bluuebunny/crossref_metadata_2025 huggingface dataset:
Multi-reactive hydrogel nanovials for temporal control of secretion capture from antibody-secreting cells
Authors: Mellody Michael , Nakagawa Yuta , James Richard , Di Carlo Dino
Published: January 2025

Antibody discovery can benefit from techniques to screen antibody-secreting cells (ASCs) at scale for the binding and functionality of a diverse set of secreted antibodies. Previously, we demonstrated the use of cavity-containing hydrogel microparticles (nanovials) coated with a single affinity agent, biotin, to capture and identify ASCs secreting antibodies against a recombinant antigen bound to the nanovial through biotin-streptavidin linkages. However, rapidly secreted antibodies from unbound cells or cells in adjacent nanovials can cause crosstalk leading to background signal. Earlier efforts address this by localizing capture sites to the nanovial cavity, emulsifying nanovials, or short secretion times to limit secreted antibodies from binding to neighboring nanovials. Here, we demonstrate a method to functionalize nanovials with moieties that impart orthogonal reactivity, enabling conjugation of cell capture antibodies and antigens at different times. We show that by using a strained alkyne moiety to attach cell-capture antibodies via click chemistry to nanovials, we can capture cells and subsequently quantify secretions via biotin-streptavidin linkages. By delaying the loading of antigens onto the nanovials until after cell capture, we were able to ensure high purity (>95%) isolation of hybridoma secreting an antigen-specific antibody in a background of other hybridoma. This approach allows tight temporal control of the secretion measurement, which is independent of the cell loading time, and requires less convective transfer steps. Click chemistry-based coupling further improved cell loading into nanovials by 58% compared to biotin-streptavidin-biotin coupling and caused no reduction in cell viability. We demonstrate an implementation of this system to improve antigen-specific hybridoma screening, yielding an 8-fold improvement in hybridoma enrichment while maintaining a similar workflow complexity. Hybridomas on nanovials sorted into well plates regrew into colonies following sorting using standard fluorescence-activated cell sorting and maintained secretion of antigen-specific antibodies with high purity (∼90%), as validated via standard enzyme-linked immunosorbent assays. This lab-on-a-particle approach can be applied more generally to decouple cell loading, treatment, or activation, from secretion measurements for single-cell functional assays.
Chronic High Intensity Interval Training (HIIT) exercise in adolescent rat’s result in cocaine place aversion and ΔFosB induction
Authors: Hammond Nikki , Rahman Nabeel , Zhan Sam , Gold Mark S , Blum Kenneth , Quattrin Teresa , Young Yim Yun , Nestler Eric J , Thanos Panayotis K.
Published: January 2025

High-Intensity Interval Training (HIIT) is a form of exercise that has been greatly popularized over the past few years for its many health benefits. Similar to other forms of exercise, HIIT may be beneficial in the prevention of substance use behaviors; however, the extent to which HIIT can impact the reinforcing effects of drugs of abuse during adolescence has not been fully evaluated. Here, we assess the effects of HIIT during adolescence on subsequent cocaine conditioned place preference (CPP) in male Lewis rats. The HIIT exercise exposed rats ran on a treadmill for 30 minutes daily (ten three-minute cycles) for six weeks with progressive speed-increased up to 0.8 mph (21.5m/min), while the sedentary rats remained in their home cage. Following the six weeks of exercise, rats were tested for cocaine (25 mg/kg) CPP. Following completion of the behavior test ΔFosB levels were measured in the brain. Results showed that the HIIT rats showed significantly attenuated place preference (-19%) in their time spent in the cocaine-paired chamber compared to the sedentary environment rats. In addition, HIIT rats had significantly higher (65%) striatum ΔFosB levels compared to the sedentary rats. Our findings show that HIIT exercise during adolescence could be protective against cocaine abuse which may be mediated by an increase in ΔFosB. This finding has important clinical implications with respect to exercise mediated protection against substance misuse and abuse. Future studies will examine this effect in females as well as the potential underlying mechanisms.
Traveling waves enhance hippocampal-parahippocampal couplings in human episodic and working memory
Authors: Chen Xinyu , Chen Zhe Sage
Published: January 2025

Multiple brain regions in the human medial temporal lobe (MTL) are coordinated in memory processing. Traveling waves is a potential mechanism to coordinate information transfer through organizing the timing or spatiotemporal patterns of wave propagation. Based on direct human intracranial EEG recordings, we detected bidirectional hippocampal and parahippocampal traveling waves (4-10 Hz) along the posterior-anterior axis during a verbal memory task. Hippocampal traveling waves enhanced hippocampal-parahippocampal and intra-hippocampal couplings in both amplitude and phase as well as hippocampal theta phase-gamma amplitude coupling, suggesting a facilitatory role of TWs. Granger causality analysis showed asymmetric information flow, with greater predictability in the parahippocampal-to-hippocampal direction and dominant peak at the beta band (20-30 Hz). Hippocampal power and bidirectional hippocampal-parahippocampal information flow at the gamma band (35-50 Hz) showed reductions during successful memory encoding trials. These results support functional significance of frequency-specific parahippocampal-hippocampal and intra-hippocampal communications during memory encoding and retrieval.

Highlights

Bidirectional hippocampal traveling waves enhance hippocampal-parahippocampal couplings in both amplitude and phase.

Hippocampal-parahippocampal gamma coherence is greater in memory retrieval than memory encoding.

Intra-hippocampal theta phase-gamma amplitude coupling is greater in memory encoding than recall.

