bluuebunny/crossref_metadata_2025 huggingface dataset:
Human Foot Force Informs Balance Control Strategies when Standing on a Narrow Beam
Authors: Shiozawa Kaymie , Russo Marta , Lee Jongwoo , Hogan Neville , Sternad Dagmar
Published: March 2024

Despite the abundance of studies on the control of standing balance, insights about the roles of biomechanics and neural control have been limited. Previous work introduced an analysis combining the direction and orientation of ground reaction forces. The “intersection point” of the lines of actions of these forces exhibited a consistent pattern across healthy, young subjects when computed for different frequency components of the center of pressure signal. To investigate the control strategy of quiet stance, we applied this intersection point analysis to experimental data of 15 healthy, young subjects balancing in tandem stance on a narrow beam and on the ground. Data from the sagittal and frontal planes were analyzed separately. The task was modeled as a double-inverted pendulum controlled by an optimal controller with torque-actuated ankle and hip joints and additive white noise. To test our prediction that the controller that minimized overall joint effort would yield the best fit across the tested conditions and planes of analyses, experimental results were compared to simulation outcomes. The controller that minimized overall effort produced the best fit in both balance conditions and planes of analyses. For some conditions, the relative penalty on the hip and ankle joints varied in a way relevant to the balance condition or to the plane of analysis. These results suggest that unimpaired quiet balance in a challenging environment can be best described by a controller that maintains minimal effort through the adjustment of relative ankle and hip joint torques.

NEW & NOTEWORTHY

This study explored balance control in humans during a challenging task using the novel intersection point analysis, based on ground reaction force direction and point of application. Experimental data of subjects standing on a narrow beam in tandem stance were compared with modeling results of a double-inverted pendulum. The analysis showed that individuals minimized effort by adjusting ankle and hip torques, shedding light on the interplay of biomechanics and neural control in maintaining balance.
High-throughput algorithm predicts F-Type ATP synthase rotor ring stoichiometries of 8 to 27 protomers
Authors: Osipov Stepan D. , Zinovev Egor V. , Anuchina Arina A. , Kuzmin Alexander S. , Minaeva Andronika V. , Ryzhykau Yury L. , Vlasov Alexey V. , Gushchin Ivan Yu.
Published: March 2024

ATP synthases are large enzymes present in every living cell. They consist of a transmembrane and a soluble domain, each comprising multiple subunits. The transmembrane part contains an oligomeric rotor ring (c-ring), whose stoichiometry defines the ratio between the number of synthesized ATP molecules and the number of ions transported through the membrane. Currently, c-rings of F-Type ATP synthases consisting of 8 to 17 (except 16) subunits have been experimentally demonstrated. Here, we present an easy-to-use high-throughput computational approach based on AlphaFold that allows us to estimate the stoichiometry of all homooligomeric c-rings, whose sequences are present in genomic databases. We validate the approach on the available experimental data, obtaining the correlation as high as 0.94 for the reference data set, and use it to predict the existence of c-rings with stoichiometry varying from 8 to 27. We then conduct molecular dynamics simulations of two c-rings with stoichiometry above 17 to corroborate the machine learning-based predictions. Our work strongly suggests existence of rotor rings with previously undescribed high stoichiometry in natural organisms and highlights the utility of AlphaFold-based approaches for studying homooligomeric proteins.
Using machine learning to count Antarctic shag (Leucocarbo bransfieldensis) nests on images captured by Remotely Piloted Aircraft Systems
Authors: Cusick Andrew , Fudala Katarzyna , Storożenko Piotr Pasza , Świeżewski Jędrzej , Kaleta Joanna , Oosthuizen W. Chris , Pfeifer Christian , Bialik Robert Józef
Published: March 2024

Using 51 orthomosaics of 11 breeding locations of the Antarctic shag, we propose a method for automating counting of shag nests. This is achieved by training an object detection model based on the YOLO architecture and identifying nests on sections of the orthomosaic, which are later combined with predictions for the entire orthomosaic. Our results show that the current use of Remotely Piloted Aircraft Systems (RPAS) to collect images of areas with shag colonies, combined with machine learning algorithms, can provide reliable and fast estimates of shag nest counts (F1 score > 0.95). By using data from only two shag colonies for training, we show that models can be obtained that generalise well to images of both spatially and temporally distinct colonies. The proposed practical application opens the possibility of using aerial imagery to perform large-scale surveys of Antarctic islands in search of undiscovered shag colonies. We discuss the conditions for optimal performance of the model as well as its limitations. The code, data and trained model allowing for full reproducibility of the results are available athttps://github.com/Appsilon/Antarctic-nests.
Rare SNP in theHELBgene interferes with RPA interaction and cellular function of HELB
Authors: Osei Bertha , May Benjamin H. , Stiefel Clara M. , West Kirk L. , Zafar Maroof Khan , Thompson Matthew D. , Bergstrom Erik , Leung Justin W. , Enemark Eric J. , Byrd Alicia K.
Published: March 2024

HELB is a human helicase involved in initiation of DNA replication, the replication stress response, and regulation of double-strand DNA break repair. rs75770066 is a rare SNP in the HELB gene that affects age at natural menopause. rs75770066 results in a D506G substitution in an acidic patch within the 1A domain of the helicase that is known to interact with RPA. We found that this amino acid change dramatically impairs the cellular function of HELB. D506G-HELB exhibits impaired interaction with RPA, which likely results in the effects of rs75770066 as this reduces recruitment of HELB to sites of DNA damage. Reduced recruitment of D506G-HELB to double-strand DNA breaks and the concomitant increase in homologous recombination likely alters the levels of meiotic recombination, which affects the viability of gametes. Because menopause occurs when oocyte levels drop below a minimum threshold, altered repair of meiotic double-stranded DNA breaks has the potential to directly affect the age at natural menopause.
Single-nucleus Multiomic Analyses Identifies Gene Regulatory Dynamics of Phenotypic Modulation in Human Aneurysmal Aortic Root
Authors: Liu Xuanyu , Zeng Qingyi , Yang Hang , Li Wenke , Chen Qianlong , Yin Kunlun , Pan Zihang , Wang Kai , Luo Mingyao , Shu Chang , Zhou Zhou
Published: March 2024

