bluuebunny/crossref_metadata_2025 huggingface dataset:
Dynamics of cell states and alternative splicing following kidney ischemia-reperfusion injury
Authors: Markiewitz Dana , Goldberger Jacob , Kalisky Tomer
Published: October 2024

The progression of kidney damage in chronic kidney disease (CKD) involves multiple post-injury stages and complex cellular and molecular mechanisms that are not yet fully understood. In our study we set to characterize the dynamics of mRNA splicing following kidney injury. To this end, we analyzed publicly available bulk RNA-seq data covering nine time points following a kidney ischemia-reperfusion injury (IRI) mouse experiment. Using topic modeling we discerned five distinct temporal phases corresponding to the following cell states: “early injury response”, “injury”, “repairing”, “failed recovery”, and “healthy proximal tubule”. Additionally, we discovered a set of genes that are alternatively spliced between selected time points associated with these cell states, some of which are related to injury, stress, EMT, and apoptosis. Finally, we found several putative splicing regulators that are differentially expressed between the different time points and whose binding motifs are enriched in the vicinity of alternatively spliced exons, indicating that they may play critical roles in mRNA splicing dynamics following kidney injury and repair. These findings enhance our understanding of the molecular mechanisms involved in kidney injury and repair, offering potential avenues for developing targeted therapeutic strategies for acute kidney injury (AKI) and its progression to CKD.
NLGN3 contributes to angiogenesis in myocardial infarction via activation of the Gαi1/3-Akt Pathway
Authors: Qiao Shunsong , Tang Chao , Zhu Jingjing , Cao Cong , Feng Yu , Gu Xiaosong
Published: October 2024

BACKGROUND

Angiogenesis is an important repair mechanism for myocardial infarction. Neuroligin-3 (NLGN3) can promote angiogenesis by activating Gαi1/3-Akt signaling following ischemic brain injury. This study investigated the role of NLGN3 in myocardial infarction (MI).

METHODS AND RESULTS

On the 7thday after MI, the plasma level of NLGN3 in patients was significantly higher than in the control group. A mouse model of MI also showed significantly increased expression of NLGN3 in heart tissue. Single-nucleus transcriptome analysis revealed that NLGN3 was located predominantly in cardiac fibroblasts and endothelial cells (ECs). Endothelial-specific knockdown of NLGN3, or inhibition of NLGN3 using ADAM10i, significantly increased the ischemic area, reduced angiogenesis, and worsened cardiac function. Co-immunoprecipitation (Co-IP) experiments showed that NLGN3 interacted with Gαi1/3. The Gαi1/3 knockout (Gαi1/3-KO) mouse model of MI showed an increased ischemic area, decreased angiogenesis, and impaired cardiac function. Mechanistic studies showed that the NLGN3-Gαi1/3 signaling pathway exerts cardioprotective effects by promoting EC proliferation and tube formation through the PI3K-Akt-mTOR pathway. Silencing of Gαi1/3 largely eliminated the ability of NLGN3-promoting cardiac ECs to proliferate and form tubes.

CONCLUSION

Our findings suggest the endothelial NLGN3-Gαi1/3 signaling pathway promotes angiogenesis and reduces the ischemic area following MI, which is critical for maintaining cardiac function and repairing tissues. Targeting of the NLGN3-Gαi1/3 signaling pathway may have clinical therapeutic potential in protecting the heart from ischemic injury.

What Is Known?

Myocardial infarction (MI) leads to a significantly increased risk of recurrent events in individually vascularized regions, which then stimulates adverse systemic vascular effects.

Gαi proteins play a key role in receptor tyrosine kinase (RTK) and other non-GPCR receptor-mediated signaling.

Neuroligin 3 (NLGN3) is a major member of the NLGN family of proteins that help to form synapses between neurons. It is known be cleaved and secreted in an activity-dependent manner, but its function in heart disease is still poorly understood.

What New Information Does This Article Contribute?

NLGN3 expression was found to be markedly increased in the heart after MI, especially in cardiac ECs.

Knockdown of Gαi1/3 significantly reduced angiogenesis and impaired cardiac function, while knockdown of NLGN3 increased the ischemic area and reduced angiogenesis through Gαi1/3.

NLGN3-Gαi1/3 signaling promotes vascular EC proliferation and tube formation through the PI3K-Akt-mTOR pathway, thereby protecting the ischemic heart.
Complex Properties of Training Stimuli Affect Brain Alignment in a Deep Network Model of Mouse Visual Cortex
Authors: Torabian Parsa , Chen Yinghan , Ng Candace , Mihalas Stefan , Buice Michael , Bakhtiari Shahab , Tripp Bryan
Published: October 2024

Deep convolutional neural networks are important models of the visual cortex that ac-count relatively well for brain activity and are able to perform ethologically relevant functions. However, it is unknown which combination of factors, such as network ar-chitecture, training objectives, and data best align this family of models with the brain. Here we investigate the statistics of training data. We hypothesized that stimuli that are naturalistic for mice would lead to higher similarity between deep network models and activity in mouse visual cortex. We used a video-game engine to create training datasets in which we varied the naturalism of the environment, the movement statis-tics, and the optics of the modelled eye. The naturalistic environment substantially and consistently led to greater brain similarity, while the other factors had more subtle and area-specific effects. We then hypothesized that differences in brain similarity between the two environments arose due to differences in spatial frequency spectra, distribu-tions of color and orientation, and/or temporal autocorrelations. To test this, we created abstract environments, composed of cubes and spheres, that resembled the naturalis-tic and non-naturalistic environments in these respects. Contrary to our expectations, these factors accounted poorly for differences in brain similarity due to the naturalis-tic and non-naturalistic environments. This suggests that the higher brain similarities we observed after training with the naturalistic environment were due to more complex factors.
A Comparative Study on Deep Convolutional Neural Networks and Histogram Equalization Techniques for Glaucoma Detection From Fundus Images
Authors: Kulkarni Ashish , Ahmed H Shafeeq
Published: October 2024

Aims

To evaluate the performance of eight different convolutional neural network (CNN) models and histogram equalization techniques for glaucoma detection from fundus images.