Hippocampal power and hippocampal-parahippocampal granger causality in gamma band reduced in successful trials.
4-Phenylbutyric acid modulates Connexin 43 expression restricting murine-β-coronavirus infectivity and virus-induced demyelination
Authors: Kasle Grishma , Sharma Madhav , Kumar Saurav , Das Pranati , Das Sarma Subhajit , Koval Michael , Mahadevan Anita , Kenyon Lawrence C. , Das Sarma Jayasri
Published: January 2025

Gap junction intercellular communication, particularly involving Connexin 43 and Connexin 47, plays a critical role in maintaining CNS homeostasis and has been implicated in Multiple Sclerosis (MS) pathology. Thus, warranting further studies in experimental animal models to understand how modulation of Cx43 expression can influence MS pathology. Intracranial infection with murine-β-coronavirus Mouse Hepatitis Virus (MHV-A59) in mice results in acute pathology characterized by high viral titers, glial activation in the brain and chronic neuroinflammatory demyelination, effectively mimicking key pathological hallmarks of MS and serving as a robust model to investigate its viral etiology. MHV-A59 infection leads to a pronounced downregulation of Cx43 during the acute phase, emphasizing its critical role in virus-induced CNS pathology. In this study, we investigated the potential ofin vivo4-phenylbutyric acid (4-PBA) administration in modulating Cx43 expression in this MHV-induced model and its consequence on chronic virus-induced demyelination. Our results reveal that 4-PBA treatment reduced acute MHV-A59 infectivity and viral spread in the brain while modulating the glial cell response, mounting host immunity. Treatment with 4-PBA effectively preserved the expression of both Cx43 and Cx47 in infected CNS cells, counteracting their infection-induced downregulation. Furthermore, MHV-A59 infection downregulated the expression of ER-resident thioredoxin family protein (ERp29), a well-known molecular chaperone of Cx43, which was rescued by 4-PBA treatment. We further validated if such downregulation of ERp29 is also evident in MS demyelinating plaques. In human MS patient-derived brain tissue, reduced Cx43 and ERp29 staining was observed in demyelinating plaques. Our studies revealed that 4-PBA treatment not only limits viral replication and spread throughout the brain but also protects the mice against severe chronic neuroinflammatory demyelination. These findings suggest that targeting Cx43 with 4-PBA holds significant therapeutic potential for addressing virus-induced neuroinflammatory demyelination and MS by preserving gap junction intercellular communication.
Resource partitioning in organosulfonate utilization by free-living heterotrophic bacteria during a North Sea microalgal bloom
Authors: Sidhu Chandni , Bartosik Daniel , Kale Vaikhari , Trautwein-Schult Anke , Becher Dörte , Schweder Thomas , Amann Rudolf I. , Teeling Hanno
Published: January 2025

Blooming microalgae (phytoplankton) release diverse organic molecules that fuel the marine pools of dissolved and particulate organic matter. A highly specialized community of heterotrophic bacteria rapidly remineralizes substantial parts of this organic matter in the sun-lit upper ocean. In particular, microalgae produce large quantities of various organosulfur compounds that can serve as carbon and sulfur sources for bacteria.

Here, we report on the analyses of a time series of previously generated 30 long-read metagenomes, 30 corresponding deeply sequenced short-read metatranscriptomes and 15 metaproteomes from 0.2-3 µm size fractions that we sampled in 2020 during a biphasic phytoplankton bloom in the German Bight (Southern North Sea). We analyzed the assembled contigs as well as 70 bacterial metagenome-assembled genomes that recruited the highest transcript numbers with respect to the utilization of methyl sulfur compounds (dimethylsulfoniopropionate (DMSP), dimethyl sulfide (DMS), dimethyl sulfone (DMSO2)), C3-sulfonates (2,3-dihydroxypropane-1-sulfonate (DHPS), 3-sulfolactate, 3-sulfopyruvate) and 2-aminoethanesulfonic acid (taurine).

We observed a pronounced resource partitioning among bacterial clades that utilize distinct organosulfur compounds, which may explain successions of these clades during the studied bloom.Alphaproteobacteriawere the most active and degraded a variety of organosulfonates via various metabolic routes. However, we also found previously underreported roles of members of theBacteroidotaandGammaproteobacteriaas efficient degraders of DMSP, DMS, and DMSO2. One striking observation was a strong preference for DMSP cleavage inBacteroidotaas opposed to DMSP demethylation inAlphaproteobacteriaand indications for a particular proficiency for taurine utilization in Ilumatobacter_A andAcidimicrobiia.

Importance

Sulfur-containing low-molecular-weight algal metabolites play an important role in overall marine carbon and sulfur fluxes. This study highlights that such compounds may play a crucial role in governing the succession of distinct bacterioplankton clades in response to phytoplankton blooms in coastal shelf areas of the temperate zone, such as the German Bight of the North Sea. WhileAlphaproteobacteriaare the most versatile and competitive degraders of dissolved organosulfur compounds during such blooms, this study repositions clades previously thought to play only a more limited role in dissolved organosulfur metabolismin situ, such asGammaproteobacteria,Bacteroidota, andAcidimicrobiia, as crucial contributors to the remineralization of organosulfur compounds in the upper ocean. This study also highlights the high level of interconnectedness of bacterial carbon and sulfur cycling during phytoplankton blooms.
cancerSimCraft: A Multi-resolution Cancer Genome Simulator with Comprehensive Ground Truth Tracking
Authors: Jin Haijing , Navin Nicholas , Chen Ken
Published: January 2025

