bluuebunny/crossref_metadata_2025 huggingface dataset:
Transcriptional modulation unique to vulnerable motor neurons predicts ALS across species and SOD1 mutations
Authors: Mei Irene , Nichterwitz Susanne , Leboeuf Melanie , Nijssen Jik , Lenoel Isadora , Repsilber Dirk , Lobsiger Christian S. , Hedlund Eva
Published: March 2024

Amyotrophic lateral sclerosis (ALS) is characterized by the progressive loss of somatic motor neurons (MNs), which innervate skeletal muscles. However, certain MN groups including ocular MNs that regulate eye movement are relatively resilient to ALS. To reveal mechanisms of differential susceptibility, we investigate the transcriptional dynamics of two vulnerable and two resilient MN populations in SOD1G93A ALS mice. Analysis of differentially expressed genes (DEGs) shows that each neuron type displays a largely unique spatial and temporal response to disease. Baseline gene expression of resilient MNs is clearly divergent from vulnerable MNs and their response to mutant SOD1 is minor, with regulation of few genes, some with known neuroprotective properties, including e.g.Ucn,CckandPostn. EASE, fGSEA and ANUBIX enrichment analysis demonstrate that vulnerable MN groups share pathway activation, including regulation of neuronal death, inflammatory response, ERK and MAPK cascades, cell adhesion and synaptic signaling. These pathways are largely driven by 11 upregulated genes, includingAtf3, Nupr1, Cd44, Gadd45a, Ngfr, Ccl2, Ccl7, Gal, Timp1, Serpinb1aandChl1, and indicate that cell death occurs through similar mechanisms across vulnerable MNs albeit with distinct timing. Machine learning using DEGs upregulated in our SOD1G93A spinal MNs predict disease in human stem cell-derived MNs harboring theSOD1E100Gmutation, and show that dysregulation ofVGF,INA, PENKandNTSare strong disease-predictors across SOD1 mutations and species. Meta-analysis across SOD1 transcriptome datasets identifies a shared vulnerability code of 32 genes including e.gAtf3,Nupr1,Vgf,Ina,Sprr1a, Fgf21, C1qb, Gap43, Adcyap1,andMt1. In conclusion our study reveals cell-type-specific gene expression and dynamic disease-induced regulation that may act to preserve neurons and can be used to predict disease.
Functional regimes define the response of the soil microbiome to environmental change
Authors: Lee Kiseok Keith , Liu Siqi , Crocker Kyle , Huggins David R. , Tikhonov Mikhail , Mani Madhav , Kuehn Seppe
Published: March 2024

The metabolic activity of soil microbiomes plays a central role in carbon and nitrogen cycling. Given the changing climate, it is important to understand how the metabolism of natural communities responds to environmental change. However, the ecological, spatial, and chemical complexity of soils makes understanding the mechanisms governing the response of these communities to perturbations challenging. Here, we overcome this complexity by using dynamic measurements of metabolism in microcosms and modeling to reveal regimes where a few key mechanisms govern the response of soils to environmental change. We sample soils along a natural pH gradient, construct >1500 microcosms to perturb the pH, and quantify the dynamics of respiratory nitrate utilization, a key process in the nitrogen cycle. Despite the complexity of the soil microbiome, a minimal mathematical model with two variables, the quantity of active biomass in the community and the availability of a growth-limiting nutrient, quantifies observed nitrate utilization dynamics across soils and pH perturbations. Across environmental perturbations, changes in these two variables give rise to three functional regimes each with qualitatively distinct dynamics of nitrate utilization over time: a regime where acidic perturbations induce cell death that limits metabolic activity, a nutrientlimiting regime where nitrate uptake is performed by dominant taxa that utilize nutrients released from the soil matrix, and a resurgent growth regime in basic conditions, where excess nutrients enable growth of initially rare taxa. The underlying mechanism of each regime is predicted by our interpretable model and tested via amendment experiments, nutrient measurements, and sequencing. Further, our data suggest that the long-term history of environmental variation in the wild influences the transitions between functional regimes. Therefore, quantitative measurements and a mathematical model reveal the existence of qualitative regimes that capture the mechanisms and dynamics of a community responding to environmental change.
Correspondence of fentanyl brain pharmacokinetics and behavior measured via engineering opioids biosensors and computational ethology
Authors: Muthusamy Anand K. , Rosenberg Matthew H. , Kim Charlene H. , Wang Alex Z. , Ebisu Haruka , Chin Theodore M. , Koranne Ashil , Marvin Jonathan S. , Cohen Bruce N. , Looger Loren L. , Oka Yuki , Meister Markus , Lester Henry A.
Published: March 2024