Material and Methods

The study utilized the ACRIMA database, comprising 705 fundus images (396 glaucomatous and 309 normal). The CNN architectures evaluated include VGG-16, VGG-19, ResNet-50, ResNet-152, Inception v3, Xception, DenseNet-121, and EfficientNetB7. Two histogram-based preprocessing methods were applied: histogram equalization (HE) and contrast limited adaptive histogram equalization (CLAHE). The models were trained using supervised learning, with image augmentation techniques applied. Performance metrics such as accuracy, sensitivity, specificity, precision, negative predictive value, Dice Similarity Coefficient, and Area Under the Receiver Operating Characteristics Curve (AUC ROC) were used for evaluation.

Results

VGG-19 achieved the highest accuracy (97.9%) in the raw data setting, followed closely by VGG-16 (97.2%). ResNet-50 and ResNet-152 showed the highest specificity scores (98.4%). The sensitivity of the models ranged from 91.3% to 98.8%, with VGG-16 and VGG-19 demonstrating the highest values. CLAHE preprocessing resulted in improved performance, particularly for ResNet-152, which achieved an accuracy of 97.5% and sensitivity of 97.9%. The AUC ROC values varied from 0.937 to 0.996, with VGG-16 having the highest value (0.998).

Conclusion

The study highlights the importance of selecting suitable CNN architectures and preprocessing techniques in developing effective glaucoma detection systems. While VGG-19 exhibited the highest accuracy with raw data, VGG-16 and ResNet-50 offered consistent performance across different preprocessing techniques, making them reliable options for clinical applications.
Aberrant iron deposition in the multiple sclerosis spinal cord relates to neurodegeneration
Authors: Pisa Marco , Lockhart Andrew , Angell Thomas , Avery Aimee , Waldman Alex , Rodriguez Carla , Duarte Tiago , Pansieri Jonathan , Dhari Zaenab , Bailey Mary , Hametner Simon , Drakesmith Hal , Hofer Monika , Limbaeck Clara , DeLuca Gabriele C
Published: October 2024

Background

Iron accumulates in microglia-macrophages at the edge of multiple sclerosis (MS) lesions in the brain. Iron-rimmed brain lesions strongly predict disability accumulation, supporting iron metabolism is crucial in MS pathology. Little is known about iron distribution in the spinal cord.

Methods

Autopsy cervical, thoracic and lumbar spinal cord samples from 9 controls and 46 MS donors of whom a subset (n=36) had mesiofrontal motor cortical tissue available for study, were labelled and systematically assessed for iron (DAB-enhanced Turnbull), myelin (PLP), axons (Palmgren silver), microglia-macrophages (TMEM119, Iba1, CD68), astroglia (GFAP), oligodendroglia (OLIG2), acute axonal injury (B-APP, SMI-32, NPY-1R) and oxidative stress (E06). MS lesional and non-lesional areas were considered. Total non-haem iron was quantified by inductively coupled plasma optical emission spectroscopy (ICP-OES).

Results

In controls, iron predominantly localised to oligodendrocytes with total non-haem iron relating to total myelin fraction, which markedly differed in MS where iron accumulated in microglia-macrophages, subpial astrocytes, and axons in non-lesional areas. Iron laden microglia-macrophages were over-represented relative to total microglial-macrophages and displayed dysmorphic features. Iron-positive axons showed a disto-proximal gradient (highest at lumbar level) with a predilection for the corticospinal tracts. The extent of iron axon positivity related to smaller spinal cord area, lower total axonal counts, and greater oxidative stress. Iron positivity in each cellular compartment (i.e. subpial astrocytes, microglia-macrophage and axons) related to one-another and total non-haem iron correlated with axonal counts in MS. No iron-rimmed lesions were detected in the spinal cord unlike in cortical grey and subcortical white matter of the same cases where 22% and 80% of iron-rimmed lesions, respectively, were seen.

Conclusions

Despite the conspicuous absence of iron-rimmed lesions in the MS spinal cord, we demonstrate widespread aberrant iron distribution in the MS spinal cord that relates to oxidative stress and neurodegeneration independent of demyelination. The distal cord predominant and corticospinal tract specific accumulation of iron in axons mirrors the pattern of length-dependent motoric disability commonly encountered in progressive MS. These findings implicate aberrant iron accumulation as a novel, clinically relevant, feature of MS spinal cord pathology.
CD90 identifies distinct fractions of muscle stem cells with different modalities of activation and quiescence maintenance
Authors: Kheir Eyemen , Libergoli Michela , Metti Samuele , Jensen Jonas Brorson , Wang Jakob , Jessen Niels , Florio Francesca , Murganti Francesca , Di Savino Serena , Belicchi Marzia , Villa Chiara , Vissing Kristian , Verma Mayank , Giordani Lorenzo , Bonaldo Paolo , Torrente Yvan , Farup Jean , Biressi Stefano
Published: October 2024

