological contexts.
Perceptual biases, camouflage patterns, and the origin of sexual signals
Authors: Héjja-Brichard Yseult , Raymond Michel , Cuthill Innes C. , Mendelson Tamra C. , Renoult Julien P.
Published: September 2023

Natural and sexual selection can be in conflict in driving the evolution of sexual ornamentation. Sexual selection typically favors detectability to potential mates, whereas natural selection penalizes detectability to avoid predators. Considering signal efficiency in addition to detectability, however, suggests that natural and sexual selection need not be antagonistic. In the visual domain, considerable evidence supports a “processing bias” by which people prefer images that match the spatial statistics of natural scenes, likely because the brain has evolved to process such scenes efficiently. A direct but untested prediction of this bias is that background-matching camouflage can be favored by natural and sexual selection. We conducted an online experiment where we show for the first time human preference for camouflaged patterning and confirm a preference for the average image statistics of natural scenes. Because many of the underlying visual mechanisms are shared across vertebrates, our results suggest that camouflage patterns can serve as evolutionary precursors of sexual signals.
Inhibiting Glutamine Metabolism Blocks Coronavirus Replication in Mammalian Cells
Authors: Greene Kai Su , Choi Annette , Chen Matthew , Yang Nianhui , Li Ruizhi , Qiu Yijian , Lukey Michael J , Rojas Katherine S , Shen Jonathan , Wilson Kristin F , Katt William P , Whittaker Gary R , Cerione Richard A
Published: September 2023

Developing therapeutic strategies against COVID-19 has gained widespread interest given the likelihood that new viral variants will continue to emerge. Here we describe one potential therapeutic strategy which involves targeting members of the glutaminase family of mitochondrial metabolic enzymes (GLS and GLS2), which catalyze the first step in glutamine metabolism, the hydrolysis of glutamine to glutamate. We show three examples where GLS expression increases during coronavirus infection of host cells, and another in which GLS2 is upregulated. The viruses hijack the metabolic machinery responsible for glutamine metabolism to generate the building blocks for biosynthetic processes and satisfy the bioenergetic requirements demanded by the ‘glutamine addiction’ of virus-infected host cells. We demonstrate how genetic silencing of glutaminase enzymes reduces coronavirus infection and that newer members of two classes of small molecule allosteric inhibitors targeting these enzymes, designated as SU1, a pan-GLS/GLS2 inhibitor, and UP4, which is specific for GLS, block viral replication in mammalian epithelial cells. Overall, these findings highlight the importance of glutamine metabolism for coronavirus replication in human cells and show that glutaminase inhibitors can block coronavirus infection and thereby may represent a novel class of anti-viral drug candidates.

Teaser

Inhibitors targeting glutaminase enzymes block coronavirus replication and may represent a new class of anti-viral drugs.
The FW2.2/CNR protein regulates cell-to-cell communication in tomato by modulating callose deposition at plasmodesmata
Authors: Beauchet Arthur , Bollier Norbert , Grison Magali , Rofidal Valérie , Gévaudant Frédéric , Bayer Emmanuelle , Gonzalez Nathalie , Chevalier Christian
Published: September 2023

TheFW2.2gene is the founding member of theCELL NUMBER REGULATOR(CNR) gene family. More than 20 years ago,FW2.2was the first cloned gene underlying a Quantitative Trait Locus (QTL) governing fruit size/weight in tomato. However, despite this discovery, the molecular mechanisms by which FW2.2 acts as a negative regulator of cell divisions during fruit growth remain undeciphered. In the present study, we confirm that FW2.2 is a transmembrane spanning protein, whose both N- and C-terminal ends are facing the apoplast. We unexpectedly found that FW2.2 is located at plasmodesmata (PD). FW2.2 participates in the spatiotemporal regulation of callose deposition at PD via an interaction with Callose Synthases, which suggests a regulatory role in cell-to-cell communication by modulating PD transport capacity and trafficking of signaling molecules during fruit development.
Leveraging GPT-4 for Identifying Cancer Phenotypes in Electronic Health Records: A Performance Comparison between GPT-4, GPT-3.5-turbo, Flan-T5 and spaCy’s Rule-based & Machine Learning-based methods
Authors: Bhattarai Kriti , Oh Inez Y. , Sierra Jonathan Moran , Tang Jonathan , Payne Philip R.O. , Abrams Zachary B. , Lai Albert M.
Published: September 2023