Cancer evolution follows complex trajectories involving diverse genomic alterations and clonal dynamics, making it challenging to validate computational methods for single-cell DNA sequencing analysis. Here we present cancerSimCraft, a comprehensive framework for simulating cancer genome data at both clonal and single-cell resolution. cancerSimCraft combines deterministic rules with stochastic processes to model various genomic events including CNVs, SNVs, and WGDs. The framework enables integration of real cancer genome patterns with user-defined parameters, supporting customizable simulation designs that reflect both empirical data and theoretical models. Through systematic benchmarking, we demonstrate cancerSimCraft’s utility in evaluating computational methods under various conditions, particularly focusing on the impact of dataset size and parameter sensitivity. We further demonstrated its application in exploring clonal evolution and mutation patterns through controlledin silicoexperiments. These comprehensive simulation capabilities make cancerSimCraft a valuable resource for both computational method development and theoretical studies in cancer genomics.
Modulating Prestimulus Alpha and Beta Power with tRNS Establishes Their Causal Role in Visual Perception
Authors: Wei Jinwen , Zou Huiru , Tang Qianyuan , Yao Ziqing , Huang Gan , Liang Zhen , Zhang Li , Zhou Changsong , Zhang Zhiguo
Published: January 2025

Variability in visual perception in response to consistent stimuli is a fundamental phenomenon linked to fluctuations in prestimulus low-frequency neural oscillations—particularly in the alpha (8–13 Hz) and beta (13–30 Hz) bands—typically measured by their power in electroencephalography (EEG) signals. However, the causal role of these prestimulus alpha and beta power fluctuations in visual perception remains unestablished. In this study, we investigated whether prestimulus alpha and beta power causally affect visual perception using transcranial random noise stimulation (tRNS). In a sham-controlled, single-blind, within-subject design, 29 participants performed a visual detection task while receiving occipital tRNS. Online functional near-infrared spectroscopy (fNIRS) was used to measure cortical excitability during stimulation, and offline EEG signals were collected after stimulation. Mental fatigue was incorporated as a state-dependent factor influencing tRNS effects. Our findings demonstrate that, primarily under low fatigue states, tRNS increased cortical excitability during stimulation (indicated by increased fNIRS oxyhemoglobin amplitude), decreased subsequent prestimulus EEG alpha and beta power, and consequently reduced the visual contrast threshold (VCT), indicating enhanced visual perception. Sensitivity analysis revealed that alpha oscillations contributed more significantly to visual perception than beta oscillations under low fatigue. Additionally, the state-dependent effects of tRNS may result from different sensitivities of VCT to neural oscillations across fatigue states. These results provide causal evidence linking prestimulus alpha and beta power to visual perception and underscore the importance of considering brain states in neuromodulation research. Our study advances the understanding of the neural mechanisms underlying visual perception and suggests potential therapeutic applications targeting neural oscillations.

Significance statement

Understanding why we perceive identical visual stimuli differently is a fundamental question in neuroscience. This study provides causal evidence that prestimulus alpha and beta neural oscillations directly influence visual perception, particularly under low mental fatigue state. By using tRNS alongside fNIRS and EEG recordings, we demonstrate that modulating neural excitability can alter perceptual outcomes. Our findings highlight the importance of considering brain state—such as fatigue levels—in neuromodulation research. This work advances our understanding of the neural mechanisms underlying visual perception and opens avenues for developing targeted interventions to enhance sensory processing and cognitive functions, potentially benefiting individuals with perceptual or attentional disorders.
T-type calcium channels participate in intrinsic and synaptic activity of PKCγ neurons of the dorsal horn of the spinal cord during chronic pain
Authors: Cuculière Célia , Bourdon-Alonzeau Rachel , Rulhe Coline , Chemin Jean , François Amaury , Blanchard Marie-Pierre , Fontanaud Pierre , Deval Emmanuel , Mangoni Matteo , Lingueglia Eric , Bourinet Emmanuel , Méry Pierre-François
Published: January 2025

ABSTRACT

The disinhibition of the excitatory PKCγ interneurons plays a central role during mechanical allodynia in the dorsal horn of the spinal cord, routing harmless information to nociceptive pathways. The T-type calcium channel Cav3.2, necessary for mechanical and cold allodynia, is found in most PKCγ neurons of the spinal cord. In this study, the role of Cav3.2 in PKCγ neurons was studied after its pharmacological inhibition and its conditional deletion (KO) in Cav3.2GFP-FloxKI x PKCγ-CreERT2 x Ai14 mice in normal conditions and in the spared-nerve-injury (SNI) model of neuropathic pain. Conditional deletion of Cav3.2 increased the hind-paw basal mechanical sensitivity before surgery, and decreased mechanical pain 7 days, but not 28 days, after surgery. At the cellular level, Cav3.2 participated in the low-threshold currents of PKCγ neurons and the T-type calcium current of PKCγ neurons was decreased in KO mice as compared to wild-type (WT). This loss did not convert into proportional alterations in subthreshold properties including “rebound” potentials, suggesting the involvement of other T-type channels. Action potential kinetics and firing properties seemed similar in WT and KO mice too, but rebound potentials were diminished in the SNI model in WT but not in KO mice. In addition, the modulations of firing properties induced by T-type channel pharmacological blocker Z944 observed in WT mice were absent in KO mice and after SNI. Furthermore, the pairing of action potentials was modified after SNI in WT mice, and not in KO mice. At the synaptic level, excitatory currents were lowered 7- and 28-days after surgery, while inhibitory currents were lowered only at 28 days. These changes were not found in Cav3.2-ablated neurons. Miniature currents analysis indicated that Cav3.2 was involved in both excitatory and inhibitory synaptic transmissions at the level of PKCγ neurons. Surprisingly, Z944 did not mimic the effects of Cav3.2 ablation in PKCγ neurons, suggesting distinct and eventually opposite roles of other T-type calcium channels. Altogether, our results show that Cav3.2 is not mandatory for firing of PKCγ neurons of the dorsal horn of the spinal cord, but that it participates to the SNI-induced changes in their intrinsic and synaptic activity, including changes in their excitatory and inhibitory controls.