Stem cell transition from quiescence to activation is crucial to guarantee productive tissue regeneration. Here we show that CD90 diversifies quiescent muscle stem cells (MuSCs) in murine and human muscle into two subpopulations differing in the kinetics of activation, CD90+veMuSCs exhibiting a faster exit quiescence and predominating the initial phases of regeneration compared to CD90-veMuSCs. In the absence of injury, the CD90+vefraction is primed toward activation through an active CD90-AMPK axis but is maintained in quiescence through signals from the extracellular matrix. Our studies show that Collagen VI, which is preferentially expressed by CD90+veMuSCs, binds to the Calcitonin receptor and plays a role in this context. Moreover, while the number of CD90+veand CD90-vesubpopulations is similar in healthy muscles, the CD90-vefraction predominates in the muscles of murine models of Duchenne and Ullrich congenital muscular dystrophies. These findings provide novel insights into the mechanistic determinants of MuSCs functional heterogeneity and have implications for understanding the stimulation of repair in dystrophic muscle.
Mechanical regulation of tissue flatness inMarchantia
Authors: Ferria Jordan , Fournié Carla J.A. , Jankowska Magdalena H. , Grossman Doron , Roeder Adrienne H.K. , Drevensek Stéphanie , Boudaoud Arezki
Published: October 2024

The production of flat organs involves a tight regulation of tissue expansion. During growth, the cell wall is under mechanical tension and continuous remodelling, requiring mechanisms to sense and maintain cell wall integrity (CWI). Here we studied the link between CWI sensing and flatness ofMarchantia polymorphathalli. We consideredFERONIA(FER), a CWI sensor, and assessed its role in the maintenance of thallus shape. We developed a pipeline to quantify flatness at multiple scales. We found that both abolishing and increasingFERexpression leads to loss of flatness. Based on experiments that reduce cell wall tension and partially restore wild-type cell size and thallus morphology, we ascribe loss of flatness to under- or to over-reaction of plants to mechanical stress. Altogether, our results support the concept that responses to mechanical stress contribute to the robustness of morphogenesis
Biotic interactions shape infection outcomes inArabidopsis
Authors: Mahmoudi Maryam , Almario Juliana , Hu Yiheng , Tenzer Lynn-Marie , Nieselt Kay , Kemen Eric
Published: October 2024

The plant microbiome protects plants from stresses, including pathogen attacks. However, identifying microbes that provide plant protection remains challenging in complex microbial communities. In this study, we analysed samples from naturalA. thalianapopulations, including both plants infected with the pathogenic oomyceteAlbugo laibachiiand uninfected plants, over six years. Using machine learning classification models, we achieved high accuracy in distinguishing infected and uninfected plants based on microbiome abundance. We identified 80 key taxa associated with health and disease. Among the health-associated microbes (HCom), we selected bacteria, fungi, and cercozoa that effectively reduced pathogen presence in co-inoculation assays. In comparison, disease-associated microbes (DCom) were less effective in conferring protection. Our findings highlight the complexity of plant-microbe interactions and advance our understanding of microbial roles in plant disease ecology. By integrating ecological insights with machine learning, we take a significant step towards designing robust microbial consortia that enhance plant resilience against pathogens.
Thymidine kinase-expressing yellow fever 17D reporter virus facilitates prodrug activation and bioorthogonal labelling of infected cells
Authors: Yakass Michael B. , Jansen Sander , Lemmens Viktor , Sanchez-Felipe Lorena , Neyts Johan , Luedtke Nathan W. , Quaye Osbourne , Dallmeier Kai
Published: October 2024

Tracking of viral replication and tissue tropism byin vivoimaging can help to unveil how live-attenuated vaccines such as the yellow fever 17D (17D) work, and likewise, to understand how adverse effects develop. Here we validate 17D-TK, a reporter virus derived from 17D that expresses herpes virus thymidine kinase (TK) that specifically converts nucleoside analogues such as Ganciclovir (GCV) to induce cell death, or difluoro-EdU (dF-EdU) for bioorthogonal labelling of infected cells by Click chemistry. 17D-TK induces a cytopathic effect in infected cell cultures, as well as mortality in intracranially inoculated mouse pups in a GCV dependent manner. Preferential phosphorylation of difluoro-EdU (dF-EdU) in 17D-TK infected cells allows to selectively stain cells that support 17D replication. Prospectively, 17D-TK can be used in combination with radiolabeled tracers for real-time detection and localization of sites of active viral replication in living animals using positron emission tomography (PET).
Dysregulation of FicD AMPylation causes diabetes by disrupting pancreatic endocrine homeostasis
Authors: Casey Amanda K. , Stewart Nathan M. , Zaidi Naqi , Gray Hillery F. , Fields Hazel A. , Sakurai Masahiro , Pinzon-Arteaga Carlos A. , Evers Bret M. , Wu Jun , Orth Kim
Published: October 2024

Bi-functional enzyme FicD regulates the endoplasmic reticulum chaperone BiP using AMPylation and deAMPylation during ER homeostasis and stress, respectively. Human FicD with an arginine-to-serine mutation disrupts FicD deAMPylation activity resulting in severe neonatal diabetes. We generated theFicDR371Smutation in mice to create a pre-clinical murine model for neonatal diabetes. We observed elevated BiP AMPylation levels across multiple tissues and signature markers for diabetes including glucose intolerance and reduced serum insulin levels. While the pancreas ofFicDR371Smice appeared normal at birth, adultFicDR371Smice displayed disturbed pancreatic islet organization that progressed with age.FicDR371Smice provide a preclinical mouse model for the study of UPR associated diabetes and demonstrate the essentiality of FicD for tissue resilience.
Mitochondrial cardiolipin metabolism controlled by tafazzin enables ferroptosis
Authors: Wohlfarter Yvonne , Hagenbuchner Judith , Horzum Utku , Oemer Gregor , Winter Andreas , Seifert Markus , Schwärzler Julian , Kokot Janik , Hernansanz Agustín Pablo , Juric Viktorija , Garcia Suoza Luiz Felipe , Talasz Heribert , Enríquez Domínguez José Antonio , Farhan Hesso , Adolph Timon E. , Weiss Günter , Zschocke Johannes , Keller Markus A.
Published: October 2024