Objective

Accurately identifying clinical phenotypes from Electronic Health Records (EHRs) provides additional insights into patients’ health, especially when such information is unavailable in structured data. This study evaluates the application of OpenAI’s Generative Pre-trained Transformer (GPT)-4 model to identify clinical phenotypes from EHR text in non-small cell lung cancer (NSCLC) patients. The goal was to identify disease stages, treatments and progression utilizing GPT-4, and compare its performance against GPT-3.5-turbo, Flan-T5-xl, Flan-T5-xxl, and two rule-based and machine learning-based methods, namely, scispaCy and medspaCy.

Materials and Methods

Phenotypes such as initial cancer stage, initial treatment, evidence of cancer recurrence, and affected organs during recurrence were identified from 13,646 records for 63 NSCLC patients from Washington University in St. Louis, Missouri. The performance of the GPT-4 model is evaluated against GPT-3.5-turbo, Flan-T5-xxl, Flan-T5-xl, medspaCy and scispaCy by comparing precision, recall, and micro-F1 scores.

Results

GPT-4 achieved higher F1 score, precision, and recall compared to Flan-T5-xl, Flan-T5-xxl, medspaCy and scispaCy’s models. GPT-3.5-turbo performed similarly to that of GPT-4. GPT and Flan-T5 models were not constrained by explicit rule requirements for contextual pattern recognition. SpaCy models relied on predefined patterns, leading to their suboptimal performance.

Discussion and Conclusion

GPT-4 improves clinical phenotype identification due to its robust pre-training and remarkable pattern recognition capability on the embedded tokens. It demonstrates data-driven effectiveness even with limited context in the input. While rule-based models remain useful for some tasks, GPT models offer improved contextual understanding of the text, and robust clinical phenotype extraction.
Studies of DNA ‘breathing’ by polarization-sweep single-molecule fluorescence microscopy of exciton-coupled (iCy3)2dimer-labeled DNA fork constructs
Authors: Maurer Jack , Albrecht Claire S. , Herbert Patrick , Heussman Dylan , Chang Anabel , von Hippel Peter H. , Marcus Andrew H.
Published: September 2023

Local fluctuations of the sugar-phosphate backbones and bases of DNA (often called DNA ‘breathing’) play a variety of critical roles in controlling the functional interactions of the DNA genome with the protein complexes that regulate it. Here we present a single-molecule fluorescence method that we have used to measure and characterize such conformational fluctuations at and near biologically important positions in model DNA replication fork constructs labeled with exciton-coupled cyanine [(iCy3)2] dimer probes. Previous work has shown that the constructs that we test here exhibit a broad range of spectral properties at the ensemble level, and these differences can be structurally and dynamically interpreted using our present methodology at the single-molecule level. The (iCy3)2dimer has one symmetric (+) and one anti-symmetric (–) exciton with respective transition dipole moments oriented perpendicular to one another. We excite single molecule samples using a continuous-wave linearly polarized laser with polarization direction continuously rotated at the frequency 1 MHz. The ensuing fluorescence signal is modulated as the laser polarization alternately excites the symmetric and the anti-symmetric excitons of the (iCy3)2dimer probe. Phase-sensitive detection of the modulated signal provides information about the distribution of local conformations and conformational interconversion dynamics of the (iCy3)2probe. We find that at most construct positions that we examined the (iCy3)2dimer-labeled DNA fork constructs can adopt four topologically distinct conformational macrostates. These results suggest that in addition to observing DNA breathing at and near ss-dsDNA junctions, our new methodology should be useful to determine which of these pre-existing macrostates are recognized by, bind to, and are stabilized by various genome regulatory proteins.
Human vaginal microbiota colonization is regulated by female sex hormones in a mouse model
Authors: Rahman Nuzhat , Mian Firoz , Kaushic Charu
Published: September 2023