Graphical abstract
Macrophage migration-dependent retraction fibers and migrasomes - a structural basis for spatiotemporal cytokine secretion
Authors: Li Xichun , Anbarlou Azadeh , Maste Vrushali , Condon Nicholas D. , Brown Darren , Shen Hongyu , Hsu Hao-Ruei , Ashraf Amina , Tan Jinhui Sylvia , Wang Wanyi , Chernyavskiy Oleksandr , Lucas Richard M. , Luo Lin , Koh Yvette W. H. , Murray Rachael Z. , Lieschke Graham J. , Stow Jennifer L.
Published: January 2025

Migrating cells leave behind trails of matrix-bound retraction fibers and migrasomes, but their cell-specific physiological functions are largely unknown. We comprehensively characterize retraction fibers and migrasomes of migrating macrophages in mouse and zebrafishin vitroandin vivomodels. These structures share common and macrophage-centric components of tetraspanin enriched microdomains (TEMs) including tetraspanin 4 (TSPAN4), the integrin CD11b and the pTRAP, SCIMP; like TSPAN4, we show that SCIMP expression modulates retraction fiber and migrasome formation. In immune-activated migrating macrophages, rear-positioned recycling endosomes regulate the secretory trafficking of newly synthesized inflammatory cytokines into retraction fibers and migrasomes. Transmembrane TNF is delivered via SNARE-mediated carriers to the cell surface and to the surfaces of both retraction fibers and migrasomes, from where it can be released. Retraction fibers are thus newly identified here as cytokine secretion sites, and along with migrasomes they allow migrating macrophages to secrete more cytokine compared to stationary cells. Whereas cytokine secretion has been viewed as a process ofad hocdiffusion, instead, we reveal a new mechanism of migration-dependent, localized deposition and spatiotemporal release of bioactive cytokines from RFs and migrasomes. Cytokine deployment is thus organized in a constrained manner facilitating direct influence on local inflammatory responses.
Diverse Intracellular Trafficking of Insulin Analogs by Machine Learning-based Colocalization and Diffusion Analysis
Authors: Bleshøy Sara Vogt , Kæstel-Hansen Jacob , Nielsen Annette Juma , Mishra Narendra Kumar , Jensen Knud J. , Hatzakis Nikos S.
Published: January 2025

Insulin signaling is crucial for maintaining cellular function and systemic homeostasis, with its dysregulation leading to metabolic disorders in particular diabetes. While insulin analogs are essential in type 1 diabetes treatment, their intracellular trafficking and sorting compared to endogenous insulin remains largely unresolved at the molecular level. Understanding these are important in improving therapeutics and guiding future drug development. However, current methods rely on static imaging and bulk receptor assays conducted in non-physiological conditions, which often disrupt native insulin signaling and provide only snapshots that fail to capture the temporal dynamics of insulin trafficking and characteristic sorting pathways. Here, we directly observe insulin trafficking and intracellular sorting and compare the characteristics of Atto655 labeled recombinant human insulin (HI655) with insulin aspart (IAsp655), a clinically approved, rapid-acting insulin analog. We developed a combined approach integrating Colocalization Fingerprinting, a machine learning framework for reliable, time-resolved colocalization analysis, together with our recently developed deep learning-assisted single-particle diffusional analysis (DeepSPT). Our analysis revealed subtle, yet significant differences in intracellular behavior between IAsp655and HI655, particularly in diffusional behavior and lysosomal colocalization, highlighting the potential of our approach to decipher subtle differences in intracellular trafficking and sorting characteristics. In addition to contributing to a more detailed understanding of biology of insulin analogs and intracellular sorting, we provide a reliable machine-learning methodology to study intricate cellular processes.
Eye movements organize excitability state, information coding and network connectivity in the human hippocampus
Authors: Leszczynski Marcin , Espinal Elizabeth , Smith Elliot , Schevon Catherine , Sheth Sameer , Schroeder Charles E.
Published: January 2025

Natural vision is an active sensing process that entails frequent eye movements to sample the environment. Nonetheless vision is often studied using passive viewing with eye position held constant. Using closed-loop eye-tracking, with saccade-contingent stimulation and simultaneous intracranial recordings in surgical epilepsy patients, we tested the critical role of eye movement signals during natural visual processing in the hippocampus and hippocampal-amygdala circuit. Prior work shows that saccades elicit phase reset of ongoing neural excitability fluctuations across a broad array of cortical and subcortical areas. Here we show that saccade-related reset systematically modulates neuronal ensemble responses to visual input, enables phase-coding of information across the saccade-fixation cycle and modulates network connectivity between hippocampus and amygdala. The saccade-fixation cycle thus emerges as a fundamental sampling unit, organizing a range of neural operations including input representation, network connectivity and information coding.