Aortic root aneurysm is a potentially life-threatening condition that may lead to aortic rupture and is often associated with genetic syndromes, such as Marfan syndrome (MFS). Although studies with MFS animal models have provided valuable insights into the pathogenesis of aortic root aneurysms, our understanding of the transcriptomic and epigenomic landscape in human aortic root tissue remains incomplete. This knowledge gap has impeded the development of effective targeted therapies. Here, this study performs the first integrative analysis of single-nucleus multiomic (gene expression and chromatin accessibility) and spatial transcriptomic sequencing data of human aortic root tissue under healthy and MFS conditions. Cell-type-specific transcriptomic and cis-regulatory profiles in the human aortic root are identified. Regulatory and spatial dynamics during phenotypic modulation of vascular smooth muscle cells (VSMCs), the cardinal cell type, are delineated. Moreover, candidate key regulators driving the phenotypic modulation of VSMC, such asFOXN3,TEAD1,BACH2, andBACH1, are identified.In vitroexperiments demonstrate that FOXN3 functions as a novel key regulator for maintaining the contractile phenotype of human aortic VSMCs through targeting ACTA2. These findings provide novel insights into the regulatory and spatial dynamics during phenotypic modulation in the aneurysmal aortic root of humans.
The Predictive Value of Diaphragm Muscle Ultrasound for Ventilator Weaning Outcomes after Cervical Spinal Cord Injury: A Retrospective Case Series
Authors: Bhatia Natasha S. , Kunzweiler Stephany , Conley Christopher , Kim Ki H. , Adewuyi Adenike A. , Mondriguez-Gonzalez Antonio , Wolfe Lisa F. , Kwasny Mary , Franz Colin K.
Published: March 2024

Objectives

Neuromuscular respiratory failure after cervical spinal cord injury (cSCI) can lead to dependence on an invasive mechanical ventilator. Ventilator-free breathing after cSCI is associated with improved morbidity, mortality, and quality of life. We investigated the use of diaphragm muscle ultrasound to predict ventilator weaning outcomes after cSCI.

Methods

This is a retrospective case series conducted at a university-affiliated freestanding inpatient rehabilitation facility. We identified patients with cSCI who had a tracheostomy and were dependent on an invasive mechanical ventilator at the time of admission to inpatient rehabilitation. A diaphragm muscle ultrasound was performed, which included measurements of the thickness of the diaphragm and a calculation of the thickening ratio (TR), which reflects diaphragm muscle contraction. The primary outcome measure was the need for mechanical ventilation at time of discharge from the inpatient rehabilitation facility. Successful ventilator weaning was defined as either daytime or full 24-hour ventilator-free breathing.

Results

Of the 21 patients enrolled, 11 (52%) were able to wean successfully (partially or fully) from the ventilator. Of the ultrasound measurements that were taken, the TR was the optimal predictor for ventilator weaning outcomes. A threshold of TR≥1.2 as the maximum hemidiaphragm measurement had a sensitivity of 1.0 and specificity of 0.90 for predicting ventilator weaning.

Conclusion

Normal diaphragm contractility (TR ≥ 1.2) as determined by diaphragm muscle ultrasound is a strong positive predictor for successful ventilator weaning in patients with cSCI.

Utilizing diaphragm ultrasound, rehabilitation physicians can set precision rehabilitation goals regarding ventilator weaning for inpatients with respiratory failure after cSCI, potentially improving both outcomes and quality of life.
Sex-Specific Effect of MTSS1 Downregulation on Dilated Cardiomyopathy
Authors: Choe Dongwook , Burke Megan , Brandimarto Jeffrey A , Marti-Pamies Ingrid , Yob Jaime , Yang Yifan , Morley Michael P , Drivas Theodore G. , Day Sharlene , Damrauer Scott , Wang Xiao , Cappola Thomas P
Published: March 2024

MTSS1 (metastasis suppressor 1) is an I-BAR protein that regulates cytoskeleton dynamics through interactions with actin, Rac, and actin-associated proteins. In a prior study, we identified genetic variants in a cardiac-specific enhancer upstream ofMTSS1that reduce human left ventricular (LV) MTSS1 expression and associate with protection against dilated cardiomyopathy (DCM). We sought to probe these effects further using population genomics andin vivomurine models. We crossedMtss1-/-mice with a transgenic (Tg) murine model of human DCM caused by the D230N pathogenic mutation inTpm1(tropomyosin 1). In females,Tg/Mtss1+/-mice had significantly increased LV ejection fraction and reduced LV volumes relative to theirTg/Mtss1+/+counterparts, signifying partial rescue of DCM due toMtss1haploinsufficiency. No differences were observed in males. To study effects in humans, we fine-mapped theMTSS1locus with 82 cardiac magnetic resonance (CMR) traits in 22,381 UK Biobank participants.MTSS1enhancer variants showed interaction with biological sex in their associations with several CMR traits. After stratification by biological sex, associations with CMR traits and colocalization withMTSS1expression in the Genotype-Tissue Expression (GTEx) Project were observed principally in women and were substantially weaker in men. These findings suggest sex dimorphism in the effects of MTSS1-lowering alleles, and parallel the increased LV ejection fraction and reduced LV volumes observed femaleTg/Mtss1+/-mice. Together, our findings at theMTSS1locus suggest a genetic basis for sex dimorphism in cardiac remodeling and motivate sex-specific study of common variants associated with cardiac traits and disease.
Metabolic phenotyping of healthy and diseased human RPE cells
Authors: Rizwan Saira , Toothman Beverly , Li Bo , Engel Abbi J. , Lim Rayne R. , Niernberger Sheldon , Lu Jinyu , Ratliff Cloe , Xiang Yinxiao , Eminhizer Mark , Chao Jennifer R. , Du Jianhai
Published: March 2024