Mitochondria are important producers of reactive oxygen species, which are involved in triggering ferroptosis, a lipid peroxidation driven form of cell death. Paradoxically, in the rare inherited metabolic disease Barth Syndrome, we discovered a protection from erastin-induced ferroptosis, despite intrinsically elevated mitochondrial ROS levels. The affected transacylase tafazzin, which is mutated in Barth Syndrome, is pivotal for remodeling of the dimeric phospholipid cardiolipin. They unique to mitochondria and essential for shaping their membrane functionalities. We investigated which downstream effects of the pathogenic membrane alterations are responsible for the protective effect against ferroptosis. We found that while iron metabolism, the unsaturation of membrane lipids, and the metabolic activity of the cells were modifying factors, they were not causal. However, we observed that cardiolipin abnormalities are not limited to impair only inner, but also outer mitochondrial membrane protein complexes. Specifically, they impact abundance and oligomerization of voltage-dependent anion channels (VDAC) in response to oxidative stress. We found that tafazzin deficiency via alteration of cardiolipins affects VDAC functionality, thereby modulating small molecule transport and signaling between mitochondria and the remaining cell. This is in line with a reduction of mitochondria-associated membranes (MAM) sites that are formed through VDACs and trapping ROS in mitochondria where they are unable to contribute to ferroptosis. These findings demonstrate that the mitochondrial membrane architecture impacting on subcellular small molecule distribution crucially impact on the manifestation of cell fate decisions, including ferroptosis.
Structure of human MUTYH and functional profiling of cancer-associated variants reveal an allosteric network between its [4Fe-4S] cluster cofactor and active site required for DNA repair
Authors: Trasviña-Arenas Carlos H. , Dissanayake Upeksha C. , Tamayo Nikole , Hashemian Mohammad , Lin Wen-Jen , Demir Merve , Hoyos-Gonzalez Nallely , Fisher Andrew J. , Cisneros G. Andrés , Horvath Martin P. , David Sheila S.
Published: October 2024

MUTYH is a clinically important DNA glycosylase that thwarts mutations by initiating base-excision repair at 8-oxoguanine (OG):A lesions. The roles for its [4Fe-4S] cofactor in DNA repair remain enigmatic. Functional profiling of cancer-associated variants near the [4Fe-4S] cofactor revealed that most variations abrogate both retention of the cofactor and enzyme activity. Surprisingly, R241Q and N238S retained the metal center and bound substrate DNA tightly but were completely inactive. We solved the crystal structure of human MUTYH bound to a transition state mimic and this revealed that Arg241 and Asn238 build an H-bond network connecting the [4Fe-4S] cluster to the catalytic Asp236 that mediates base excision. The structure of the bacterial MutY variant R149Q, along with molecular dynamics simulations of the human enzyme, show the cofactor functions to position and activate the catalytic Asp. These results suggest that allosteric cross-talk between the DNA binding [4Fe-4S] cofactor and the base excision site regulate DNA repair.
LSD1 inhibition attenuates targeted therapy-induced lineage plasticity inBRAFV600Ecolorectal cancer
Authors: Ladaika Christopher A. , Chakraborty Averi , Masood Ashiq , Hostetter Galen , Yi Joo Mi , O’Hagan Heather M.
Published: October 2024

BRAFactivating mutations occur in approximately 10% of metastatic colorectal cancer (CRCs) and are associated with worse prognosis due to an inferior response to standard chemotherapy. Standard of care for patients with refractory metastaticBRAFV600ECRC is treatment with BRAF and EGFR inhibitors. However, responses are not durable. Lineage plasticity to neuroendocrine cancer is an emerging mechanism of targeted therapy resistance in several cancer types.

Enteroendocrine cells (EECs), the neuroendocrine cell of the intestine, are uniquely present inBRAFV600ECRC as compared toBRAFwildtype CRC. Here, we demonstrated that combined BRAF and EGFR inhibition enriches for EECs in several models ofBRAFV600ECRC. Additionally, EECs and other secretory cell types were enriched in a subset ofBRAFV600ECRC patient samples following targeted therapy. Importantly, inhibition of the lysine demethylase LSD1 with a clinically relevant inhibitor attenuated targeted therapy-induced EEC enrichment through blocking the interaction of LSD1, CoREST2 and STAT3.