One-Sentence Summary

Saccade-fixation cycle: a fundamental sampling unit, organizing input representation, information coding and network activation
Structures of the human adult muscle-type nicotinic receptor in resting and desensitised states
Authors: Li Anna , Pike Ashley C. W. , Webster Richard , Maxwell Susan , Liu Wei-Wei , Chi Gamma , Palace Jacqueline , Beeson David , Sauer David B. , Dong Yin Yao
Published: January 2025

Muscle-type nicotinic acetylcholine receptor (AChR) is the key signalling molecule in neuromuscular junctions. Here we present the structures of full-length human adult receptors in complex with Fab35 in α-bungarotoxin (αBuTx)-bound resting states, and in acetylcholine (ACh)-bound desensitised states. In addition to identifying the conformational changes during recovery from desensitisation, we also used electrophysiology to probe the effects of eight previously unstudied AChR genetic variants found in congenital myasthenic syndrome (CMS) patients, revealing they cause either slow- or fast-channel CMS characterised by prolonged or abbreviated ion channel bursts. The combined kinetic and structural data offer a better understanding of both AChR state transition and the pathogenic mechanisms of disease variants.
Identification of specific lipid-protein interactions in dividing cells using lipid-trap mass spectrometry
Authors: Paquola Andrea , Ozbalci Cagakan , Storck Elisabeth M. , Terry Stephen J. , Benson Clare E. , Eggert Ulrike S.
Published: January 2025

Cells actively maintain complex lipidomes that encompass thousands of lipids, however, many of the roles of these lipids remain unexplored. Specific interactions between lipids and membrane proteins are a likely reason for the evolutionary conservation of complex lipidomes. We report the development of a technique, named lipid-trap mass spectrometry (LTMS), to systematically study protein-lipid interactions directly captured from mammalian cells. LTMS uses immunoprecipitation of GFP-tagged proteins expressed in HeLa, followed by lipidomic analysis of lipids bound to the GFP-tagged protein. We applied LTMS to cell division to illustrate the technique. We chose this process because membranes regulate their lipid composition as they undergo major changes during cytokinesis and many cytokinetic proteins, including RACGAP1 and ESCRTIII components CHMP4B and CHMP2A, are membrane-associated. Using LTMS, we found that RACGAP1 and CHMP4B associate with specific lipid species in dividing compared to non-dividing cells. We expand our understanding of lipid diversity during cell division and present a general approach to explore lipid-protein interactions to further our understanding of the roles of lipids in mammalian cells.
Reduced enteric BDNF-TrkB signaling drives glucocorticoid-mediated GI dysmotility
Authors: Slosberg Jared , Puttapaka Srinivas N. , Seika Philippa , Hong Su Min , Singh Alpana , Kulkarni Subhash
Published: January 2025

Stress affects gastrointestinal (GI) function causing dysmotility, especially indisorders of gut-brain interactions (DGBI)patients. GI motility is regulated by the enteric nervous system (ENS), suggesting that stress alters ENS biology to cause dysmotility. While stress increases glucocorticoid levels through the hypothalamus-pituitary-adrenal axis, how glucocorticoids affect GI motility is not known. Glucocorticoid signaling reduces expression of specific transcriptional isoforms of brain-derived neurotrophic factor (BDNF) in the central nervous system, altering signaling through its receptor Tropomyosin-related kinase B (TrkB) to cause behavioral defects. However, since the nature of ENS-specificBdnfisoforms and their response to glucocorticoids remains unknown, we are limited in studying how stress impacts the ENS to cause dysmotility. Here, in male and female mice, we establish that stress-responsiveBdnfisoforms that are transcriptionally regulated at exons 4 and 6 represent >85% of allBdnfisoforms in the post-natal ENS, and thatBdnfandNtrk2(TrkB) are expressed by enteric neurons. We further show using male mice dosed with a synthetic glucocorticoid receptor (GR) agonist dexamethasone (Dexa), that increased glucocorticoid signaling in ENS significantly reduces the expression ofBdnftranscripts and protein and that it significantly reduces GI motility. Finally, by using HIOC, a specific synthetic agonist of TrkB, we observe that HIOC treatment significantly improved GI motility of a cohort of Dexa-treated male mice, when compared to Dexa-treated and HIOC-untreated mice. Our results implicate BDNF- TrkB signaling in the etiology of stress-associated dysmotility and suggest that TrkB is a putative therapeutic target for dysmotility in DGBI patients.

Significance statement

The mechanism through which stress-associated increase in glucocorticoid signaling causes gastrointestinal (GI) dysmotility is not well understood. GI motility is regulated by the enteric nervous system, which depends on optimal signaling between brain-derived neurotrophic factor (BDNF) and its receptor tropomyosin related kinase B (TrkB). The lack of clarity on whether glucocorticoid impact BDNF signaling in the ENS, analogous to the manner they affect BDNF in brain, has limited our understanding of stress-associated dysmotility. Here, by identifying the nature ofBdnfisoforms expressed in ENS, studying their response to increased glucocorticoid signaling, and testing the effect of TrkB agonist to promote healthy gut motility in a model of glucocorticoid-driven dysmotility, we implicate altered BDNF-TrkB signaling as the mechanism driving stress-associated dysmotility.
Bouncer’s receptor on sperm: Investigating sperm-egg compatibility in fish
Authors: Blaha Andreas , Pauli Andrea
Published: January 2025

Fertilization requires the successful binding and fusion of sperm and egg. In zebrafish, sperm-egg binding is mediated by the Spaca6-Izumo1-Tmem81 complex on sperm interacting with Bouncer on the egg. We previously found that expressing medaka Bouncer on zebrafish eggs andvice versaallows hybridization between medaka and zebrafish, two species that can normally not hybridize. Here, we tested whether providing zebrafish Spaca6 and Izumo1 on medaka sperm and vice versa enables cross-species compatibility from the side of the sperm. To this end, we generatedspaca6andizumo1knock-out (KO) lines in medaka, which are male sterile, and introduced zebrafishspaca6andizumo1transgenes. Transgenic medaka males did not fertilize zebrafish or medaka eggs with zebrafish Bouncer. Similarly, zebrafish males expressing medaka Spaca6 and Izumo1 failed to fertilize zebrafish eggs presenting medaka Bouncer. Unexpectedly, providing either full-length medaka Spaca6 or the Bouncer binding site of medaka Izumo1 in zebrafish sperm rescued the sterility ofspaca6andizumo1KO, respectively. Therefore, medaka Spaca6 and Izumo1 can interact with zebrafish Bouncer when paired with their zebrafish sperm complex members underscoring the nuanced interplay between molecular restrictions and compatibilities during sperm-egg interaction across teleosts.
The dual coding geneSLC35A4protects against oxidative stress
Authors: Ajala Ikram , Tiaiba Imadeddine , Grondin Benoît , Lipuma Damien , Jmii Souleimen , Benlyamani Hiba , Cappadocia Laurent , Vanderperre Benoît
Published: January 2025