Despite the ongoing epidemic of opioid use disorder and death by fentanyl overdose, opioids remain the gold standard for analgesics. Pharmacokinetics (PK) dictates the individual’s experience and utility of drugs; however, PK and behavioral outcomes have been conventionally studied in separate groups, even in preclinical models. To bridge this gap, we developed the first class of sensitive, selective, and genetically encodable fluorescent opioid biosensors, iOpioidSnFRs, including the fentanyl sensor, iFentanylSnFR. We expressed iFentanylSnFR in the ventral tegmental area of mice and recorded [fentanyl] alongside videos of behaviors before and after administration. We developed a machine vision routine to quantify the effects of the behavior on locomotor activity. We found that mice receiving fentanyl exhibited a repetitive locomotor pattern that paralleled the [fentanyl] time course. In a separate experiment, mice navigating a complex maze for water showed a dose-dependent impairment in navigation, in which animals repeated incorrect paths to the exclusion of most of the unexplored maze for the duration of the average fentanyl time course. This approach complements classical operant conditioning experiments and introduces a key feature of human addiction, the ability to carry out an ethologically relevant survival task, only now quantified in rodents. Finally, we demonstrate the utility of iFentanylSnFR in detecting fentanyl spiked into human biofluids and the generalizability of engineering methods to evolve selective biosensors of other opioids, such as tapentadol and levorphanol. These results encourage diagnostic and continuous monitoring approaches to personalizing opioid regimens for humans.
Cohesin regulates promoter-proximal pausing of RNA Polymerase II by limiting recruitment of super elongation complex
Authors: Tei Shoin , Sakata Toyonori , Yoshimura Atsunori , Natsume Toyoaki , Kanemaki Masato T , Bando Masashige , Shirahige Katsuhiko
Published: March 2024

Cohesin is a ring-shaped complex, responsible for establishing sister chromatid cohesion and forming topologically associating domains (TADs) and chromatin loops. Loss-of-function mutations in cohesin subunits and its regulatory factors can cause Cornelia de Lange syndrome (CdLS). Because dysregulated gene expression was observed in CdLS, it has long been thought that cohesin plays a regulatory role in transcription. Here, we investigated the effect of acute cohesin depletion on transcription and observed that a small number of genes exhibited differential expression. Analysis of RNA polymerase II (Pol II) distribution revealed that the depletion reduced Pol II promoter binding and pausing simultaneously at the majority of genes. This implies that at most genes, the two decreases counterbalance each other, resulting in unchanged gene expression. Additionally, we find that cohesin loss increased promoter binding of super elongation complex (SEC), which mediates the release of Pol II from paused state. Moreover, the reduction in pausing caused by cohesin depletion was no longer observed when SEC was inhibited. These observations suggest that cohesin regulates Pol II pausing by restricting SEC recruitment to promoters. Together, our study demonstrates the involvement of cohesin in transcriptional regulation, particularly in Pol II pause and release.
C1q/MASP complexes – hybrid complexes of classical and lectin pathway proteins are found in the circulation
Authors: Rosbjerg Anne , Plchová Tereza Alica , Bayarri-Olmos Rafael , Holm Bettina Eide , Pedersen Ida Sandau , Skjoedt Mikkel-Ole , Garred Peter
Published: March 2024

Complement pathways, traditionally regarded as separate entities in vitro, are increasingly noted for cross-communication and bypass mechanisms. Among these, the MBL/Ficolin/CL associated serine protease-3 (MASP-3) — a component of lectin pathway pattern recognition receptors (PRMs) — has shown the ability to process critical substrates like pro-factor D and insulin growth factor binding protein-5 (IGFBP-5). Given shared features between lectin pathway PRMs and C1q from the classical pathway, we hypothesized that C1q might be a viable in vivo binding partner for the MASPs.

We used microscale thermophoresis, ELISA and immunoprecipitation assays to detect C1q/MASP complexes and functionally assessed the complexes through enzymatic cleavage assays.

C1q/MASP-3 complexes were detected in human serum and correlated well with MASP-3 serum levels in healthy individuals. The binding affinity between MASP-3 and C1q in vitro was in the nanomolar range, and the interaction was calcium-dependent, as demonstrated by their dissociation in the presence of EDTA. Furthermore, most of the circulating C1q-bound MASP-3 was activated. Based on immunoprecipitatin, also C1q/MASP-2 complexes appereared to be present in serum. Finally, C1q/MASP-2 and C1q/MASP-3 in vitro complexes were able to cleave C4 and pro-factor D, respectively.

Our study reveals the existence of C1q/MASP complexes in the circulation of healthy individuals and both C1q/MASP-2 and C1q/MASP-3 complexes display proteolytic activity. Hence, this study uncovers a crosstalk route between complement pathways not previously described.
RNA 3’end tailing safeguards cells against products of pervasive transcription termination
Authors: Wu Guifen , Rouvière Jérôme O. , Schmid Manfred , Jensen Torben Heick
Published: March 2024

Premature transcription termination yields a wealth of unadenylated (pA-) RNA. Although this can be targeted for degradation by the Nuclear EXosome Targeting (NEXT) complex, possible back-up pathways remain poorly understood. Here, we find RNA 3’end uridylation and adenylation upon NEXT inactivation. U-tailed RNAs are generally short and modified by the cytoplasmic tailing enzymes, TUT4/7, following their PHAX-dependent nuclear export and prior to their degradation by the cytoplasmic exosome or the exoribonuclease DIS3L2. Longer RNAs are instead adenylated redundantly by enzymes TENT2, PAPOLA and PAPOLG. These transcripts are either degraded via the nuclear Poly(A) tail eXosome Targeting (PAXT) connection or exported by conventional factors and removed by the cytoplasmic exosome in a translation-dependent manner. Failure to do so decreases global translation and induces cell death. We conclude that post-transcriptional 3’end modification and removal of excess pA-RNA is achieved by tailing enzymes and export factors shared with productive RNA pathways.
Stereochemically-aware bioactivity descriptors for uncharacterized chemical compounds
Authors: Comajuncosa-Creus Arnau , Lenes Aksel , Sánchez-Palomino Miguel , Aloy Patrick
Published: March 2024