Statement of Significance

Our findings that BRAF plus EGFR inhibition induces lineage plasticity inBRAFV600ECRC represents a new paradigm for how resistance to BRAF plus EGFR inhibition occurs and our finding that LSD1 inhibition blocks lineage plasticity has the potential to improve responses to BRAF plus EGFR inhibitor therapy in patients.
Shifts in embryonic oxygen levels cue heterochrony in limb initiation
Authors: Zhu Meng , Catta-Preta Rinaldo , Lee ChangHee , Tabin Clifford
Published: October 2024

Heterochrony, or the alteration of developmental timing, is an important mechanism of evolutionary change. Avian species display synchronized growth of the forelimbs and hindlimbs, while mammalian species show delayed hindlimb development. We find that mammalian limb heterochrony is evident from the start of limb bud formation, and is associated with heterochronic expression of T-box transcription factors. This heterochronic shift is not due to changes in cis-regulatory sequences controlling T-box gene expression, but unexpectedly, is dependent upon differential oxygen levels to which avian and mammalian embryos are exposed prior to limb initiation, mediated, at least partially, by an NFKB transcription factor, cRel. Together, these results provide mechanistic understanding of an important example of developmental heterochrony and exemplify how the maternal environment regulates timing during embryonic development.
Disentangling the contributions of stress fibres and the unbundled actin meshwork to the anisotropy of cortical tension in response to cell shape
Authors: Fage F. , Asnacios S. , Pluta A. , Richert A. , Vias C. , Fouchard J. , Enslen H. , Etienne J. , Callan-Jones A. , Thery M. , Pereira D. , Asnacios A.
Published: October 2024

Many fundamental biological processes, in particular development and morphogenetic movements, involve tissue and cell deformation, as well as the generation of anisotropic mechanical stresses. They are often accompanied by the appearance of oriented contractile actomyosin structures resembling the stress fibres (SF) observedin vitro. Here, we investigate, at the single cell level, how cell shape — by itself — could control the structure and tension of the actomyosin cortex. Using a unique combination of 3D micropatterning, single peripheral SF (PSF) tension measurement, laser ablation and image analysis, we show that cell shape anisotropy, e.g. its 2D aspect ratio, is indeed sufficient to induce anisotropy of the cortical structure and tension. In particular, taking into account the experimentally measured anisotropy of the cortical meshwork, we could quantify cortical tension and decouple the contribution originating from bundled actin (oriented cortical stress fibres, CSF) and the contribution of the unbundled actin meshwork (UAM). We show that the increase of cortical tension anisotropy with the cell’s aspect ratio depends on the CSF alignment and orientation, the contribution of the isotropic mesh being independent of cell shape. Remarkably, while experimental data from single stress fibre measurements and laser ablation were analysed through different theoretical frameworks, namely that of negative pressure in nematics and hole drilling in prestressed materials, we found quantitatively the same composite material behaviour. In sum, we decipher here the very material properties of the actomyosin cortex, and its sensitivity to cell shape which is at the root of many mechanobiological processes, in particular morphogenesis.
Deep structural analysis of rhamnogalacturonan-I (RG-I) in Arabidopsis reveals new insights into pectin diversity
Authors: Zhang Liang , Vlach Jiri , Black Ian M. , Archer-Hartmann Stephanie , Heiss Christian , Azadi Parastoo , Urbanowicz Breeanna R.
Published: October 2024

Pectic polysaccharides are an integral part of primary plant cell walls, where they are perfrom crucial structural and biological functions. Pectin is generally divided into four distinct structural categories, including homogalacturonan, xylogalacturonan, rhamnogalacturonan I (RG-I) and rhamnogalacturonan II. Among the four main pectin domains, the structural intricacies of rhamnogalacturonan-I (RG-I) remain the least understood, especially in widely employed plant models. We employed multiple complementary analytical techniques to present a detailed structural analysis of RG-I in the model systemArabidopsis thaliana.Using highly purified RG-I from different tissues, we conducted comparative linkage and NMR analyses, complemented by mass spectrometry of enzymatically digested RG-I oligosaccharides. Our findings present the most comprehensive structural overview of Arabidopsis RG-I to date, revealing novel structural features. Notably, we identifiedO-acetylation of rhamnose backbone residues as a predominant feature, a modification previously unreported in this species. The combined results present a comprehensive structural overview ofArabidopsis thalianaRG-I that will serve as a roadmap for studying pectin biosynthesis and function.
Identification and characterization of UBA5 inhibitors
Authors: Singh Vigyasa , Wan Yuansong , Ahmed Tahsin Reaz , Wang Si-Yu , Samrat Subodh Kumar , Li Zhong , Sinha Subhash , Gan Li , Li Hongmin
Published: October 2024

UBA5 is a critical E1-activating enzyme in the UFMylation pathway, a post-translational modification process implicated in neurodegenerative diseases and cancers. In this study, we developed a high-throughput screening (HTS) assay to identify inhibitors of UBA5 from a library of blood-brain barrier (BBB)-permeable compounds. The assay, based on the AMP-Glo™ kit, enabled the identification of five novel UBA5 inhibitors belonging to three distinct chemical scaffolds with low micromolarIC50values. These inhibitors demonstrated selectivity for UBA5 over other E1 enzymes, such as UBA1, and showed efficacy in inhibiting endogenous UFMylation in HEK293T cells. The identified inhibitors not only provided valuable tools for studying UFMylation but also represent potential therapeutic candidates for diseases associated with dysregulated UFMylation, such as Alzheimer’s disease and cancer. Further optimization of these compounds is necessary to enhance their potency and pharmacokinetic properties. Our findings highlight UBA5 as a promising therapeutic target and pave the way for the development of selective inhibitors that can modulate UFMylation in human diseases.
Automated Harmonization and Large-Scale Integration of Heterogeneous Biomedical Sample Metadata Using Large Language Models
Authors: Higashi Koichi , Nakagawa Zenichi , Yamada Takuji , Mori Hiroshi
Published: October 2024