Clinically, aLactobacillusrich vaginal microbiota (VMB) is considered optimal for reproductive outcomes, while a VMB populated by anaerobes is associated with dysbiosis and the clinical condition bacterial vaginosis (BV), which is linked to increased susceptibility to sexually transmitted infections and adverse reproductive outcomes. Mouse models that mimic eubiotic and dysbiotic VMB are currently lacking but could play a critical role in improving protective interventions. In this study, probiotic, eubiotic, and dysbiotic models were first developed in normal mice, using probiotic strainsLactobacillus rhamnosusGR-1 andLactobacillus reuteriRC-14, eubioticLactobacillus crispatus, or dysbioticGardnerella vaginalisstrains. Following a single administration,L. rhamnosusandL. reuteripersisted in the mouse vaginal tract for up to eight days,L. crispatuspersisted for up to three days, andG. vaginalispersisted for up to two days, as measured by quantitative plating assays and qPCR. Colonization ofG. vaginaliswas facilitated by the presence of mucin. Endogenous sex hormones were manipulated by either ovariectomizing (OVX) mice or administering 17β-estradiol and progesterone pellets in OVX mice. The lack of endogenous hormones in OVX mice dramatically decreased VMB bacterial load compared to normal mice. None of the exogenous bacteria includingLactobacillicould colonize OVX mice for more than 24 hours. Treatment with 17β-estradiol but not progesterone restored the endogenous microbiota and colonization withLactobacilliandG. vaginalis. Interestingly, 17β-estradiol treated mice had significantly increased levels of glycogen, compared to OVX and progesterone-treated mice. These results suggest there is a dynamic interaction between sex hormones and the VMB, which can affect bacterial diversity and the ability for a VMB to colonize.
MultipleXed Population Selection and Enrichment single nucleus RNA sequencing (XPoSE-seq) enables sample identity retention during transcriptional profiling of rare populations
Authors: Savell Katherine E. , Madangopal Rajtarun , Saravanan Padmashri , Palaganas Ryan G. , Woods Kareem D. , Thompson Drake J. , Drake Olivia R. , Brenner Megan B. , Weber Sophia J. , Van Leer Elise , Choi Jae H. , Martin Toni L. , Martin Jody C. , Steinberg Mia K. , Austin James W. , Charendoff Chloé I. , Hope Bruce T.
Published: September 2023

Single nucleus RNA-sequencing is critical in deciphering tissue heterogeneity and identifying rare populations. However, current high throughput techniques are not optimized for rare target populations and require tradeoffs in design due to feasibility. We provide a novel snRNA pipeline, MulipleXedPopulationSelection andEnrichment snRNA-sequencing (XPoSE-seq), to enable targeted snRNA-seq experiments and in-depth transcriptomic characterization of rare target populations while retaining individual sample identity.
Tracking genome evolution in single cell clones reveals the rates and features of copy number alterations generated by ongoing chromosomal instability in cancer
Authors: Guscott Molly A. , Gómez-Peregrina David , Andersen Alexander Malling , Soliman Tanya N. , Horrach Caterina Vidal , Bakker Bjorn , Johanna Spierings Diana Carolina , Wardenaar René , Foijer Floris , Serrano Cesar , Schwarz Roland , McClelland Sarah E.
Published: September 2023

Cancer genomes exhibit extensive chromosomal alterations caused by ongoing Chromosomal Instability (CIN). The ensuing cell-cell heterogeneity facilitates evolution and cancer cell plasticity that can drive therapy resistance, yet cancer CIN driver mechanisms remain essentially uncharacterised. This lack of knowledge presents an untapped opportunity to target vulnerabilities associated with ongoing CIN for therapy. Existing methods to investigate the cellular mechanisms responsible for CIN rely on laborious functional assays, or inference from genomic alteration patterns from sequencing data. Current bulk sequencing derived copy number alteration pattern signatures lack the cell-cell resolution that would reveal recent genomic alterations caused by CIN. Large-scale single cell sequencing of cancer cell populations is now emerging. However, it is not known whether the effects of selection still obscure the spectrum of genomic alterations caused by recent CIN. To address this, we employed a single-cell whole-genome sequencing (scWGS) clonal outgrowth technique, that allows us to track the real-time evolution of cancer genomes at the single-cell level. Single cancer cells surprisingly re-establish heterogeneity that matches their parental population within ∼22 generations. By comparing the features of copy number alterations at different evolutionary timepoints we reveal that some alteration types are likely under negative selection and are thus only apparent in the most recent cell divisions, and not in the parental population. In one cell line we identify a particular chromosome subject to recurrent chromosomal deletions, and validated that this chromosome wasinvolved frequently in mis-segregation events during anaphase using fluorescenceIn-Situhybridisation.
Highly Robust DNA Data Storage Based on Controllable GC Content and homopolymer of 64-Element Coded Tables
Authors: Yunfei Lu , Xuncai Zhang
Published: September 2023