Alternative proteins (AltProts) represent a newly recognized class of biologically active proteins encoded from alternative open reading frames (AltORFs) within already annotated genes. This study focuses on theSLC35A4gene, which encodes both the reference protein SLC35A4 and the alternative protein AltSLC35A4. Using a combination of microscopy and biochemical analyses, we confirmed the presence of AltSLC35A4 in the inner mitochondrial membrane, resolving previous conflicting reports. Previous studies employing ribosome profiling have revealed that during oxidative stress induced by sodium arsenite, the reference coding sequence ofSLC35A4exhibits the largest increase in translational efficiency among all cellular mRNAs. Our results confirmed this translational upregulation, with the emergence of SLC35A4 protein isoforms during oxidative stress in an upstream ORF-dependent manner. Notably, the expression of AltSLC35A4 remained unchanged during oxidative stress. Knock- out of SLC35A4 enhanced sensitivity to oxidative stress in a rescuable manner, indicating a direct implication for SLC35A4 in stress resistance. In conclusion, our research provides compelling evidence for the functional significance of the dual-coding nature ofSLC35A4for resistance to oxidative stress and highlights the importance of considering AltProts in the functional study of eukaryotic genes.
Short RNA chaperones promote aggregation-resistant TDP-43 conformers to mitigate neurodegeneration
Authors: Copley Katie E. , Mauna Jocelyn C. , Danielson Helen , Ngo Marilyn , Xie Longxin , Smirnov Ashleigh , Davis Matt , Mayne Leland , Linsenmeier Miriam , Rubien Jack D. , Portz Bede , Lee Bo Lim , Odeh Hana M. , Hallegger Martina , Ule Jernej , Pasinelli Piera , Poon Yan , Fawzi Nicolas L. , Black Ben E. , Donnelly Christopher J. , Jensen Brigid K. , Shorter James
Published: January 2025

Aberrant aggregation of the prion-like, RNA-binding protein TDP-43 underlies several debilitating neurodegenerative proteinopathies, including amyotrophic lateral sclerosis (ALS). Here, we define how short, specific RNAs antagonize TDP-43 aggregation. Short, specific RNAs engage and stabilize the TDP-43 RNA-recognition motifs, which allosterically destabilizes a conserved helical region in the prion-like domain, thereby promoting aggregation-resistant conformers. By mining sequence space, we uncover short RNAs with enhanced activity against TDP-43 and diverse disease-linked variants. The solubilizing activity of enhanced short RNA chaperones corrects aberrant TDP-43 phenotypes in optogenetic models and ALS patient-derived neurons. Remarkably, an enhanced short RNA chaperone mitigates TDP-43 proteinopathy and neurodegeneration in mice. Our studies reveal mechanisms of short RNA chaperones and pave the way for the development of short RNA therapeutics for fatal TDP-43 proteinopathies.
The 8-nm spaghetti: well-structured glycans coating linear tetrapeptide repeats discovered from freshwater with CryoSeek
Authors: Wang Tongtong , Sun Yitong , Li Zhangqiang , Yan Nieng
Published: January 2025

We recently developed a research strategy, termed CryoSeek, to identify uncharacterized bio-entities from natural or endogenous resources using cryo-electron microscopy (cryo-EM). Here we report the discovery of a glycofibril whose primary molecular mass is attributed to a thick glycan shell. The 3.3-Å resolution cryo-EM reconstruction reveals that the only protein component of the glycofibril, which is approximately 8 nm in diameter, is a linear chain of tetrapeptide repeats. Each tetrapeptide repeat consists of a 3,4-dihydroxyproline (diHyp), a Ser or Thr, and two less conserved residues. Two and one glycan chains are respectively O-linked to the diHyp and Ser/Thr residues. The protein sequence pattern of this glycofibril is similar to that of our recently observed TLP-4, although the glycan chains are different. We rename the previously characterized glycofibril as TLP-4a and designate this one as TLP-4b. Our discoveries reveal the critical role of glycans in structural folding of glycoconjugates and shed light on understanding the carbon/nitrogen ratio in biospheres.
Reproduction and preference to macronutrients have different relations to biological or chronological age inDrosophila
Authors: Lushchak Oleh , Strilbytska Olha , Petakh Pavlo , Kamyshnyi Oleksandr , Koliada Oleksandr , Semaniuk Uliana
Published: January 2025