Purpose

Metabolic defects in the retinal pigment epithelium (RPE) underlie many retinal degenerative diseases. This study aims to identify the nutrient requirements of healthy and diseased human RPE cells.

Methods

We profiled nutrient utilization of various human RPE cells, including differentiated and dedifferentiated fetal RPE (fRPE), induced pluripotent stem cell derived-RPE (iPSC RPE), Sorsby fundus dystrophy (SFD) patient-derived iPSC RPE, CRISPR-corrected isogenic SFD (cSFD) iPSC RPE, and ARPE-19 cell lines using Biolog Phenotype MicroArray Assays.

Results

Differentiated fRPE cells and healthy iPSC RPE cells can utilize 51 and 48 nutrients respectively, including sugars, intermediates from glycolysis and tricarboxylic acid (TCA) cycle, fatty acids, ketone bodies, amino acids, and dipeptides. However, when fRPE cells lose their epithelial phenotype through dedifferentiation, nutrient utilization becomes restricted to 17 nutrients, primarily sugar and glutamine-related amino acids. SFD RPE cells can utilize 37 nutrients; however, compared to cSFD RPE and healthy iPSC RPE, they are unable to utilize lactate, some TCA cycle intermediates, and short-chain fatty acids. Nonetheless, they show increased utilization of branch-chain amino acids (BCAAs) and BCAA-containing dipeptides. Dedifferentiated ARPE-19 cells grown in traditional culture media cannot utilize lactate and ketone bodies. In contrast, nicotinamide supplementation promotes differentiation towards an epithelial phenotype, restoring the ability to use these nutrients.

Conclusions

Epithelial phenotype confers metabolic flexibility to healthy RPE for utilizing various nutrients. SFD RPE cells have reduced metabolic flexibility, relying on the oxidation of BCAAs. Our findings highlight the potentially important roles of nutrient availability and utilization in RPE differentiation and diseases.
Erythropoietin production in embryonic neural cells is controlled by hypoxia-inducible factors and histone deacetylases in an undifferentiated state
Authors: Iwamura Yuma , Nakai Taku , Kato Koichiro , Ishioka Hirotaka , Yamamoto Masayuki , Hirano Ikuo , Suzuki Norio
Published: March 2024

During mammalian development, production sites of the erythroid growth factor erythropoietin (EPO) shift from the neural tissues to the liver in embryos and to the adult kidneys. Embryonic neural EPO-producing (NEP) cells, a subpopulation of neuroepithelial and neural crest cells, express theEpogene between embryonic day (E) 8.5 and E11.5 to promote primitive erythropoiesis in mice. WhileEpogene expression in the liver and kidney is induced under hypoxic conditions through hypoxia-inducible transcription factor (HIF) 2α, theEpogene regulatory mechanisms in NEP cells remain to be elucidated. This study confirms the presence of cells coexpressing the genes encoding EPO and HIF2α in E9.5 neural tubes, where the hypoxic microenvironment activates HIF1α. In human neural progenitors and mouse embryonic neural tissues, a HIF-activating compound upregulatedEPOexpression, and this induction was blocked by inhibiting HIFs. Additionally, a cell line of NEP cell derivatives that no longer expressed theEpogene demonstrated that histone deacetylase inhibitors (HDACIs) reactivate EPO production while rejuvenating the cells. HDACIs also inducedEPOgene expression in SK-N-BE(2)c human neuroblastoma cells and mouse primary neural crest cells. Thus, EPO production is controlled by epigenetic mechanisms and hypoxia signaling in an immature state of hypoxic NEP cells.
Under the radar: differential responses of bed bugs to an entomopathogen, environmental bacteria, and a human pathogen
Authors: Walt Hunter K. , Bronzato-Badial Aline , Maedo Sophie E. , Hinton Joseph A. , King Jonas G. , Pietri Jose E. , Hoffmann Federico G.
Published: March 2024

Background

Bed bugs (Hemiptera: Cimicidae) are a widely distributed, obligately blood-feeding insect, but they have never been linked to pathogen transmission in humans. Most other hematophagous insects that frequently bite humans transmit pathogens, and it is unclear why bed bugs do not. One hypothesis is that bed bugs have evolved a highly robust immune system because their mating system, traumatic insemination, exposes females to consistent wounding and bacterial infections. Although this has been proposed, very little is known about the bed bug immune system and how bed bugs respond to microbial challenges. Understanding the bed bug immune system could give insight to why bed bugs are not known to transmit disease and under what circumstances they could, while also facilitating biological control efforts involving microbes.