In this paper, we propose a DNA storage encoding scheme based on a 64-element coding table combined with forward error correction. The method encodes the data into DNA sequences by LZW compression of the original text, adding error correction codes and scrambling codes. In the encoding process, the effects of GC content limitation and long homopolymers on DNA sequences are considered. At the same time, RS error correction code is introduced to correct the DNA sequence to improve the accuracy of decoding. Finally, the feasibility and effectiveness of the program were verified by simulation experiments on Shakespeare’s sonnets. The data results show that the GC content of DNA sequences encoded by the program is kept at 50%, the homologous multimer length is not more than 2, and the original information can be recovered from the data of 10-fold sequencing depth without error with an error rate of 0.3%. We conducted simulation experiments of primer design, DNA sequence recombination, PCR amplification, and sequence reading on DNA sequences loaded with design information, which further proved the concrete feasibility of the scheme. This scheme provides a reliable and efficient encoding scheme for DNA information storage.
Requirement of microtubules for secretion of a micronemal protein CpTSP4 in the invasive stage of the apicomplexanCryptosporidium parvum
Authors: Wang Dongqiang , Jiang Peng , Wu Xiaodong , Zhang Ying , Wang Chenchen , Li Meng , Liu Mingxiao , Yin Jigang , Zhu Guan
Published: September 2023

The zoonoticCryptosporidium parvumis a global contributor to infantile diarrheal diseases and opportunistic infections in immunocompromised/weakened individuals. Like other apicomplexans, it possesses several specialized secretory organelles, including micronemes, rhoptry and dense granules. However, the understanding of cryptosporidial micronemal composition and secretory pathway remains limited. Here we report a new micronemal protein inC. parvum, namely thrombospondin repeat domain-containing protein-4 (CpTSP4), providing insights into these ambiguities. Immunostaining and enzyme-linked assays show that CpTSP4 is prestored in the micronemes of unexcysted sporozoites but secreted during sporozoite excystation, gliding and invasion. In excysted sporozoites, CpTSP4 is also distributed on the two central microtubules unique toCryptosporidium. The secretion and microtubular distribution could be completely blocked by selective kinesin-5 inhibitors, SB-743921 and SB-715992, resulting in the accumulation of CpTSP4 in micronemes. These support the kinesin-dependent microtubular trafficking of CpTSP4 for secretion. We also localize γ-tubulin, consistent with kinesin-dependent anterograde trafficking. Additionally, recombinant CpTSP4 displays nanomolar binding affinity to host cell surface, for which heparin acts as one of the host ligands. A novel heparin-binding motif is identified and biochemically validated for its contribution to the adhesive property of CpTSP4 by peptide competition assays and site-directed mutagenesis. These findings shed light on the mechanisms of intracellular trafficking and secretion of a cryptosporidial micronemal protein and the interaction of a TSP-family protein with host cells.

Importance

Cryptosporidium parvumis a globally distributed apicomplexan parasite infecting humans and/or animals. Like other apicomplexans, it possesses specialized secretory organelles in the zoites, in which micronemes discharge molecules to facilitate the movement and invasion of zoites. Although past and recent studies have identified several proteins in cryptosporidial micronemes, our understanding on the composition, secretory pathways and domain-ligand interactions of micronemal proteins remains limited. This study identifies a new micronemal protein, namely CpTSP4, that is discharged during excystation, gliding and invasion ofC. parvumsporozoites. The CpTSP4 secretion depends on the intracellular trafficking on the twoCryptosporidium-unique microtubes that could be blocked by kinesin-5/Eg5 inhibitors. Additionally, a novel heparin-binding motif is identified and biochemically validated, which contributes to the nanomolar binding affinity of CpTSP4 to host cells. These findings indicate that kinesin-dependent microtubular trafficking is critical to CpTSP4 secretion and heparin/heparan sulfate is one of the ligands for this micronemal protein.
Germline mutations and developmental mosaicism underlyingEGFR-mutant lung cancer
Authors: Burr Risa , Leshchiner Ignaty , Costantino Christina L , Blohmer Martin , Sundaresan Tilak , Cha Justin , Seeger Karsen , Guay Sara , Danysh Brian P , Gore Ira , Jacobs Raquel A , Slowik Kara , Utro Filippo , Rhrissorrakrai Kahn , Levovitz Chaya , Barth Jaimie L , Dubash Taronish , Chirn Brian , Parida Laxmi , Sequist Lecia V , Lennerz Jochen K , Mino-Kenudson Mari , Maheswaran Shyamala , Naxerova Kamila , Getz Gad , Haber Daniel A
Published: September 2023