The exponential growth of biomedical data has created an urgent need for efficient integration and analysis of heterogeneous sample metadata across studies. However, current methods for harmonizing and standardizing these metadata are largely manual, time-consuming, and prone to inconsistencies. Here, we present a novel computational framework that leverages large language models (LLMs) to automate the harmonization and large-scale integration of diverse biomedical sample metadata. Our approach combines semantic clustering techniques with LLM-driven natural language processing to extract, interpret, and standardize metadata from various sources, including research papers, supplementary tables, and text data from public databases. We demonstrate the efficacy of our framework by applying it to thousands of human gut microbiome papers, successfully extracting and integrating metadata from over 400,000 samples. Our method achieved a 50% recovery rate of manually curated metadata, significantly outperforming traditional rule-based methods. Furthermore, our framework enabled the creation of a unified, searchable database of standardized metadata, facilitating cross-study analyses and revealing previously obscured patterns in microbiome composition across diverse populations and conditions. The scalability and adaptability of our approach suggest its potential applicability to a wide range of biomedical fields, potentially accelerating meta-analyses and fostering new insights from existing data. This work represents a significant advancement in biomedical data integration, offering a powerful tool for researchers to unlock the full potential of accumulated scientific knowledge.
C9orf72expansion creates the unstable folate-sensitive fragile site FRA9A
Authors: Mirceta Mila , Schmidt Monika H.M. , Shum Natalie , Prasolava Tanya K. , Meikle Bryanna , Lanni Stella , Mohiuddin Mohiuddin , Mckeever Paul M. , Zhang Ming , Liang Minggao , van der Werf Ilse , Scheers Stefaan , Dion Patrick A. , Wang Peixiang , Wilson Michael D. , Abell Theresa , Philips Elliot A. , Sznajder Łukasz J. , Swanson Maurice S. , Mehkary Mustafa , Khan Mahreen , Yokoi Katsuyuki , Jung Christine , de Jong Pieter J. , Freudenreich Catherine H. , McGoldrick Philip , Yuen Ryan K.C. , Abrahão Agessandro , Keith Julia , Zinman Lorne , Robertson Janice , Rogaeva Ekaterina , Rouleau Guy A. , Kooy R. Frank , Pearson Christopher E.
Published: October 2024

The hyper-unstable Chr9p21 locus, harbouring the interferon gene cluster, oncogenes andC9orf72,is linked to multiple diseases.C9orf72(GGGGCC)n expansions (C9orf72Exp) are associated with incompletely penetrant amyotrophic lateral sclerosis, frontotemporal dementia and autoimmune disorders.C9orf72Exp patients display hyperactive cGAS-STING-linked interferon immune and DNA damage responses, but the source of immuno-stimulatory or damaged DNA is unknown. Here, we showC9orf72Exp in pre-symptomatic and ALS-FTD patient cells and brains cause the folate-sensitive chromosomal fragile site, FRA9A. FRA9A centers on >33kb ofC9orf72as highly-compacted chromatin embedded in an 8.2Mb fragility zone spanning 9p21, encompassing 46 genes, making FRA9A one of the largest fragile sites.C9orf72Exp cells show chromosomal instability, heightened global- and Chr9p-enriched sister-chromatid exchanges, truncated-Chr9s, acentric-Chr9s and Chr9-containing micronuclei, providing endogenous sources of damaged and immunostimulatory DNA. Cells from oneC9orf72Exp patient contained highly-rearranged FRA9A-expressing Chr9 with Chr9-wide dysregulated gene expression. SomaticC9orf72Exp repeat instability and chromosomal fragility are sensitive to folate-deficiency. Age-dependent repeat instability, chromosomal fragility, and chromosomal instability can be transferred to CNS and peripheral tissues of transgenicC9orf72Exp mice, implicatingC9orf72Exp as the source. Our results highlight unappreciated effects ofC9orf72expansions that trigger vitamin-sensitive chromosome fragility, adding structural variations to the disease-enriched 9p21 locus, and likely elsewhere.
Effects of one-night partial sleep deprivation on perivascular space volume fraction: Findings from the Stockholm Sleepy Brain Study
Authors: Custer Rachel M. , Lynch Kirsten M. , Barisano Giuseppe , Herting Megan M. , Åkerstedt Torbjörn , Nilsonne Gustav , Ahmadi Hedyeh , Choupan Jeiran
Published: October 2024

Increased waste clearance in the brain is thought to occur most readily during late-stage sleep (stage N3). Sleep deprivation disrupts time spent in deeper sleep stages, fragmenting the clearance process. Here, we have utilized the publicly available Stockholm Sleepy Brain Study to investigate whether various sleep-related measures are associated with changes in perivascular space (PVS) volume fraction following a late-night short-sleep experiment. Our sample consisted of 60 participants divided into old (65-75 years) and young (20-30 years) age groups. We found that partial sleep deprivation was not significantly associated with major PVS changes. In the centrum semiovale, we observed an interaction between percentage of total sleep time spent in N3 and sleep deprivation status on PVS volume fraction. In the basal ganglia, we saw an interaction between N2 (both percentage of total sleep time and absolute time in minutes) and sleep deprivation status. However, the significance of these findings did not survive multiple comparisons corrections. This work highlights the need for future longitudinal studies of PVS and sleep, allowing for quantification of within-subject morphological changes occurring in PVS due to patterns of poor sleep. Our findings here provide insight on the impacts that a single night of late-night short-sleep has on the perivascular waste clearance system.