Methods

To investigate the immune response of bed bugs to bacterial challenges, we exposed female bed bugs to three bacterial challenges. 1.)Pseudomonas fluorescens, an entomopathogen known to have harmful effects to bed bugs, 2.) bacteria cultured from a bed bug enclosure likely encountered during traumatic insemination, and 3.)Borrelia duttoni, a human vector-borne pathogen that causes relapsing fever. We compared the transcriptomes of infected bed bugs with uninfected bed bugs, focusing on immune-related genes. We also conducted phylogenetic analyses to understand patterns of gene duplication and function of potentially immune-related genes.

Results

We found many known immune effector genes upregulated in response toP. fluorescensand traumatic insemination-associated bacteria, but interestingly, not in response toB. duttoni. Furthermore, we found significant overlap in the genes differentially expressed in response toP. fluorescensand the traumatic insemination associated bacteria, and betweenP. fluorescensandB. duttoni, but no significant overlap between traumatic insemination bacteria andB. duttoni. We also show that bed bug diptericin-like antimicrobial peptides underwent a lineage-specific gene duplication, and that they may have further functional specialization. Finally, we identify previously overlooked candidates for future study of immune function in bed bugs, including some putative cuticle-associated genes, a laccase-like gene, and a mucin-like gene.

Conclusions

By taking comprehensive transcriptomic approach, our study is an important step in understanding how bed bugs respond to diverse immune challenges.
Rewinding the developmental tape shows how bears break a developmental rule
Authors: Stenberg Otto E. , Moustakas-Verho Jacqueline E. , Jernvall Jukka
Published: March 2024

Mammals have evolved a broad variety of dental morphologies. Nevertheless, the development of the mammalian dentition is considered highly conserved. Molar size proportions exemplify this as a system where small changes in shared developmental mechanisms yield a defined range of morphological outcomes. The Inhibitory Cascade (IC) model states that as molars develop in a sequence, the first developing anterior molars inhibit the development of subsequent posterior ones. The IC model thus predicts a trend of linear tooth size change along the molar row, as has been observed in a wide range of mammalian taxa with otherwise differing dental morphology. Perhaps the starkest exceptions to the IC rule are bears, in which the second molar is the largest and the third molar is disproportionally small. Here we sought to illuminate when and how during development the bear dentition falls of the IC prediction. We examined molar proportions in seven bear species. The results indicate that development of bear molars deviates from IC expectation already during patterning. Yet, during the earlier cap stage, size proportions of bear molars still seem to adhere to the IC model predictions. Overall, these analyses are suggestive that irrespective of the final outcome, the process of initial splitting of the molar-forming region into individual teeth is conserved and follows the IC rule.
Direct pharmacological AMPK activation inhibits mucosal SARS-CoV-2 infection by reducing lipid metabolism, restoring autophagy flux and the type I IFN response
Authors: Cottignies-Calamarte Andrea , Marteau Flora , He Feifan , Belouzard Sandrine , Dubuisson Jean , Tudor Daniela , Viollet Benoit , Bomsel Morgane
Published: March 2024

AMP-activated protein kinase (AMPK) plays a central role in regulating cell energy balance. When activated, AMPK supresses energy-consuming pathways such as lipid and protein synthesis while increasing nutrient availability through the activation of autophagy. These pathways downstream AMPK activation contribute to SARS-CoV-2 infection, which hijacks autophagy and accumulates lipid droplets in viral factories to support viral replication. Here, we assessed the antiviral activity of the direct pan-AMPK allosteric activator MK-8722in vitro.MK-8722 efficiently inhibited infection of Alpha and Omicron SARS-CoV-2 variants in Vero76 and human bronchial epithelial Calu-3 cells at micromolar concentration. This inhibition relied on restoring the autophagic flux, which redirected newly synthesized viral proteins for degradation, and on a reduction in lipid metabolism, which affected the viral factories. Furthermore, MK-8722 treatment increased the type I interferon (IFN-I) response. Post-infection treatment with MK-8722 was enough to inhibit efficiently viral replication and restore the IFN-I response. Finally, MK-8722 treatment did not alter the SARS-CoV-2-specific CD8+T cell response mounted upon Spike vaccination. Overall, by activating AMPK, MK-8722 acts as an effective antiviral against SARS-CoV-2 infection, even when applied post-exposure, paving the way for preclinical tests aimed at inhibiting viral replication and improving patients’ symptoms.

Graphical abstract

Highlights

MK-8722 exerts post-exposure antiviral activity

MK-8722 induces a decrease in cellular lipid content

MK-8722 promotes an increase in the autophagic flux of viral components

MK-8722 promotes the restoration of the IFN-I activity

MK-8722 antiviral activity is compatible with virus-specific T cell responses
Siderophore piracy enables the nasal commensalStaphylococcus lugdunensisto antagonize the pathogenStaphylococcus aureus
Authors: Rosenstein Ralf , Salazar Benjamin O. Torres , Sauer Claudia , Heilbronner Simon , Krismer Bernhard , Peschel Andreas
Published: March 2024