While the development of multiple primary tumors in smokers with lung cancer can be attributed to carcinogen-induced field cancerization, the occurrence of multiple primary tumors in individuals withEGFR-mutant lung cancer who lack known environmental exposures remains unexplained. We identified ten patients with early-stage, resectable non-small cell lung cancer who presented with multiple anatomically distinctEGFR-mutant tumors. We analyzed the phylogenetic relationships among multiple tumors from each patient using whole exome sequencing (WES) and hypermutable poly-guanine (poly-G) repeat genotyping, as orthogonal methods for lineage tracing. In two patients, we identified germlineEGFRvariants, which confer moderately enhanced signaling when modeledin vitro. In four other patients, developmental mosaicism is supported by the poly-G lineage tracing and WES, indicating a common non-germline cell-of-origin. Thus, developmental mosaicism and germline variants define two distinct mechanisms of genetic predisposition to multipleEGFR-mutant primary tumors, with implications for understanding their etiology and clinical management.
Natriuretic Peptides Improve Classification of People at Low Risk of Atrial Fibrillation after Stroke
Authors: Cameron Alan , Arnold Markus , Katsas Georgios , Yang Jason , Quinn Terence J , Abdul-Rahim Azmil H , Campbell Ross , Docherty Kieran , De Marchis Gian Marco , Arnold Marcel , Kahles Timo , Nedeltchev Krassen , Cereda Carlo , Kaegi Georg , Bustamante Alejandro , Montaner Joan , Ntaios George , Foerch Christian , Spanaus Katharina , Von Eckardstein Arnold , Dawson Jesse , Katan Mira
Published: September 2023

Background

Prolonged cardiac monitoring (PCM) increases atrial fibrillation detection after stroke (AFDAS) but access is limited. We aimed to assess the utility of midregional pro-atrial natriuretic peptide (MR-proANP) and N-terminal pro-B-type natriuretic peptide (NT-proBNP) to identify people who are unlikely to have AFDAS and improve healthcare resource allocation for PCM..

Methods

We analysed people from the BIOSIGNAL (Biomarker Signature of Stroke Aetiology) study with ischaemic stroke, no known AF and ≥3 days cardiac monitoring. External validation was in the PRECISE (Preventing Recurrent Cardioembolic Stroke: Right Approach, Right Patient) study of 28-days cardiac monitoring after stroke.

The main outcome is no AFDAS. We assessed the discriminatory value of MR-proANP and NT-proBNP combined with clinical variables to identify people with no AFDAS. We determined the net reduction in people who would undergo PCM using the models with 15% AFDAS threshold probability.

Results

We included 621 people from BIOSIGNAL. The clinical model included age, National Institutes of Health Stroke Scale score, lipid-lowering therapy, creatinine and smoking status. The AUROC was 0.68 (95%CI 0.62-0.74) with clinical variables, which improved with log10MR-proANP (0.72,0.66-0.78;p=0.001) or log10NT-proBNP (0.71,0.65-0.77;p=0.009). Performance was similar for log10MR-proANP versus log10NT-proBNP (p=0.28).

In 239 people from PRECISE, the AUROC for clinical variables was 0.68 (0.59-0.76), which improved with log10NT-proBNP (0.73,0.65-0.82;p<0.001) or log10MR-proANP (0.79,0.72-0.86;p<0.001). Performance was better with log10MR-proANP versus log10NT-proBNP (p=0.03).