Statement of Significance

Sleep issues have been known to be associated with a variety of health risks. Recent research has shown the importance of sleep for brain clearance, with increases in brain clearance occurring during sleep. PVS seen on MRI are known to represent both influx and efflux pathways for the exchange of cerebrospinal fluid (CSF) and interstitial fluid (ISF) in the brain parenchyma, a key process underlying the waste clearance system.1Numerous recent studies have found links between sleep disruption and PVS volume changes. However, no study to date has been able to examine within-subject changes in PVS resulting from experimental manipulation of sleep. Our study represents the first within-subject study looking at the effects of partial sleep deprivation on PVS volume fraction using an automated PVS segmentation technique.
Benchmarking unsupervised methods for inferring TCR specificity
Authors: Jouannet Charline , Vantomme Hélène , Klatzmann David , Mariotti-Ferrandiz Encarnita
Published: October 2024

Identifying T cell receptor (TCR) specificity is crucial for advancing the understanding of adaptive immunity. Despite the development of computational methods to infer TCR specificity, their performance has not been thoroughly compared. We addressed this by curating a unified database of 190,670 human TCRs with known specificities for 2,313 epitopes across 121 organisms, combining data from IEDB, McPAS-TCR, and VDJdb. Nine methods for clustering TCRs based on similarity were benchmarked against this dataset. DeepTCR demonstrated the best retention, while ClusTCR, TCRMatch, and GLIPH2 excelled in cluster purity but had lower retention. DeepTCR, Levenshtein distance, and TCRdist3 generated large clusters, with DeepTCR showing high purity. Smaller, antigen-specific clusters were produced by Hamming distance, GIANA, and iSMART. GLIPH2 and DeepTCR were the most sensitive in capturing antigen-specific TCRs. This study offers a unified TCR database and a benchmark of specificity inference methods, guiding researchers in selecting appropriate tools.
Spatial quantitative metabolomics enables identification of remote and sustained ipsilateral cortical metabolic reprogramming after stroke
Authors: Wang Gangqi , van den Berg Bernard M. , Kostidis Sarantos , Pinkham Kelsey , Jacobs Marleen E. , Liesz Arthur , Giera Martin , Rabelink Ton J.
Published: October 2024

Mass spectrometry imaging (MSI) has become a cornerstone of spatial biology research. However, various factors that are intrinsic to the technology limit quantitative capacity of MSI-based spatial metabolomics and thus reliable interpretation. Here, we developed a quantitative MSI workflow (Q-MSI), based on isotopically13C-labeled yeast extract as internal standards, to overcome these pitfalls. Using brain and kidney tissue, we demonstrate that this approach allows for (absolute) quantification of hundreds of metabolites and lipids. Applying our workflow to a stroke model allowed us to not only map metabolic remodeling of the infarct and peri-infarct area over time, but also discover hitherto unnoted remote metabolic remodeling in the histologically unaffected ipsilateral sensorimotor cortex. At day 7 post-stroke, increased levels of neuroprotective lysine and reduced excitatory glutamate levels were found compared to the contralateral cortex. By day 28 post-stroke, lysine and glutamate levels had been normalized, while decreased precursor pools of UDP-GlcNAc and linoleate persisted that have previously been associated with vulnerability. Importantly, traditional normalization strategies not employing internal standards were unable to visualize these differences. Using13C-labeled yeast extracts as a normalization strategy establishes a new paradigm in quantitative MSI-based spatial metabolomics that greatly enhances reliability and interpretive strength.
B4GALT5-deficient CHO-Lec2 cells expressing human α1,4-galactosyltransferase: a glycoengineered cell model for studying Shiga toxin receptors
Authors: Mikolajczyk Krzysztof
Published: October 2024

Human α1,4-galactosyltransferase (A4galt) is a glycosyltransferase existing in humans as two isoforms, widespread A4galt (named A4G) and its rare variant with p.Q211E substitution (A4Gmut). Both isoforms produce Gb3 (Galα1-4Galβ1-4Glc-Cer) on glycosphingolipids and P1 glycotope (Galα1-4Galβ1-4GlcNAc-R) on glycoproteins, which serve as receptors for Shiga toxin types 1 and 2 (Stx1 and Stx2). Stx1 is bound by Gb3 and P1 glycotope, while Stx2 is recognized solely by Gb3. To elucidate the role of these receptors, CHO-Lec2 cells expressing human A4G and A4Gmut were modified by disrupting the hamster B4GALT5 gene using CRISPR/Cas9 technology. The B4GALT5 gene encodes β1,4-galactosyltransferase 5 (B4galt5), synthesizing lactosylceramide, the key substrate for Gb3 synthesis. Consequently, B4GALT5-deficient CHO-Lec2-expressing A4G and A4Gmut cells lacked Gb3 glycosphingolipid but retained the ability to synthesize glycoprotein-based P1 glycotope. Both B4GALT5-deficient CHO-Lec2 cells expressing A4G and A4Gmut demonstrated significantly decreased binding of Stx1B and Stx2B. However, cytotoxicity assay showed that B4GALT5-deficient CHO-Lec2 cells expressing A4G were completely resistant to Stx1 holotoxin while A4Gmut-expressing cells revealed reduced sensitivity to Stx2. The obtained in this study glycoengineered CHO-Lec2 cells provide a valuable model for studying receptors for Stxs, enabling a detailed assessment of their roles in toxin binding and cytotoxicity.
Gb3 trisaccharide-bearing exosomes as a novel neutralizer for Shiga toxin type 1
Authors: Mikołajczyk Krzysztof
Published: October 2024