Bacterial pathogens such asStaphylococcus aureuscolonize body surfaces of part of the human population, which represents a critical risk factor for skin disorders and invasive infections. However, such pathogens do not belong to the human core microbiomes – beneficial ‘commensal’ bacteria can often prevent the persistence of certain pathogens – using molecular strategies that are only superficially understood. We recently reported that the commensal bacteriumStaphylococcus lugdunensisproduces the novel antibiotic lugdunin, which eradicatesS. aureusfrom nasal microbiomes of hospitalized patients. However, it has remained unclear ifS. lugdunensismay affectS. aureuscarriage in the general population and howS. lugdunensiscarriage could be promoted to enhance itsS. aureus-eliminating capacity. We could cultivateS. lugdunensisfrom the noses of 6.3% of healthy human volunteers. In addition,S. lugdunensisDNA could be identified in metagenomes of many culture-negative nasal samples indicating that cultivation success depends on a specific bacterial threshold density. HealthyS. lugdunensiscarriers had a 5.2-fold lower propensity to be colonized byS. aureusindicating that lugdunin can eliminateS. aureusalso in healthy humans.S. lugdunensis-positive microbiomes were dominated by eitherStaphylococcus epidermidis,Corynebacteriumspecies, orDolosigranulum pigrum. These and further bacterial commensals, whose abundance was positively associated withS. lugdunensis, promotedS. lugdunensisgrowth in co-culture. Such mutualistic interactions depended on the production of iron-scavenging siderophores by supporting commensals, and on the capacity ofS. lugdunensisto import siderophores. These findings underscore the importance of microbiome homeostasis for eliminating pathogen colonization. Elucidating mechanisms that drive microbiome interactions will become crucial for microbiome-precision editing approaches.
Differential gene expression and chromatin accessibility in the medial prefrontal cortex associated with individual differences in rat behavioral models of opioid use disorder
Authors: Liu Shirelle X. , Muelken Peter , Maxim Zia L. , Ramakrishnan Aarthi , Estill Molly S. , LeSage Mark G. , Smethells John R. , Shen Li , Tran Phu V. , Harris Andrew C. , Gewirtz Jonathan C.
Published: March 2024

Opioid use disorder (OUD) is a neuropsychological disease that has a devastating impact on public health. Substantial individual differences in vulnerability exist, the neurobiological substrates of which remain unclear. To address this question, we investigated genome-wide gene transcription (RNA-seq) and chromatin accessibility (ATAC-seq) in the medial prefrontal cortex (mPFC) of male and female rats exhibiting differential vulnerability in behavioral paradigms modeling different phases of OUD: Withdrawal-Induced Anhedonia (WIA), Demand, and Reinstatement. Ingenuity Pathway Analysis (IPA) of RNA-seq revealed greater changes in canonical pathways in Resilient (vs. Saline) rats in comparison to Vulnerable (vs. Saline) rats across 3 paradigms, suggesting brain adaptations that might contribute to resilience to OUD across its trajectory. Analyses of gene networks and upstream regulators implicated processes involved in oligodendrocyte maturation and myelination in WIA, neuroinflammation in Demand, and metabolism in Reinstatement. Motif analysis of ATAC-seq showed changes in chromatin accessibility to a small set of transcription factor (TF) binding sites as a function either of opioid exposure (i.e., morphine versus saline) generally or of individual vulnerability specifically. Some of these were shared across the 3 paradigms and others were unique to each. In conclusion, we have identified changes in biological pathways, TFs, and their binding motifs that vary with paradigm and OUD vulnerability. These findings point to the involvement of distinct transcriptional and epigenetic mechanisms in response to opioid exposure, vulnerability to OUD, and different stages of the disorder.
Gluk4-containing kainate receptors regulate synaptic communication in the motor cortex and reduce axon degeneration in adult mice
Authors: Ricci Raphael , Fletcher Jessica L , Makowiecki Kalina , Pepper Renee E , Fortune Alastair , Cullen Carlie L , Connelly William M , Charlesworth Jac , Blackburn Nicholas B , Pitman Kimberley A , Young Kaylene M
Published: March 2024

Glutamate-gated kainate receptors comprising the Gluk4 subunit (encoded byGrik4) are highly expressed by neurons in the central nervous system. We report thatGrik4mRNA is widely expressed by neurons in the adult mouse motor cortex, where GluK4-containing kainate receptors account for ∼60% of the kainate evoked current in layer V pyramidal neurons. To elucidate their role in motor circuit regulation, we analysed the behaviour of mice that lacked the pore forming domain of the GluK4 subunit (Grik4-/-mice).Grik4-/-mice were hyperactive, had an abnormal gait, and impaired motor coordination. At postnatal day (P)60, layer V pyramidal neurons received fewer miniature excitatory post synaptic currents, had a reduced density of thin spines on their basal dendrites, and a reduced density of VGlut1 puncta at the soma, consistent with neurons receiving fewer excitatory synaptic connections.Grik4-/-mice also lost ∼44% of their callosal axons between P60 and P180 and the amplitude of the callosal compound action potential was reduced by ∼25-30%. RNA sequencing data support the capacity forGrik4to modulate synaptic and neuroprotective signalling pathways.
Label-free imaging of intracellular structures in living mammalian cells via external apodization phase-contrast microscopy
Authors: Ohno Hiroshi , Nishimura Takenori , Kainoh Kenta , Ohashi Yoshitaka , Onodera Naoko , Kano Mayuko , Sari Lay Nurhana , Masuda Masato , Tamura Yoshiaki , Hayashi Yusuke , Yamamoto Yusuke , Takahashi Shin-Ichiro , Mishima Yuta , Yoneyama Yosuke , Takeuchi Yoshinori , Sekiya Motohiro , Matsuzaka Takashi , Miyamoto Takafumi , Shimano Hitoshi
Published: March 2024

Developing techniques to visualize intracellular structures, which influence the spatiotemporal functionality of biomolecules, is essential for elucidating mechanisms governing cellular behavior. In this study, we demonstrate that label-free external apodization phase-contrast (ExAPC) microscopy serves as a valuable tool for the simultaneous observation of various intracellular structures with high spatiotemporal resolution, while successfully mitigating halo artifacts. Additionally, through quantitative analysis of images obtained by combining ExAPC microscopy with fluorescence microscopy, we identified distinct heterogeneities in biomolecular condensates, lipid droplets, and mitochondria. Our findings highlight the potential of ExAPC microscopy to provide detailed insights into alterations in intracellular structures associated with diverse cellular processes, corroborating the existing knowledge and potentially contributing to the discovery of novel cellular mechanisms.
Methionine cycle in a pair of serotonergic neurons regulates diet-dependent behavior and longevity through a neuron-gut signaling
Authors: Rahman Sabnam Sahin , Bhattacharjee Shreya , Prakash Govind , Motwani Simran , Nair Tripti , Keerthana Rachamadugu Sai , Mukhopadhyay Arnab
Published: March 2024