The models could reduce the number who would undergo PCM by 30% (clinical+log10MR-proANP), 27% (clinical+log10NT-proBNP) or 20% (clinical).

Conclusions

MR-proANP and NT-proBNP help classify people who are unlikely to have AFDAS and could reduce the number who need PCM by 30%.
Beneficial metabolic effects of PAHSAs depend on the gut microbiota in diet-induced obese mice
Authors: Lee Jennifer , Wellenstein Kerry , Rahnavard Ali , Nelson Andrew T. , Holter Marlena M. , Cummings Bethany , Yeliseyev Vladimir , Castoldi Angela , Clish Clary B. , Bry Lynn , Siegel Dionicio , Kahn Barbara B.
Published: September 2023

Dietary lipids play an essential role in regulating the function of the gut microbiota and gastrointestinal tract, and these luminal interactions contribute to mediating host metabolism. PAHSAs are a class of lipids with anti-diabetic and anti-inflammatory properties, but whether the gut microbiota contributes to their beneficial effects on host metabolism is unknown. Here, we report that treating high fat diet (HFD)-fed germ-free mice with PAHSAs does not improve insulin sensitivity. However, transfer of feces from PAHSA-treated, but not Vehicle-treated, chow-fed mice increases insulin-sensitivity in HFD-fed germ free mice. Thus, the gut microbiota is necessary for and can transmit the insulin-sensitizing effects of PAHSAs in HFD-fed germ-free mice. Functional analyses of the cecal metagenome and lipidome of PAHSA-treated mice identified multiple lipid species that associate with the gut commensalBacteroides thetaiotaomicron(Bt) and with insulin sensitivity resulting from PAHSA treatment.Btsupplementation in HFD-fed female mice prevented weight gain, reduced adiposity, improved glucose tolerance, fortified the colonic mucus barrier and reduced systemic inflammation versus chow-fed controls, effects that were not observed in HFD-fed male mice. Furthermore, ovariectomy partially reversed the beneficialBteffects on host metabolism, indicating a role for sex hormones in mediating probiotic effects. Altogether, these studies highlight the fact that lipids can modulate the gut microbiota resulting in improvement in host metabolism and that PAHSA-induced changes in the microbiota result in at least some of their insulin-sensitizing effects in female mice.
Enhanced spine stability and survival lead to increases in dendritic spine density as an early response to local alpha-synuclein overexpression in mouse prefrontal cortex
Authors: Bosch Peter J. , Kerr Gemma , Cole Rachel , Warwick Charles A. , Wendt Linder H , Pradeep Akash , Bagnall Emma , Aldridge Georgina M.
Published: September 2023

Lewy Body Dementias (LBD), including Parkinson’s disease dementia and Dementia with Lewy Bodies, are characterized by widespread accumulation of intracellular alpha-Synuclein protein deposits in regions beyond the brainstem, including in the cortex. Patients with LBDs develop cognitive changes, including abnormalities in executive function, attention, hallucinations, slowed processing, and cognitive fluctuations. The causes of these non-motor symptoms remain unclear; however, accumulation of alpha-Synuclein aggregates in the cortex and subsequent interference of synaptic and cellular function could contribute to psychiatric and cognitive symptoms. It is unknown how the cortex responds to local pathology in the absence of significant secondary effects of alpha-Synuclein pathology in the brainstem. To investigate this, we employed viral overexpression of human alpha-Synuclein protein targeting the mouse prefrontal cortex (PFC). We then usedin vivo2-photon microscopy to image awake head-fixed mice via an implanted chronic cranial window to assess the early consequences of alpha-Synuclein overexpression in the weeks following overexpression. We imaged apical tufts of Layer V pyramidal neurons in the PFC ofThy1-YFPtransgenic mice at 1-week intervals from 1-2 weeks before and 9 weeks following viral overexpression, allowing analysis of dynamic changes in dendritic spines. We found an increase in the relative dendritic spine density following local overexpression of alpha-Synuclein, beginning at 5 weeks post-injection, and persisting for the remainder of the study. We found that alpha-Synuclein overexpression led to an increased percentage and longevity of newly-persistent spines, without significant changes in the total density of newly formed or eliminated spines. A follow up study utilizing confocal microscopy revealed that the increased spine density is found in cortical cells within the alpha-Synuclein injection site, but negative for alpha-Synuclein phosphorylation at Serine-129, highlighting the potential for effects of dose and local circuits on spine survival. These findings have important implications for the physiological role and early pathological stages of alpha-Synuclein in the cortex.
Engineered bacteria launch and control an oncolytic virus
Authors: Singer Zakary S. , Pabón Jonathan , Huang Hsinyen , Sun William , Luo Hongsheng , Grant Kailyn Rhyah , Obi Ijeoma , Coker Courtney , Rice Charles M , Danino Tal
Published: September 2023