Shiga toxin types 1 (Stx1) and 2 (Stx2), produced by Shiga toxin-producingEscherichia coli(STEC) andShigella dysenteriae, are key virulence factors responsible for severe foodborne diseases, such as hemorrhagic colitis and hemolytic uremic syndrome (HUS). The receptors for Stxs are Gb3 and P1 glycotope, which contain the Galα1→4Gal epitope and are synthesized by human α1,4-galactosyltransferase (A4galt). Stx-related infections pose a global public health challenge, owing to the limited therapeutic options due to the restricted use of antibiotics. Therefore, there is an urgent need to develop novel therapeutic strategies. This study proposes an innovative strategy utilizing exosomes derived from CHO-Lec2 cells, which were modified with Functional-Spacer-Lipid (FSL) conjugates bearing the Gb3 carbohydrate epitope (exo-Gb3-FSL). Flow cytometry analysis confirmed the presence of Galα1→4Gal disaccharides on exo-Gb3-FSL constructs, enabling them to bind Stx1. Moreover, using CHO-Lec2 cells evaluated the ability of exo-Gb3-FSL agents to bind Stx1 and protect these cells from Stx1-mediated cytotoxicity. For Stx1-treated CHO-Lec2 cells, increased cell survival was observed when using 25 µM exo-Gb3-FSL constructs, compared to control cells. These findings highlight the potential of exosome-based anti-Stx1 agents as promising alternatives to conventional therapies. This innovative strategy may provide novel directions for studies on Stx1 neutralization, offering a valuable strategy for the treatment of Stx-related diseases.

Highlights

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Exo-Gb3-FSL constructs hold Stx1 binding Galα1→4Gal epitope.

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Stx1 was effectively bound by exo-Gb3-FSL constructs.

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Exo-Gb3-FSL protected CHO-Lec2 cells from Stx1.

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Exo-Gb3-FSL is a promising anti-Stx1 agent.
Genomic landscape of antiviral defense systems in prokaryotes and phages
Authors: Li Jinquan , Gu Jiazheng , Xia Runyue , Li Meng
Published: October 2024

Prokaryotes and their viruses have co-evolved for billions of years, resulting in emergence of numerous antiviral defense systems. With the development of bioinformatic technologies and experimental studies, more and more novel defense systems have been discovered in the near decades. However, the origin and mechanism of these systems are still largely unknown. This study provides a systematic analysis of 132 defense systems within 212,599 prokaryotic genomes, which should be the largest analyzed data so far, revealing the diversity and distribution of these systems across different taxonomic units. Our findings also reveal that not only well-studied bacteria, but also archaea and virus encode diverse antiviral defense systems with specific features. In summary, this work classify 132 known defense systems, provides a comprehensive view of prokaryotic defense systems and insights into the evolution of immune responses.
Low Intestinal Doses of Botulinum Neurotoxins types A and B favour infection bySalmonellaandShigellawithout the flaccid paralysis of botulism
Authors: Fabris Federico , Brun Paola , Megighian Aram , Bernabè G , Castagliuolo Ignazio , Drigo Ilenia , Bano Luca , Lista Florigio , Bernardini Maria Lina , Montecucco Cesare , Rossetto Ornella
Published: October 2024

Botulism is a life-threatening disease characterized by a descending flaccid paralysis caused by a protein neurotoxin (BoNT) released by different anaerobic bacterial species of the genusClostridium. The paralysis results from blockade of neurotransmitter release from the terminals of peripheral cholinergic, skeletal and autonomic neurons exerted by BoNT through the cleavage of SNARE proteins, which are essential for neuroexocytosis. Here, we investigated the effect of different doses of BoNT serotypes A and B, the serotypes most commonly associated with human botulism, on enteric nervous system neurons which play an important role in gut health and physiology. We found that BoNT/A and BoNT/B enter cholinergic neurons where they cleave SNARE proteins even at doses that do not cause signs of flaccid neuroparalysis. However, these low BoNT doses favour the invasion and infection of the mouse body bySalmonella thyphimuriumandShigella flexneri. This may have significant animal health implications.
Spatially global effects of feature-based attention in functional subdivisions of human subcortical nuclei
Authors: Lin Weiru , Zhang Yan-Yu , Zhang Xilin , Fang Fang , Qian Chencan , Zhang Peng
Published: October 2024

Attention can prioritize the processing of non-spatial features throughout the visual field. While human subcortical nuclei are known to play important roles in spatial attention, the subcortical mechanisms of feature-based attention remain elusive. Using high-resolution 7T fMRI, we investigated the spatially global effects of color-based attention in functional subdivisions of human subcortical nuclei. Paying attention to a color matching the unattended stimulus across the visual field selectively modulated BOLD signals in the parvocellular (P) layers of the lateral geniculate nucleus (LGN) of the thalamus, enhancing the response to the unattended stimulus while reducing the response to the attended stimulus. The spatially global effects of color-based attention were also found in the deeper layers of the superior colliculus (SC), early visual cortices, and the intraparietal sulcus (IPS) of the parietal lobe. Effective connectivity analyses further revealed enhanced feedforward and feedback connectivity between LGN and V1, along with top-down modulation from IPS through the SC and ventral pulvinar. Finally, attended color can be decoded from multi-voxel response patterns in frontoparietal regions. These findings demonstrate that color-based attention selectively modulates color processing in P subdivisions of the LGN of the thalamus throughout the visual field, controlled by top-down signals from the parietal cortex through the SC and pulvinar.

Highlights

Color-based attention selectively modulates parvocellular activity in the LGN throughout the visual field.

Spatially global effect of feature-based attention significantly modulates activity in the deeper layers of the SC.

Feature-based attention enhances both feedforward and feedback connectivity between the LGN and V1.

Top-down signals from IPS through the SC and pulvinar controls feature-based attention.