The folate-methionine cycle (Met-C) is a central metabolic pathway that is regulated by vitamin B12 (B12), a micronutrient obtained exclusively from diet and microbiota. This metabolic hub supports amino acid, nucleotide and lipid biosynthesis apart from its central role of providing one carbon (-CH3) moiety for methylation reactions. While deficiency of B12 as well as polymorphism in enzymes of the Met-C has been clinically attributed to neurological and metabolic disorders, how this pathway cell non-autonomously regulates systemic physiological processes is less understood. Using a B12-sensitive mutant ofCaenorhabditis elegans, we show that the neuronal Met-C responds to differential B12 content in diet to regulate p38-MAPK activation in intestinal cells, thereby modulating cytoprotective gene expression, stress tolerance and longevity. Mechanistically, B12-driven changes in the metabolic flux through the Met-C in the serotonergic ADF neurons of the mutant lead to the release of serotonin (5-hydroxytryptamine, 5-HT). 5-HT activates its receptor, MOD-1, in the post-synaptic interneurons that then secretes the neuropeptide FLR-2. FLR-2 binds to FSHR-1, its cognate receptor in the intestine, and induces the phase transition of the SARM domain protein TIR-1, thereby activating the p38-MAPK pathway. Importantly, this cascade influences the foraging behaviour of the mutant worms such that they prefer a B12-rich diet. Together, our study reveals a dynamic neuron-gut signaling axis that helps an organism modulate behaviour and life history traits based on the neuronal Met-C metabolic flux determined by B12 availability in its diet. Understandably, disruption of the optimum functioning of this axis may have debilitating effects on the health of an organism and the survival of the species.
Sticking together: Independent evolution of biofilm formation in different species of staphylococci has occurred multiple times via different pathways
Authors: Crossman Lisa , Sims Leanne , Dean Rachael , Felgate Heather , Calvo Teresa Diaz , Hill Claire , McNamara Iain , Webber Mark A , Wain John
Published: March 2024

Various species of staphylococci cause a wide range of infections, including implant-associated infections which are often difficult to treat due to the presence of biofilms. Whilst some proteins involved in biofilm formation are known, the differences in biofilm production between staphylococcal species remains understudied. Currently biofilm formation byStaphylococcus aureusis better understood than for other members of the genus as more research effort has focused on this species.

We assembled a panel of 385 non-aureus Staphylococcusisolates of 19 species from prosthetic joint infection as well as other clinical sources and reference strains. We assessed the biofilm forming ability of all strains using a high-throughput crystal violet assay. This identified distinct biofilm formation categories and we then compared the prevalence of Pfam domains and identified those which distinguished the categories as well as using machine learning to identify amino acid 20-mers linked to biofilm formation.

This identified some domains within proteins already positively linked to biofilm formation but we also identified important domains not previously linked to biofilm formation. RT-qPCR confirmed the expression of selected genes predicted to encode important domains within biofilms inStaphylococcus epidermidis.

The prevalence and distribution of biofilm associated domains showed a link to phylogeny, suggesting differentStaphylococcusspecies have independently evolved different mechanisms of biofilm production.

This work has identified different routes to biofilm formation in diverse species ofStaphylococcusas well as suggesting independent evolution of biofilm has occurred multiple times across the genus. Understanding the mechanisms of biofilm formation in any given species is likely to require detailed study of relevant strains and the ability to generalise across the genus may be limited.
A quantitative pipeline to assess secretion of human leptin coding variants reveals mechanisms underlying leptin deficiencies
Authors: Baird Harry J. M. , Shun-Shion Amber S. , de Oliveira Edson Mendes , Stalder Danièle , Eden Jessica , Chambers Joseph E. , Farooqi I. Sadaf , Gershlick David C. , Fazakerley Daniel J.
Published: March 2024

The hormone leptin, primarily secreted by adipocytes, plays a crucial role in regulating whole-body energy homeostasis. Homozygous loss-of-function mutations in the leptin gene(LEP)cause hyperphagia and severe obesity, primarily through alterations in leptin’s affinity for its receptor or changes in serum leptin concentrations. Although serum concentrations are influenced by various factors (e.g., gene expression, protein synthesis, stability in the serum), proper delivery of leptin from its site of synthesis in the endoplasmic reticulum via the secretory pathway to the extracellular serum is a critical step. However, the regulatory mechanisms and specific machinery involved in this trafficking route, particularly in the context of humanLEPmutations, remain largely unexplored.

We have employed the Retention Using Selective Hooks (RUSH) system to elucidate the secretory pathway of leptin. We have refined this system into a medium-throughput assay for examining the pathophysiology of a range of obesity-associatedLEPvariants. Our results reveal that leptin follows the default secretory pathway, with no additional regulatory steps identified prior to secretion.