Abstract

The ability of bacteria and viruses to selectively replicate in tumors has led to synthetic engineering of new microbial therapies. Here we design a cooperative strategy wherebyS. typhimuriumbacteria transcribe and deliver the Senecavirus A RNA genome inside host cells, launching a potent oncolytic viral infection. “Encapsidated” by bacteria, the viral genome can further bypass circulating antiviral antibodies to reach the tumor and initiate replication and spread within immune mice. Finally, we engineer the virus to require a bacterially delivered protease to achieve virion maturation, demonstrating bacterial control over the virus. This work extends bacterially delivered therapeutics to viral genomes, and shows how a consortium of microbes can achieve a cooperative aim.
The impact of HTLV-1 expression on the 3D structure and expression of host chromatin
Authors: Yaguchi Hiroko , Melamed Anat , Ramanayake Saumya , Kiik Helen , Witkover Aviva , Bangham Charles R M
Published: September 2023

A typical HTLV-1-infected individual carries >104different HTLV-1-infected T cell clones, each with a single-copy provirus integrated in a unique genomic site. We previously showed that the HTLV-1 provirus causes aberrant transcription in the flanking host genome and, by binding the chromatin architectural protein CTCF, forms abnormal chromatin loops with the host genome. However, it remained unknown whether these effects were exerted simply by the presence of the provirus or were induced by its transcription. To answer this question, we sorted HTLV-1-infected T-cell clones into cells positive or negative for proviral plus-strand expression, and then quantified host and provirus transcription using RNA-seq and aberrant chromatin looping using quantitative chromosome conformation capture (q4C) in each cell population. We found that proviral plus-strand transcription induces aberrant transcription and splicing in the flanking genome but suppresses aberrant chromatin loop formation with the nearby host chromatin. Reducing provirus-induced host transcription with an inhibitor of transcriptional elongation allows recovery of chromatin loops in the plus-strand-expressing population. We conclude that aberrant host transcription induced by proviral expression causes temporary, reversible disruption of chromatin looping in the vicinity of the provirus.
Plasma of COVID-19 patients does not alter electrical resistance of human endothelial blood-brain barrierin vitro
Authors: Pociūtė Agnė , Kriaučiūnaitė Karolina , Kaušylė Aida , Zablockienė Birutė , Alčauskas Tadas , Jelinskaitė Augustė , Rudėnaitė Akvilė , Jančorienė Ligita , Ročka Saulius , Verkhratsky Alexei , Pivoriūnas Augustas
Published: September 2023

The pandemic of Coronavirus Disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) instigated the most serious global health crisis. Clinical presentation of COVID-19 frequently includes severe neurological and neuropsychiatric symptoms. However, it is presently unknown whether and to which extent pathological impairment of blood-brain barrier (BBB) contributes to the development of neuropathology during COVID-19 progression.

In the present study we used human induced pluripotent stem cells-derived brain endothelial cells (iBECs) to study the effects of blood plasma derived from COVID-19 patients on the BBB integrityin vitro. We also performed a comprehensive analysis of the cytokine and chemokine profiles in the plasma of COVID-19 patients, healthy and recovered individuals.

We found significantly increased levels of interferon γ-induced protein 10 kDa (IP-10), hepatocyte growth factor (HGF), and interleukin-18 (IL-18) in the plasma of COVID-19 patients. However, blood plasma from COVID-19 patients did not affect transendothelial electrical resistance (TEER) in iBEC monolayers.

Our results demonstrate that COVID-19-associated blood plasma inflammatory factors do not impair BBB integrity directly and suggest that pathological remodelling of BBB during COVID-19 may occur through indirect mechanisms.