Genetic manipulations, dietary composition and supplementations with varied drugs and natural compounds were shown to extend both the life- and health-span in different model organisms. An understanding of the mechanisms behind the beneficial properties of intervention includes the evaluation of physiological and molecular traits at certain time points that reflect the values at distinguished chronological age. Thus, if one cohort has a long-lived phenotype than measurements at certain time points represent the difference between organisms of different biological ages. We have compared reproduction and consumption of specific macronutrients for flies of the same cohort but related to different quartiles of chronological and biological age. We found that the decline in carbohydrate or protein consumption was stronger in the case of chronological rather than biological age. However, flies of biological or chronological quartile 4 consumed virtually the same amounts of macronutrients. The decline in reproduction was significantly reduced in relation to biological age. Thus, the decline was about 38-68% when within chronological quartiles 2 and 1 but only 4-31% for biological ones. The reproductive capacity was reduced by 86-93% in flies of chronological Q4 as compared to a 60-77% decrease for those of biological. Starting from quartile 2 biologically aged flies laid significantly higher number of eggs than flies of the same chronological quartile. Our results point out the significant difference in flies of the same biological and chronological quartile and raise the question about the suitability of comparison traits of organisms with different lifespans same chronological age.
Lifespan Normative Modeling of Brain Microstructure
Authors: Villalón-Reina Julio E. , Zhu Alyssa H. , Benavidez Sebastian , Moreau Clara A. , Feng Yixue , Chattopadhyay Tamoghna , Nabulsi Leila , Kushan Leila , John John P. , Joshi Himanshu , Thomopoulos Sophia I. , Lawrence Katherine E. , Nir Talia M. , Jahanshad Neda , Bearden Carrie E. , Kia Seyed Mostafa , Marquand Andre F. , Thompson Paul M. ,   the Alzheimer’s Disease Neuroimaging Initiative
Published: January 2025

Normative models of brain metrics based on large populations could be extremely valuable for detecting brain abnormalities in patients with a variety of disorders, including degenerative, psychiatric and neurodevelopmental conditions, but no such models exist for the brain’s white matter (WM) microstructure. Here we present the first large-scale normative model of brain WM microstructure–based on 19 international diffusion MRI datasets covering almost the entire lifespan (totaling N=54,583 individuals; age: 4-91 years). We extracted regional diffusion tensor imaging (DTI) metrics using a standardized analysis and quality control protocol and used hierarchical Bayesian regression (HBR) to model the statistical distribution of derived WM metrics as a function of age and sex. We extracted the average lifespan trajectories and corresponding centile curves for each WM region. We illustrate the utility of the method by applying it to detect and visualize profiles of WM microstructural deviations in a variety of contexts: in mild cognitive impairment, Alzheimer’s disease, and 22q11.2 deletion syndrome – a neurogenetic condition that markedly increases risk for schizophrenia. The resulting large-scale model provides a common reference to identify disease effects on the brain’s microstructure in individuals or groups, and to compare disorders, and discover factors affecting WM abnormalities. The derived normative models are a new resource publicly available to the community, adaptable and extendable to future datasets as the available data expands.
Lysine deacetylation inhibition reverses TDP-43 mislocalization and in combination with arimoclomol ameliorates neuromuscular pathology
Authors: Scozzari Serena , Columbro Stefano Fabrizio , Favagrossa Monica , Tortarolo Massimo , Cagnotto Alfredo , Salmona Mario , De Marco Giovanni , Bendotti Caterina , Calvo Andrea , Pasetto Laura , Bonetto Valentina
Published: January 2025

Background

Cytoplasmic inclusions containing TAR DNA-binding protein 43 kDa (TDP-43) are recognized as a major pathological feature of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia, but more recently have been associated with several neurodegenerative conditions. Cyclophilin A (PPIA), a foldase and molecular chaperone, interacts with TDP-43 and influences its aggregation and function. The interaction between PPIA and TDP-43 is facilitated by PPIA Lys-acetylation, which is reduced in peripheral blood mononuclear cells (PBMCs) of ALS patients showing signs of TDP-43 proteinopathy. In this study, we investigated the effect of lysine deacetylation inhibition to uncover the underlying mechanisms of TDP-43 proteinopathy in relation to PPIA acetylation, and to identify new therapeutic avenues.

Methods

Through a screening of histone deacetylase (HDAC) inhibitors in a cellular model of TDP-43 proteinopathy, we identified vorinostat/SAHA, capable of increasing PPIA acetylation, as the most effective in reversing TDP-43 mislocalization. We confirmed its effect in PBMCs from ALS patients. Next, we explored its impact on proteinopathy and PPIA acetylation in a severe and fast-progressing TDP-43-overexpressing mouse model (Thy1-hTDP-43), using several molecular biomarkers as outcome measures, including neurofilament light chain (NfL) in plasma.

Results

Thy1-hTDP-43 mice treated with SAHA showed a delayed onset of TDP-43 pathology, associated with PPIA nucleus-cytoplasm redistribution, lower levels of neurodegeneration and neuroinflammation markers, and improved neuromuscular function markers. However, over time, the broad-spectrum inhibitor SAHA was unable to counteract the two-fold overexpression of TDP-43 and led to the accumulation of side effects. When combined with the heat shock protein (HSP) co-inducer arimoclomol, a mitigation of the neurodegeneration was sustained. Moreover, a synergistic effect was observed in periphery, greatly enhancing tubulin acetylation and reducing pTDP-43 accumulation in the sciatic nerve. This resulted in a more pronounced reduction of NfL in plasma and acetylcholine receptor (AChR) γ-subunit expression in gastrocnemius muscle, indicating reduced axonal transport impairment and muscle denervation.