Through screening of leptin variants, we identified three mutations that lead to proteasomal degradation of leptin and one mutant that significantly decreased leptin secretion, likely through aberrant disulfide bond formation. These observations have identified novel pathogenic effects of leptin variants, which can be informative for therapeutics and diagnostics. Finally, our novel quantitative screening platform can be adapted for other secreted proteins.
CRISPR-Cas9 editing efficiency in fission yeast is not limited by homology search and is improved by combining gap-repair with fluoride selection
Authors: Fernandez Ronan , Berro Julien
Published: March 2024

Protocols for CRISPR-Cas9 editing have been implemented in most model organisms, including fission yeast, for which some improvements have also been later described. Here, we report an improvement to the CRISPR-Cas9 protocol in fission yeast, as we combine a cloning free gap-repair method with our previously described fluoride selection marker, which speeds up genome editing. We also report a wide variability of editing efficiencies at different loci along the genome, and we demonstrate that this variability cannot be explained by the location of the edited sequences in the genome. Lastly, our attempt at improving editing efficiency by targeting the donor DNA to the cut site using a HaloTag strategy to link the donor DNA to two proteins of the homologous recombination repair machinery (Rad51 or Rad52) fell short, which shows that editing efficiency in fission yeast is likely not limited by homology search.
Targeting Iron - Respiratory Reciprocity Promotes Bacterial Death
Authors: Sharifian Gh. Mohammad , Norouzi Fatemeh , Sorci Mirco , Zaid Tanweer S , Pier Gerald B. , Achimovich Alecia , Ongwae George M. , Liang Binyong , Ryan Margaret , Lemke Michael , Belfort Georges , Gadjeva Mihaela , Gahlmann Andreas , Pires Marcos M. , Venter Henrietta , Harris Thurl E. , Laurie Gordon W.
Published: March 2024

Discovering new bacterial signaling pathways offers unique antibiotic strategies. Here, through an unbiased resistance screen of 3,884 gene knockout strains, we uncovered a previously unknown non-lytic bactericidal mechanism that sequentially couples three transporters and downstream transcription to lethally suppress respiration of the highly virulentP. aeruginosastrain PA14 - one of three species on the WHO’s ‘Priority 1: Critical’ list. By targeting outer membrane YaiW, cationic lacritin peptide ‘N-104’ translocates into the periplasm where it ligates outer loops 4 and 2 of the inner membrane transporters FeoB and PotH, respectively, to suppress both ferrous iron and polyamine uptake. This broadly shuts down transcription of many biofilm-associated genes, including ferrous iron-dependent TauD and ExbB1. The mechanism is innate to the surface of the eye and is enhanced by synergistic coupling with thrombin peptide GKY20. This is the first example of an inhibitor of multiple bacterial transporters.
Determining the rate-limiting processes for cell division inEscherichia coli
Authors: Männik Jaana , Kar Prathitha , Amarasinghe Chathuddasie , Amir Ariel , Männik Jaan
Published: March 2024

A critical cell cycle checkpoint for most bacteria is the onset of constriction when the septal peptidoglycan synthesis starts. According to the current understanding, the arrival of FtsN to midcell triggers this checkpoint inEscherichia coli. Recent structural andin vitrodata suggests that recruitment of FtsN to the Z-ring leads to a conformational switch in actin-like FtsA, which links FtsZ protofilaments to the cell membrane and acts as a hub for the late divisome proteins. Here, we investigate this putative pathway usingin vivomeasurements and stochastic cell cycle modeling. Quantitatively upregulating protein concentrations and determining the resulting division timings shows that FtsN and FtsA numbers are not rate-limiting for the division inE. coli. However, at higher overexpression levels, they affect divisions: FtsN by accelerating and FtsA by inhibiting them. At the same time, we find that the numbers of FtsZ in the cell are rate-limiting for cell divisions in addition to other processes. Altogether, these findings suggest that instead of FtsN, FtsZ protofilaments drive the conformational switch of FtsA and lead to the onset of constriction. Our data is also suggestive that FtsA minirings are not present at any significant numbers in wild-type cells.
Analysis of Infiltrating Immune Cells Following Intervertebral Disc Injury Reveals Recruitment of Gamma-Delta (γδ) T cells in Female Mice
Authors: Clayton Sade W. , Walk Remy E. , Mpofu Laura , Easson Garrett W.D. , Tang Simon Y.
Published: March 2024

Inadequate repair of injured intervertebral discs (IVD) leads to degeneration and contributes to low back pain. Infiltrating immune cells into damaged musculoskeletal tissues are critical mediators of repair, yet little is known about their identities, roles, and temporal regulation following IVD injury. By analyzing longitudinal changes in gene expression, tissue morphology, and the dynamics of infiltrating immune cells following injury, we characterize sex-specific differences in immune cell populations and identify the involvement of previously unreported immune cell types, γδ and NKT cells. Cd3+Cd4-Cd8-T cells are the largest infiltrating lymphocyte population with injury, and we identified the presence of γδ T cells in this population in female mice specifically, and NKT cells in males. Injury-mediated IVD degeneration was prevalent in both sexes, but more severe in males. Sex-specific degeneration may be associated with the differential immune response since γδ T cells have potent anti-inflammatory roles and may mediate IVD repair.