We recently presented a set of deep neural networks to generate bioactivity descriptors associated to small molecules (i.e.Signaturizers), capturing their effects at increasing levels of biological complexity (i.e. from protein targets to clinical outcomes)1. However, such models were trained on 2D representations of molecules and are thus unable to capture key differences in the activity of stereoisomers. Now, we systematically assess the relationship between stereoisomerism and bioactivity on over 1M compounds, finding that a very significant fraction (∼40%) of spatial isomer pairs show, to some extent, distinct bioactivities. We then used these data to train a second generation of Signaturizers, which are now stereochemically-aware, and provide an even more faithful description of complex small molecule bioactivity properties.
Elongation capacity of polyunsaturated fatty acids in the annelidPlatynereis dumerilii
Authors: Ramos-Llorens Marc , Bainour Khalida , Adelmann Leonie , Hontoria Francisco , Navarro Juan C. , Raible Florian , Monroig Óscar
Published: March 2024

In animals, elongation of very long-chain fatty acid (Elovl) proteins play pivotal functions in the biosynthesis of fatty acids, including the physiologically essential long-chain polyunsaturated fatty acids (LC-PUFA). Polychaetes have important roles in marine ecosystems, contributing not only to nutrient recycling but also exhibiting a distinctive capacity for biosynthesising LC-PUFA, as emphasised in previous research. In order to expand our current understanding of the LC-PUFA biosynthesis in polychaetes, the present study conducted a thorough molecular and functional characterisation of Elovl occurring in the model organismPlatynereis dumerilii. In this study, we identify six Elovl proteins in the genome ofP. dumerilii. The sequence and phylogenetic analyses established that four Elovl, identified as Elovl2/5, Elovl4 (two genes) and Elovl1/7, have putative functions in the LC-PUFA biosynthesis. Functional characterisation in yeast confirmed the roles of these elongases in the LC-PUFA biosynthetic pathways, demonstrating thatP. dumeriliipossesses a varied and functionally diverse complement of Elovl enzymes that, along the enzymatic specificities of previously characterised desaturases, enableP. dumeriliito perform all the reactions required for the biosynthesis of the LC-PUFA. Importantly, we uncovered that one of the two Elovl4-encoding genes fromP. dumeriliiis remarkably long in comparison with any other animals’ Elovl, which contains a C terminal KH domain unique among Elovl enzymes. The distinctive expression pattern of this protein in photoreceptors strongly suggests a central role in vision.
Dorsomedial Striatal Glutamatergic Transmission Inhibits Binge Drinking in Selectively Bred Crossed High Alcohol Preferring Mice
Authors: Bauer Meredith R. , McVey Megan M. , Zhang Yanping , Boehm Stephen L.
Published: March 2024

Crossed high alcohol preferring (cHAP) mice have been selectively bred to consume considerable amounts of alcohol resulting in binge drinking. The dorsal striatum (DS) is a brain region involved in action selection where the dorsomedial striatum (DMS) is involved in goal-directed action selection and dorsolateral striatum (DLS) is involved in habitual action selection. Alcohol use disorder (AUD) may involve a disruption in the balance between the DMS and DLS. While the DLS is involved in binge drinking, the reliance on the DMS and DLS in binge drinking has not been investigated in cHAP mice. We have previously demonstrated that glutamatergic activity in the DLS is necessary for binge-like alcohol drinking in C57BL/6J mice, another high drinking mouse. Because of this, we hypothesized that DLS glutamatergic activity would gate binge-like alcohol drinking in cHAP mice. cHAP mice underwent bilateral cannulation into the DMS or DLS and were allowed free-access to 20% alcohol for two-hours each day for 11 days. Mice were microinjected with the AMPA receptor (AMPAR) antagonist, NBQX, into the DMS or DLS immediately prior to alcohol access. AMPAR protein expression was also assessed in a separate group of animals in DS subregions following an 11-day drinking history. We found that intra-DMS (but not intra-DLS) NBQX, alters binge alcohol drinking, with intra-DMS NBQX increasing alcohol consumption. We also found that the ratio of GluA1 to GluA2 differs across DS subregions. Together, these findings suggest that glutamatergic activity in the DMS may serve to limit binge drinking in cHAP mice.
A Comparison of Antibody-Antigen Complex Sequence-to-Structure Prediction Methods and their Systematic Biases
Authors: McCoy Katherine Maia , Ackerman Margaret E. , Grigoryan Gevorg
Published: March 2024

The ability to accurately predict antibody-antigen complex structures from their sequences could greatly advance our understanding of the immune system and would aid in the development of novel antibody therapeutics. There have been considerable recent advancements in predicting protein-protein interactions (PPIs) fueled by progress in machine learning (ML). To understand the current state of the field, we compare six representative methods for predicting antibody-antigen complexes from sequence, including two deep learning approaches trained to predict PPIs in general (AlphaFold-Multimer, RoseTTAFold), two composite methods that initially predict antibody and antigen structures separately and dock them (using antibody-mode ClusPro), local refinement in Rosetta (SnugDock) of globally docked poses from ClusPro, and a pipeline combining homology modeling with rigid-body docking informed by ML-based epitope and paratope prediction (AbAdapt). We find that AlphaFold-Multimer outperformed other methods, although the absolute performance leaves considerable room for improvement. AlphaFold-Multimer models of lower-quality display significant structural biases at the level of tertiary motifs (TERMs) towards having fewer structural matches in non-antibody containing structures from the Protein Data Bank (PDB). Specifically, better models exhibit more common PDB-like TERMs at the antibody-antigen interface than worse ones. Importantly, the clear relationship between performance and the commonness of interfacial TERMs suggests that scarcity of interfacial geometry data in the structural database may currently limit application of machine learning to the prediction of antibody-antigen interactions.
A bacterial type III effector hijacks plant ubiquitin proteases to evade degradation
Authors: Yu Wenjia , Li Meng , Wang Wenjun , Zhuang Haiyan , Luo Jiamin , Sang Yuying , Segonzac Cecile , Macho Alberto P.
Published: March 2024