Conclusions

This study suggests that HDAC inhibition, by increasing acetylated PPIA, could be beneficial in restoring TDP-43 localization and function. The combination of lysine deacetylation inhibition and HSP induction shows a synergistic effectin vivoand has potential as a therapeutic approach for patients.
Mechanistic Insights into Dengue Virus Inhibition by a Clinical Trial Compound NITD-688
Authors: Wang Yan , Sun Long , Fernandes Luciana , Wang Yu-Hsiu , Zou Jing , Franklin Samuel J. , Hu Yanping , Palmer Lee K. , Yeung Jason , Barriga Daniela , Russell William K. , Moquin Stephanie A. , Shi Pei-Yong , Skepper Colin , Xie Xuping
Published: January 2025

Dengue, caused by the dengue virus (DENV), presents a significant public health challenge with limited effective treatments. NITD-688 is a potent pan-serotype DENV inhibitor currently in Phase II clinical trials. However, its mechanism of action is not fully understood. Here, we present the molecular details of how NITD-688 inhibits DENV. NITD-688 binds directly to the nonstructural protein 4B (NS4B) with nanomolar affinities across all four DENV serotypes and specifically disrupts the interaction between NS4B and nonstructural protein 3 (NS3) without significantly changing the interactions between NS4B and other viral or host proteins. NS4B mutations that confer resistance to NITD-688 reduce both NITD-688 binding to NS4B and disruption of the NS4B/NS3 interaction. Specifically, NITD-688 blocks the interaction of NS3 with a cytosolic loop within NS4B. This inhibits the formation of new NS4B/NS3 complexes and disrupts pre-existing complexes invitroand DENV-infected cells, ultimately inhibiting viral replication. Consistent with this mechanism, NITD-688 retains greater potency in cellular assays with delayed treatment compared to JNJ-1802, another NS4B inhibitor that has been studied in Phase II clinical trials. Together, these findings provide critical insights into the mechanism of action of NITD-688, facilitating the development of novel flavivirus NS4B inhibitors and informing future clinical interventions against DENV.
A novel pan class-I glucose transporter inhibitor DRB18 exhibits synergistic effectsin vitroandin vivowith paclitaxel against human non-small cell lung cancer
Authors: Shriwas Pratik , Bachmann Lindsey , Ward Ryan , Adhicary Subhodip , Nielsen Corinne M. , Li Yunsheng , Zhang Haiyun , Song Jingwen , Roberts Dennis , Bergmeier Stephen , Chen Xiaozhuo
Published: January 2025

Cancer cells depend on glucose for biomass synthesis, cell proliferation, and drug resistance. Glucose transporter (GLUT) transcripts as well as proteins are upregulated in human lungs and other cancers and are negatively correlated with patient survival. Thus, inhibiting GLUT function has been an attractive anticancer strategy. We previously characterized WZB117 and DRB18, first- and second-generation pan-class I GLUT inhibitors, respectively. DRB18 strongly inhibits glucose transport mediated by GLUT1-4 in non-small lung cancer (NSCLC) A549 cellsin vitroandin vivo. Here, we report DRB18 as a more stable and potent anticancer compound, compared to WZB117. DRB18 reduced glucose uptake in NSCLC A549, pancreatic Panc1, and cervical Hela cancer cells with varied but strong anticancer potencies in the presence or absence of extracellular nutrients such as ATP and glucose. Combined with different clinical and pre-clinical anticancer compounds such as V9302, CB839, Sutent, Brigatinib, DRB18 significantly increased death of A549 and Panc1 cells. Noteworthy, DRB18 exhibited strong anticancer synergy with paclitaxel, an approved chemo drug for NSCLC, drastically reducing cancer cell proliferationin vitroand growth of A549 tumors grafted on the flank of nude mice without increased side effects, compared to single drug treatments. Mechanistically, dual treatment of DRB18 with paclitaxel elevated the expression of Caspases 3 and 9, suggesting GLUT-inhibiting and apoptosis-inducing anticancer mechanisms of DRB18 with paclitaxel. Collectively, our results demonstrate anticancer efficacy of DRB18 either working alone or in combination with paclitaxel, providing a potentially more efficacious therapeutic strategy for treating advanced NSCLC and other cancers.
Diverse and Location-Specific Roles of PlexinA2, PlexinA4, and NCAM in Developing Hippocampal Mossy Fibers
Authors: Zhao Xiao-Feng , Kohen Rafi , Van Battum Eljo Y. , Zeng Ying , Zhang Xiaolu , Johnson Craig N. , Wang Karen , Lim Brian C. , Oses-Prieto Juan A. , Rasband Joshua M. , Burlingame Alma L. , Pasterkamp R. Jeroen , Rasband Matthew N. , Giger Roman J.
Published: January 2025

Mossy fibers (MFs) originate from dentate granule cells and innervate area CA3 of the hippocampus. Upon entry of CA3, MFs partition into two prominent axon bundles, the suprapyramidal tract (SPT) and infrapyramidal tract (IPT) and form lamina specific synaptic contacts in the stratum lucidum (SL) and stratum oriens (SO), respectively. Here we employed new mouse lines to dissect the function of Sema6A and its receptors, PlexinA2 (PlxnA2) and PlxnA4, in developing MFs. InSema6a−/−mice, MF partitioning into SPT and IPT bundles is incomplete and IPT axons in the SO are overextended, while the SPT correctly innervates the SL. Loss of neuronalSema6aresults in defective MF patterning and we show that this involves Sema6A reverse signaling.Plxna4controls MF partitioning, SPT axon bundling and laminar targeting to the SL, as well as IPT length. Many of these defects are replicated in mice deficient for PlxnA4 GAP catalytic activity, underscoring the importance of this GAP domain. MFs are tightly fasciculated inPlxna2−/−mice and fail to separate into SPT and IPT bundles, and defects are significantly reduced in PlxnA2 GAP mutants, highlighting the involvement of GAP-independent signaling events. To further explore the molecular basis of aberrant axon fasciculation, we employed anti-PlxnA2 dependent proximity biotinylation and identified several PlxnA2-associated Ig-CAM family members. We observed a genetic interaction betweenPlxna2andNcam1,but notPlxna4andNcam1, for SPT and IPT formation and positioning in CA3. Together, our studies provide insights into the multifaceted and overlapping, yet distinct, functions of PlxnA family members in orchestrating specific guidance decisions in developing MFs.