Abstract Figure

Graphical Abstract:

Schematic of the workflow to obtain longitudinal analyses of the acute IVD injury response

Injured caudal IVDs, CC5/6-CC9/10, were bilaterally punctured with a 30G needle to induce a traumatic injury. Injured IVDs and neighboring uninjured internal Control IVDs, CC12/13-CC16/17, were isolated during the acute injury response every 2-3 days until 21 days post injury (dpi) and a chronic injury time point at 42 dpi in female mice. Male mice samples were only collected at key injury response time points (red numbers): 3, 7, 12, 19, and 42 dpi. Longitudinal analyses of the temporal regulation of immune cell gene expression and cell infiltration were measured with qPCR, flow cytometry, immune fluorescence, and histology analyses to identify a sex-divergent immune response.
Omega-3 polyunsaturated fatty acids modify glucose metabolism in THP-1 monocytes
Authors: Byun Michael J. , Armon Roni , Souza Tamaris , Anderson Hope D. , Saleem Ayesha , Pauls Samantha D.
Published: March 2024

Chronic inflammation is a driving factor in diseases like obesity and type 2 diabetes. Enhanced glucose metabolism, includingviaoxidative phosphorylation, may contribute to heightened immune activation. A recent clinical trial showed that supplementation with the n-3 fatty acid α-linolenic acid (ALA) reduced oxidative phosphorylation rates in human monocytes. However, the mechanism remains unknown. Therefore, our objective was to explore the direct effects of ALA and docosahexaenoic acid (DHA) on glucose metabolism in a cell culture model and to explore the molecular mechanism.

THP-1 monocytes were treated for 48h with 10-40 μM of ALA or DHA and compared with vehicle and oleic acid controls. The Seahorse XFe24 system was used to approximate catabolic rates in the presence of glucose, including glycolysis and oxidative phosphorylation. The latter was validated by respirometry using an Oroboros O2k Oxygraph. Both ALA and DHA treatments reduced oxidative phosphorylation and increased glycolytic rates relative to control conditions. We identified pyruvate dehydrogenase kinase 4 (PDK4), an enzyme that inhibits the conversion of pyruvate to acetyl-CoA, as a possible mechanistic candidate. This gene was significantly upregulated by ALA and, to a greater extent, by DHA. Using fluorescent indicators, we also found that DHA increased reactive oxygen species while ALA had no effect.

Our data suggest that ALA and DHA trigger a re-wiring of bioenergetic pathways in monocytes, possiblyviathe upregulation of PKD4. Given the close relationship between cell metabolism and immune cell activation, this may represent a novel mechanism by which n-3 fatty acids modulate immune function and inflammation.
Investigating cross-sectional and longitudinal relationships between brain structure and distinct dimensions of externalizing psychopathology in the ABCD Sample
Authors: Propp Lee , Nakua Hajer , Bedard Anne-Claude V. , Sanches Marcos , Ameis Stephanie H. , Andrade Brendan F.
Published: March 2024

Externalizing psychopathology in childhood is a predictor of poor outcomes across the lifespan. Children exhibiting elevated externalizing psychopathology also commonly show emotion dysregulation and callous-unemotional (CU) traits. Examining cross-sectional and longitudinal neural correlates across dimensions linked to externalizing psychopathology during childhood may clarify shared or distinct neurobiological vulnerability for psychopathological impairment later in life. We used tabulated brain structure and behavioural data from baseline, year 1, and year 2 timepoints of the Adolescent Brain Cognitive Development Study (ABCD; baseline n=10,534). We fit separate linear mixed effect models to examine whether baseline brain structures in frontolimbic and striatal regions (cortical thickness or subcortical volume) were associated with externalizing symptoms, emotion dysregulation, and/or CU traits at baseline and over a two-year period. At baseline, cortical thickness in the right rostral middle frontal gyrus and bilateral pars orbitalis was positively associated with CU traits (β=|0.027-0.033|,pcorrected=0.009-0.03). Subcortical volume in the left caudate, right amygdala, and bilateral nucleus accumbens was negatively associated with emotion dysregulation (β=|0.026 – 0.037|,pcorrected=<0.001-0.02). Over the two-year follow-up period, higher baseline cortical thickness in the left pars triangularis and rostral middle frontal gyrus predicted greater decreases in externalizing symptoms (F=6.33-6.94,pcorrected=0.014). The results of the current study suggest that unique regions within frontolimbic and striatal networks may be more strongly associated with different dimensions of externalizing psychopathology. The longitudinal findings indicate that brain structure in early childhood may provide insight into structural features that influence behaviour over time.
Optimal geographical distribution of Saudi dates exports
Authors: Ghanem Adel Mohammed , Alrwis Khaled Nahar , Alnashwan Othman S. , Alnafissa Mohamad A. , Ahamada Said Azali , Al-Nashwan Ibrahim bin Othman
Published: March 2024

Purpose

This research aimed to maximize the value of date exports for the Kingdom of Saudi Arabia.

Design/methodology/approach

To achieve its objective, this study relied on secondary data and quantitative economic analysis represented by the Linear programming model.

Findings

This study showed that Saudi Arabia exports dates to the United Arab Emirates, Yemen, Kuwait, Turkey, Somalia, Jordan, Oman, India, Indonesia, Bangladesh Morocco, Lebanon, and others. The geographical concentration coefficient for the quantity and value of date exports was 35.05% and 34.74%, respectively, during the study period. Saudi Arabia exported a quantity of dates amounting to 83.08 thousand tons, representing 40.57% of the average total amount of Saudi dates exports during the study period, to Yemen, Somalia, India, Indonesia, Bangladesh, Egypt, China, Djibouti, Bahrain, and Ethiopia, at prices lower than the average export price of 1200.31 dollars/ton, and therefore the export policy needs to restructure the geographical distribution of date exports. Based on the models of geographical distribution, Saudi date exports value can be increased by 32.76–127.12 million dollars, meaning can be increased by 13.77% – 53.44%. In light of the results of the proposed models, this study recommends the need to restructure the geographical distribution of Saudi date exports so that the value of Saudi date exports can be increased by 127.12 million dollars from the current situation for the period 2017–2021.

Originality/value

The paper’s original contribution lies in its proposal to restructure the geographical distribution of Saudi date exports to increase the value of exports.