Gram-negative bacterial pathogens inject effector proteins inside plant cells using a type III secretion system. These effectors manipulate plant cellular functions and suppress the plant immune system in order to promote bacterial proliferation. Despite the fact that bacterial effectors are exogenous threatening proteins potentially exposed to the protein degradation systems inside plant cells, effectors are relative stable and able to perform their virulence functions. In this work, we found that RipE1, an effector protein secreted by the bacterial wilt pathogen,Ralstonia solanacearum, undergoes phosphorylation of specific residues inside plant cells, and this promotes its stability. Moreover, RipE1 associates with plant ubiquitin proteases, which contribute to RipE1 deubiquitination and stabilization. The absence of those specific phosphorylation sites or specific host ubiquitin proteases leads to a substantial decrease in RipE1 protein accumulation, indicating that RipE1 hijacks plant post-translational modification regulators in order to promote its own stability. These results suggest that effector stability or degradation in plant cells constitute another molecular event subject to co-evolution between plants and pathogens.
Biophysics-based protein language models for protein engineering
Authors: Gelman Sam , Johnson Bryce , Freschlin Chase , Sharma Arnav , D’Costa Sameer , Peters John , Gitter Anthony , Romero Philip A.
Published: March 2024

Protein language models trained on evolutionary data have emerged as powerful tools for predictive problems involving protein sequence, structure, and function. However, these models overlook decades of research into biophysical factors governing protein function. We propose Mutational Effect Transfer Learning (METL), a protein language model framework that unites advanced machine learning and biophysical modeling. Using the METL framework, we pretrain transformer-based neural networks on biophysical simulation data to capture fundamental relationships between protein sequence, structure, and energetics. We finetune METL on experimental sequence-function data to harness these biophysical signals and apply them when predicting protein properties like thermostability, catalytic activity, and fluorescence. METL excels in challenging protein engineering tasks like generalizing from small training sets and position extrapolation, although existing methods that train on evolutionary signals remain powerful for many types of experimental assays. We demonstrate METL’s ability to design functional green fluorescent protein variants when trained on only 64 examples, showcasing the potential of biophysics-based protein language models for protein engineering.
Inhibition of Proteasome Activity Facilitates Definitive Endodermal Specification of pluripotent Stem Cells by influencing YAP signaling
Authors: Ashok Akshaya , Ashwathnarayan Ashwini , Bhaskar Smitha , Shekar Spandana , Prasanna Jyothi , Kumar Anujith
Published: March 2024

Understanding the molecular players that control the specification of definitive endoderm is imperative to obtain the homogenous population of pancreatic β-cells from stem cells. Though the Ubiquitin proteasome system (UPS) has been envisaged as a crucial intracellular protein degradation system, its role in germ layer specification remains elusive. In this study, using a mouse embryonic stem cells model system (mESCs) we observed decreased proteasomal activity specifically during endoderm, but not in meso- or ecto-derm differentiation. Extraneous inhibition of proteasomal activity during differentiation enhanced the expression of endodermal genes specifically. Enhancing proteasomal activity by including the activator IU1 in the induction culture, inhibited definitive endodermal differentiation. Further, inhibiting proteasomal activity at the definitive endodermal stage resulted in enhanced generation of insulin-positive cells. A similar increase in endodermal gene expression by inhibiting proteasomal activity was observed in miPSC and hiPSC differentiated towards endodermal lineage. Mechanistic insight showed no contribution of endoplasmic reticulum unfolded protein response but revealed the involvement of the YAP signaling pathway in proteasome-inhibited enhanced endodermal differentiation. Unravelling the specific involvement of UPS in endodermal cell fate specification in pluripotent stem cells paves the way for obtaining better qualitative and quantitative definitive endodermal cells for plausible cellular therapy in the future.
Microbial utilisation of maize rhizodeposits applied to an agricultural soil at a range of concentrations
Authors: Niedeggen Daniela , Rüger Lioba , Oburger Eva , Santangeli Michael , Ahmed Mutez , Vetterlein Doris , Blagodatsky Sergey , Bonkowski Michael
Published: March 2024

Rhizodeposition fuels carbon (C) and nutrient cycling in soil. However, the dynamics of microbial growth on rhizodeposits in relation to the distance from the root have not been well studied. This study investigates microbial growth on individual organic components of rhizodeposits and on maize root-derived exudates and mucilage from an agricultural soil. By creating a gradient of substrate concentrations, we simulated reduced microbial access to rhizosphere C with increasing distance to the root surface. We identified distinct C-thresholds for the activation of microbial growth, and these were significantly higher for rhizodeposits compared to singular, simple sugars. In addition, testing for stoichiometric constraints of microbial growth by supplementing N and P showed accelerated and increased microbial growth by activating a larger proportion of the microbial biomass. Early and late season exudates triggered significantly different microbial growth responses. The mineralisation of early season exudates was induced at a high C-threshold, whereas the mineralisation of late season exudates showed ’sugar-like’ properties, with a low C-threshold, high substrate affinity, and a reduced maximum respiration rate. Mucilage exhibited the highest C-threshold for the activation of microbial growth, although with a short lag-period and with an efficient mucilage degradation comparable to that of sugars. By determining kinetic parameters and turnover times for different root-derived substrates, our data enable the upscaling of micro-scale processes to the whole root system, allowing more precise predictions of how rhizodeposits drive microbial C and nutrient dynamics in soil.

Highlights

Growth thresholds for rhizodeposits were significantly higher than for singular, simple sugars.

No distinct microbial growth on root exudates was observed even at high concentrations.

Mucilage has a short lag-phase and efficient decomposition like sugars but only above a high threshold.

N and P limited microbial growth in the agricultural soil.
AGE READINGS AND ASSESSMENT IN COASTAL BATOID ELASMOBRANCHS FROM SMALL-SCALE SIZE-SELECTIVE FISHERY: THE IMPORTANCE OF DATA COMPARABILITY IN MULTISPECIFIC ASSEMBLAGES
Authors: Scacco Umberto , Zanardi Fabiana , Kroha Silvio , Mancini Emanuele , Tiralongo Francesco , Nascetti Giuseppe
Published: March 2024

Age readings and assessment of vertebral growth increments were obtained in four batoid elasmobranch species, namelyDasyatis pastinaca,Raja asterias,Torpedo marmorata, andTorpedo torpedo. Samples were obtained opportunistically from the bycatch of a size-selective fishery, such as local small-scale trammel net fishing, in the coastal waters of the Central Tyrrhenian Sea during 2019-21. We analysed the vertebrae by a simple and rapid method preventing staining phase and histological preparation to elucidate band pairs in all species studied. Consistency of age estimates was checked by controlling for agreement on band pair counts between and within observers, and by estimating the relationships between vertebral diameter and height, and body size of the animal. Based on these data, we developed a statistical routine to obtain multiple estimates of age and growth parameters for incomplete samples due to size-selective fishing. The acceptable agreement between and within readers and the increase in vertebral size with body size demonstrated the consistency of the method. Based on the results of Von Bertalanffy and Gompertz growth models, body size was a better predictor of age than vertebral size, and Gompertz models were a better estimator of age and growth parameters than Von Bertalanffy models. The estimated parameters (k and t0, kg and cg) matched the data available for the species studied in the Mediterranean Sea. In fact,D. pastinaca,T. torpedoandR. asteriasshowed the lowest (k = 0.05-0.12), intermediate (k = 0.112-0.19) and highest (k = 0.18-0.23) growth rate, respectively. Overall, the method proved effective both in delineating band pairs in vertebrae of different species by making use of only minimal optical equipment and a single reagent, and in reliably estimating the age and growth parameters of problematic samples due to size-selective fishing. The replicability of the method will help to collect comparable demographic data in similar samples from other areas of the Mediterranean, as well as in assemblages of different species from other places.
Quantitative effects of co-culture on T cell motility and cancer-T cell interactions
Authors: Li Xinyue , Jin Taoli , Wang Lisha , Li Ming , Han Weijing , Li Xuefei
Published: March 2024

One of the primary challenges in current cancer immunotherapy is the insufficient infiltration of cytotoxic T cells into solid tumors. Despite ongoing investigations, the mechanisms restricting T cell infiltration in immune-cold tumors remains elusive, hindered by the intricate tumor microenvironment. Here, we co-cultured mouse cancer cell lines with cancer-specific cytotoxic T cells to study the influence of cancer-T cell interactions on T cell motility, a crucial factor for effective tumor infiltration. By quantifying T cell motility patterns, we found that cancer-specific T cells exhibited extended contact time with cancer-cell clusters and higher directional persistence than non-specific T cells. Computational modelling suggested that T cells with stronger persistence could facilitate efficient searching for cancer clusters. Transcriptomic profiling revealed T cells recognizing cancer cells orchestrate accumulation on cancer cell clusters by activating adhesion proteins on both cancer cells and T cells, thereby fostering prolonged interaction on cancer cells. Furthermore, we observed that there were two distinct subpopulations of cancer cells after co-culturing with cancer-specific T cells: one expressing elevated levels of T-cell attractants and antigen-presentation molecules, while the other expressing immunosuppressive molecules and undergoing epithelial-to-mesenchymal transition. These dynamic insights into the complex interplay of cancer-T cell interactions and their impact on T cell motility hold implications for refining more efficacious cancer immunotherapy strategies.
Cycle-frequency content EEG analysis improves the assessment of respiratory-related cortical activity
Authors: Navarro-Sune Xavier , Raux Mathieu , Hudson Anna L. , Similowski Thomas , Chavez Mario
Published: March 2024

Time-Frequency (T-F) analysis of EEG is a common technique to characterise spectral changes in neural activity. This study explores the limitations of utilizing conventional spectral techniques in examining cyclic event-related cortical activities due to challenges, including high inter-trial variability. Introducing the Cycle-Frequency (C-F) analysis, we aim to enhance the evaluation of cycle-locked respiratory events. For synthetic EEG that mimicked cycle-locked pre-motor activity, C-F had more accurate frequency and time localization compared to conventional T-F analysis, even for a significantly reduced number of trials and a variability of breathing rhythm. Preliminary validations using real EEG data during both unloaded breathing and loaded breathing (that evokes pre-motor activity) suggest potential benefits of using the C-F method, particularly in normalizing time units to cyclic activity phases and refining baseline placement and duration. The proposed approach could provide new insights for the study of rhythmic neural activities, complementing T-F analysis.
Mitochondrial dysfunction drives a neuronal exhaustion phenotype in methylmalonic aciduria
Authors: Denley Matthew C.S. , Straub Monique S. , Marcionelli Giulio , Güra Miriam A. , Penton Ribas David , Delvendahl Igor , Poms Martin , Vekeriotaite Beata , Conte Federica , von Meyenn Ferdinand , Froese D. Sean , Baumgartner Matthias R.
Published: March 2024

Methylmalonic aciduria (MMA) is an inborn error of metabolism resulting in loss of function of the enzyme methylmalonyl-CoA mutase (MMUT). Despite acute and persistent neurological symptoms, the pathogenesis of MMA in the central nervous system is poorly understood, which has contributed to a dearth of effective brain specific treatments. Here we utilised patient-derived induced pluripotent stem cells andin vitrodifferentiation to generate a novel human neuronal model of MMA. We reveal strong evidence of mitochondrial dysfunction caused by deficiency of MMUT in patient neurons. By employing patch-clamp electrophysiology, targeted metabolomics, and bulk transcriptomics, we further expose an altered state of excitability, which we suggest may be connected to metabolic rewiring which is exacerbated by application of 2-dimethyloxoglutrate. Our work provides first evidence of mitochondrial driven neuronal dysfunction in MMA, which through our comprehensive characterisation of this paradigmatic model, enables first steps to identifying effective therapies.
Wee1 inhibition decouples Cdk1 and Plk1 activities: a role for gradual Cdk1 activation throughout G2 phase
Authors: Akopyan Karen , Lindqvist Arne
Published: March 2024

At completion of DNA replication, the mitotic kinases CDK1 and PLK1 are activated. Their activities increase slowly through early G2 phase, but the reason for this low-level activity before mitotic entry is not clear. Using a combination of experiments and mathematical modelling, we find that gradual CDK1 activation through G2 phase stimulates production of mitotic factors and coordinates activation of CDK1 and PLK1. We find that inhibition of CDK1 during G2 phase limits transcription of mitotic factors.

Conversely, the duration of premature mitosis by forced activation of CDK1 is inversely related to the time-point in G2 when mitosis is triggered. Forced CDK1 activation not only leads to a lack of mitotic factors, but also decouples CDK1 and PLK1 activation. Accordingly, we find that duration of forced mitosis by WEE1 inhibition can be partially rescued by expression of constitutively active PLK1. Our results show a function for slow mitotic kinase activation through G2 phase and suggests a mechanism for how the timing of mitotic entry is linked to preparation for mitosis.

Highlights

Slow Cdk1 activation through G2 phase coordinates expression of mitotic proteins and activation of mitotic kinases

The duration of forced mitosis by WEE1 inhibition depends on when in G2 mitosis is triggered

Forced CDK1 activation decouples CDK1 and PLK1 activation

The duration of forced mitosis can be partially rescued by expression of constitutively active PLK1
Aedes albopictuscolonies from different geographic origins differ in their sleep and activity levels but not in the time of peak activity
Authors: Wynne Nicole E. , Applebach Emilie , Chandrasegaran Karthikeyan , Ajayi Oluwaseun M. , Chakraborty Souvik , Bonizzoni Mariangela , Lahondère Chloé , Benoit Joshua B. , Vinauger Clément
Published: March 2024

Mosquitoes occupy a wide range of habitats where they experience various environmental conditions. The ability of some species, such as the tiger mosquito,Aedes albopictus, to adapt to local conditions certainly contributes to their invasive success. Among traits that remain to be examined, mosquitoes’ ability to time their activity with that of the local host population has been suggested to be of significant epidemiological importance. However, whether different populations display heritable differences in their chronotype has not been examined. Here, we compared laboratory strains originating from 8 populations from 3 continents, monitored their spontaneous locomotor activity patterns, and analyzed their sleep-like states. Overall, all strains showed conserved diurnal activity concentrated in the hours preceding the crepuscule. Similarly, they all showed increased sleep levels during the morning and night hours. However, we observed strain-specific differences in the activity levels at each phase of the day. We also observed differences in the fraction of time that each strain spends in a sleep-like state, explained by variations in the sleep architecture across strains. Human population density and the latitude of the site of geographic origin of the tested strain showed significant effects on sleep and activity patterns. Altogether, these results suggest thatAe. albopictusmosquitoes adapt to local environmental conditions via heritable adaptations of their chronotype.
A first look at the genome structure of hexaploid 'Black Mitcham' peppermint (Mentha piperita L.)
Authors: Talbot Samuel C. , Vining Kelly J. , Lange B. Markus , Pandelova Iovanna
Published: March 2024

Peppermint,Mentha xpiperitaL., is a hexaploid (2n = 6x = 72) and the predominant cultivar of commercial mint oil production in the US. This cultivar is threatened because of high susceptibility to the fungal disease Verticillium wilt, caused byVerticillium dahliae. This report details the first draft polyploid chromosome-level genome assembly for this mint species. The Black Mitcham genome resource will broaden comparative studies of disease resistance, essential oil biosynthesis, and hybridization events within the genusMentha. It will also be a valuable contribution to the body of phylogenetic studies involvingMenthaand other genera that contain species with varying ploidy levels.
Loss of the Na+/K+cation pump CATP-1 suppressesnekl-associated molting defects
Authors: Binti Shaonil , Edeen Phil T. , Fay David S.
Published: March 2024

The conservedC. elegansprotein kinases NEKL-2 and NEKL-3 regulate multiple steps of membrane trafficking and are required for larval molting. Through a forward genetic screen we identified a loss-of-function mutation incatp-1as a suppressor of molting defects in synthetically lethalnekl-2; nekl-3double mutants.catp-1is predicted to encode a membrane- associated P4-type ATPase involved in Na+–K+exchange. Moreover, a mutation predicted to abolish CATP-1 ion-pump activity also suppressednekl-2; nekl-3mutants. Endogenously tagged CATP-1 was primarily expressed in epidermal (hypodermal) cells within punctate structures located at or near the apical plasma membrane. Through whole genome sequencing, we identified two additionalnekl-2; nekl-3suppressor strains containing coding-altering mutations incatp-1but found that neither mutation, when introduced intonekl-2; nekl-3mutants using CRISPR methods, was sufficient to elicit robust suppression of molting defects. Our data also suggested that the twocatp-1isoforms,catp-1aandcatp-1b, may in some contexts be functionally redundant. On the basis of previously published studies, we tested the hypothesis that loss ofcatp-1may suppressnekl-associated defects by inducing partial entry into the dauer pathway. Contrary to expectations, however, we failed to obtain evidence that loss ofcatp-1suppressesnekl-2; nekl-3defects through a dauer-associated mechanism or that loss ofcatp-1leads to entry into the pre-dauer L2d stage. As such, loss ofcatp-1may suppressnekl-associated molting and membrane trafficking defects by altering electrochemical gradients within membrane-bound compartments.
MAGNETICALLY-DRIVEN HYDROGEL SURFACES FOR DYNAMIC STIFFNESS MODULATION FOR MODULATING MACROPHAGE BEHAVIOR
Authors: Li Lanhui , Alsema Els , Beijer Nick R.M. , Gumuscu Burcu
Published: March 2024

During the host response towards implanted biomaterials, macrophages can shift phenotype rapidly upon changes in their microenvironment within the host tissue. Exploration of this phenomenon can gain significantly from the development of adequate tools. Creating dynamic surface alterations on classical hydrogel substrates presents challenges, particularly when integrating them with cell cultivation and monitoring processes. However, having the capability to dynamically manipulate the stiffness of biomaterial surfaces holds significant potential. We introduce magnetically actuated dynamic surfaces (MadSurface) tailored to induce reversible stiffness changes on polyacrylamide hydrogel substrates with embedded magnetic microparticles in a time-controllable manner. Our investigation focused on exploring the potential ofMadSurface in dynamically modulating macrophage behavior in a programmable manner. We achieved a consistent modulation by subjecting theMadSurface to a pulsed magnetic field with a frequency of 0.1 Hz and a magnetic field flux density of 50 mT and analyzed exposed cells using flow cytometry and ELISA. At the single cell level, we identified a sub-population for which the dynamic stiffness conditions in conjunction with the pulsed magnetic field increased the expression of CD206 in M1-activated THP-1 cells, indicating a consistent shift toward M2 anti-inflammatory phenotype onMadSurface. At the population level, this effect was mostly hindered in the first 24 hours.MadSurface approach can create controlled environments to advance our understanding of the interplay between dynamic surface mechanics and macrophage behavior.
Heterogeneous and higher-order cortical connectivity undergirds efficient, robust and reliable neural codes
Authors: Egas Santander Daniela , Pokorny Christoph , Ecker András , Lazovskis Jānis , Santoro Matteo , Smith Jason P. , Hess Kathryn , Levi Ran , Reimann Michael W.
Published: March 2024

Simplified models of neural networks have demonstrated the importance of establishing a reasonable tradeoff between memory capacity and fault-tolerance in cortical coding schemes. The intensity of the tradeoff is mediated by the level of neuronal variability. Indeed, increased redundancy in neuronal activity enhances the robustness of the code at the cost of the its efficiency. We hypothesized that the heterogeneous architecture of biological neural networks provides a substrate to regulate this tradeoff, thereby allowing different subpopulations of the same network to optimize for different objectives. To distinguish between subpopulations, we developed a metric based on the mathematical theory of simplicial complexes that captures the complexity of their connectivity, by contrasting its higher-order structure to a random control. To confirm the relevance of our metric we analyzed several openly available connectomes, revealing that they all exhibited wider distributions of simplicial complexity across subpopulations than relevant controls. Using a biologically detailed cortical model and an electron microscopic data set of cortical connectivity with co-registered functional data, we showed that subpopulations with low simplicial complexity exhibit efficient activity. Conversely, subpopulations of high simplicial complexity play a supporting role in boosting the reliability of the network as a whole, softening the robustness-efficiency tradeoff. Crucially, we found that both types of subpopulations can and do coexist within a single connectome in biological neuronal networks, due to the heterogeneity of their connectivity. Our work thus suggests an avenue for resolving seemingly paradoxical previous results that assume homogeneous connectivity.
Widespread innervation of motoneurons by spinal V3 neurons globally amplifies locomotor output in mice
Authors: Zhang Han , Deska-Gauthier Dylan , MacKay Colin S. , Hari Krishnapriya , Lucas-Osma Ana M. , Borowska-Fielding Joanna , Letawsky Reese L. , Akay Turgay , Fenrich Keith K. , Bennett David J. , Zhang Ying
Published: March 2024

While considerable progress has been made in understanding the neuronal circuits that underlie the patterning of locomotor behaviours such as walking, less is known about the circuits that amplify motoneuron output to enable adaptable increases in muscle force across different locomotor intensities. Here, we demonstrate that an excitatory propriospinal neuron population (V3 neurons, Sim1+) forms a large part of the total excitatory interneuron input to motoneurons (∼20%) across all hindlimb muscles. Additionally, V3 neurons make extensive connections among themselves and with other excitatory premotor neurons (such as V2a neurons). These circuits allow local activation of V3 neurons at just one segment (via optogenetics) to rapidly depolarize and amplify locomotor-related motoneuron output at all lumbar segments in both the in vitro spinal cord and the awake adult mouse. Interestingly, despite similar innervation from V3 neurons to flexor and extensor motoneuron pools, functionally, V3 neurons exhibit a pronounced bias towards activating extensor muscles. Furthermore, the V3 neurons appear essential to extensor activity during locomotion because genetically silencing them leads to slower and weaker mice with a poor ability to increase force with locomotor intensity, without much change in the timing of locomotion. Overall, V3 neurons increase the excitability of motoneurons and premotor neurons, thereby serving as global command neurons that amplify the locomotion intensity.
HUHgle: An Interactive Substrate Design Tool for Covalent Protein-ssDNA Labeling Using HUH-tags
Authors: Smiley Adam T. , Babilonia-Díaz Natalia S. , Hughes Aspen J. , Lemmex Andrew C.D. , Anderson Michael J.M. , Tompkins Kassidy J. , Gordon Wendy R.
Published: March 2024

HUH-tags have emerged as versatile fusion partners that mediate sequence specific protein-ssDNA bioconjugation through a simple and efficient reaction. Here we present HUHgle, a python-based interactive tool for the visualization, design, and optimization of substrates for HUH-tag mediated covalent labeling of proteins of interest with ssDNA substrates of interest. HUHgle streamlines design processes by integrating an intuitive plotting interface with a search function capable of predicting and displaying protein-ssDNA bioconjugate formation efficiency and specificity in proposed HUH-tag/ssDNA sequence combinations. Validation demonstrates that HUHgle accurately predicts product formation of HUH-tag mediated bioconjugation for single- and orthogonal-labeling reactions. In order to maximize the accessibility and utility of HUHgle, we have implemented it as a user-friendly Google Colab notebook which facilitates broad use of this tool, regardless of coding expertise.

Graphical Abstract
Cell-type specialization of layer 5 excitatory neurons in tactile behavior
Authors: King Samson G. , Maire Phillip , Mergenthal Adam , Walker Stefanie , Hires Samuel Andrew
Published: March 2024

Layer 5 is the canonical output layer of sensory cortex. The two most numerous neural constituents of Layer 5 are pyramidal tract (PT) and intratelencephalic (IT) neurons. These output cell classes combine diverse sets of inputs and project to distinct locations across the brain, suggesting differing roles in sensory information processing. Here, we investigated the representation of touch and whisker motion in these two cell types within primary somatosensory cortex (S1) using optogenetically targeted single unit electrophysiology during whisker-guided object localization. PT neurons (N = 32) had much higher spike rates than IT (N=26) during behavior. Individual members of both were modulated by, but average population firing rates were stable between quiet and whisking periods. PT neurons showed greater absolute spike rate changes, but less relative modulation than IT neurons to whisking kinematic features. Touch-excited PT (N = 18) and IT neurons (N = 8) rapidly adapted to active touch. Both populations encoded the azimuthal position of touched objects, with IT neurons more sharply tuned to position. However, position was more precisely decodable from PT population activity, due to greater evoked spikes per touch. A consequence of these characteristics is that PT neurons, with their higher firing rates, may be more effective participants in rate-based neural codes, while IT neurons, with their sharp modulation, may be more effective in timing or synchrony-based codes.
Redox-dependent synaptic clustering of gephyrin
Authors: Gehling Maria-Theresa , Liebsch Filip , Jacobs Lianne , Riemer Jan , Schwarz Günter
Published: March 2024

Reactive oxygen species (ROS) play a central role in enhancing inhibitory signal transmission, thus extending their role beyond oxidative stress in disease and aging. However, the underlying molecular mechanisms mediating these functions have remained elusive. At inhibitory synapses, the scaffolding protein gephyrin clusters glycine and GABA type A receptors. Since gephyrin harbors multiple surface-exposed cysteines, we investigated the regulatory influence of ROS on gephyrin. We show that H2O2-induced oxidation of gephyrin cysteines triggered reversible, synaptic multimerization through disulfide bridge formation, which provided more receptor binding sites, lead to proteolytic protection and enhanced liquid-liquid phase separation. We identified mitochondria-derived ROS as a physiological source and observed oxidized gephyrin multimersin vivo,indicating that gephyrin can be regulated by the redox environment. Collectively, our findings suggest that cysteines in gephyrin modulate synaptic localization and clustering as regulatory redox-switches thereby establishing a link between neuronal and mitochondrial activity.