bluuebunny/crossref_metadata_2025 huggingface dataset:
The oscillation of mitotic kinase governs cell cycle latches in mammalian cells
Authors: Dragoi Calin-Mihai , Barr Alexis R. , Tyson John J. , Novák Béla
Published: May 2023

The mammalian cell cycle alternates between two phases: S-G2-M with high levels of A- and B-type cyclin-dependent kinases (CycA,B:CDK); and G1 with persistent degradation of CycA,B by Cdh1-activated APC/C (anaphase promoting complex/cyclosome). Because CDKs phosphorylate and inactivate Cdh1, these two phases are mutually exclusive. This ‘toggle switch’ is flipped from G1 to S by cyclin-E (CycE:CDK), which is not degraded by Cdh1:APC/C; and from M to G1 by Cdc20:APC/C, which is not inactivated by CycA,B:CDK. After flipping the switch, cyclin E is degraded and Cdc20:APC/C is inactivated. Combining mathematical modelling with single-cell timelapse imaging, we show that dysregulation of CycB:CDK disrupts strict alternation of the G1-S and M-G1 switches. Inhibition of CycB:CDK results in Cdc20-independent Cdh1 ‘endocycles’, and sustained activity of CycB:CDK drives Cdh1-independent Cdc20 endocycles. Our model provides one mechanistic explanation for how whole genome doubling can arise, a common event in tumorigenesis that can drive tumour evolution.
Spatio-relational inductive biases in spatial cell-type deconvolution
Authors: Viñas Ramon , Scherer Paul , Simidjievski Nikola , Jamnik Mateja , Liò Pietro
Published: May 2023

Spatial transcriptomic technologies profile gene expressionin-situ, facilitating the spatial characterisation of molecular phenomena within tissues, yet often at multi-cellular resolution. Computational approaches have been developed to infer fine-grained cell-type compositions across locations, but they frequently treat neighbouring spots independently of each other. Here we present GNN-C2L, a flexible deconvolution approach that leverages proximal inductive biases to propagate information along adjacent spots. In performance comparison on simulated and semisimulated datasets, GNN-C2L achieves increased deconvolution performance over spatial-agnostic variants. We believe that accounting for spatial inductive biases can yield improved characterisation of cell-type heterogeneity in tissues.
Bayesian Inference of Chemical Mixtures in Risk Assessment Incorporating the Hierarchical Principle
Authors: Kundu Debamita , Kim Sung Duk , Albert Paul S.
Published: May 2023

Analyzing health effects associated with exposure to environmental chemical mixtures is a challenging problem in epidemiology, toxicology, and exposure science. In particular, when there are a large number of chemicals under consideration it is difficult to estimate the interactive effects without incorporating reasonable prior information. Based on substantive considerations, researchers believe that true interactions between chemicals need to incorporate their corresponding main effects. In this paper, we use this prior knowledge through a shrinkage prior that a priori assumes an interaction term can only occur when the corresponding main effects exist. Our initial development is for logistic regression with linear chemical effects. We extend this formulation to include non-linear exposure effects and to account for exposure subject to detection limit. We develop an MCMC algorithm using a shrinkage prior that shrinks the interaction terms closer to zero as the main effects get closer to zero. We examine the performance of our methodology through simulation studies and illustrate an analysis of chemical interactions in a case-control study in cancer.
Calorie Restriction Outperforms Bariatric Surgery in a Murine Model of Obesity and Triple-Negative Breast Cancer
Authors: Coleman Michael F , Camp Kristina K , McFarlane Tori L , Doerstling Steven S , Khatib Subreen A , Rezeli Erika T , Lewis Alfor G , Pfeil Alex J , Smith Laura A , Bowers Laura W , Fouladi Farnaz , Gong Weida , Glenny Elaine M , Parker Joel S , Milne Ginger L , Carroll Ian M , Fodor Anthony A , Seeley Randy J , Hursting Stephen D
Published: May 2023

Obesity promotes triple-negative breast cancer (TNBC), and effective interventions are urgently needed to break the obesity-TNBC link. Epidemiologic studies indicate that bariatric surgery reduces TNBC risk, while evidence is limited or conflicted for weight loss via low-fat diet (LFD) or calorie restriction (CR). Using a murine model of obesity- driven TNBC, we compared the antitumor effects of vertical sleeve gastrectomy (VSG) with LFD, chronic CR, and intermittent CR. Each intervention generated weight and fat loss and suppressed tumor growth relative to obese mice (greatest suppression with CR). VSG and CR regimens exerted both similar and unique effects, as assessed using multi-omic approaches, in reversing obesity-associated transcriptional, epigenetic, secretome, and microbiota changes and restoring antitumor immunity. Thus, in a murine model of TNBC, bariatric surgery and CR each reverse obesity-driven tumor growth via shared and distinct antitumor mechanisms, and CR is superior to VSG in reversing obesity’s procancer effects.
Intensity- and frequency-specific effects of transcranial alternating current stimulation are explained by network dynamics
Authors: Zhao Zhihe , Shirinpour Sina , Tran Harry , Wischnewski Miles , Opitz Alexander
Published: May 2023

Transcranial alternating current stimulation (tACS) can be used to non-invasively entrain neural activity, and thereby cause changes in local neural oscillatory power. Despite an increased use in cognitive and clinical neuroscience, the fundamental mechanisms of tACS are still not fully understood. Here, we develop a computational neuronal network model of two-compartment pyramidal neurons and inhibitory interneurons which mimic the local cortical circuits. We model tACS with electric field strengths that are achievable in human applications. We then simulate intrinsic network activity and measure neural entrainment to investigate how tACS modulates ongoing endogenous oscillations. First, we show that intensity-specific effects of tACS are non-linear. At low intensities (<0.3 mV/mm), tACS desynchronizes neural firing relative to the endogenous oscillations. At higher intensities (>0.3 mV/mm), neurons are entrained to the exogenous electric field. We then further explore the stimulation parameter space and find that entrainment of ongoing cortical oscillations also depends on frequency by following an Arnold tongue. Moreover, neuronal networks can amplify the tACS induced entrainment via excitation-inhibition balance. Our model shows that pyramidal neurons are directly entrained by the exogenous electric field and drive the inhibitory neurons. Our findings can thus provide a mechanistic framework for understanding the intensity- and frequency- specific effects of oscillating electric fields on neuronal networks. This is crucial for rational parameters selection for tACS in cognitive studies and clinical applications.
Changes in Cortical Directional Connectivity during Difficult Listening in Younger and Older Adults
Authors: Soleimani Behrad , Karunathilake I.M. Dushyanthi , Das Proloy , Kuchinsky Stefanie E. , Babadi Behtash , Simon Jonathan Z.
Published: May 2023

One way to investigate the mechanisms that underlie speech comprehension under difficult listening conditions is via cortical connectivity. The innovative Network Localized Granger Causality (NLGC) framework was applied to magnetoencephalography (MEG) data, obtained from older and younger subjects performing a speech listening task in noisy conditions, in delta and theta frequency bands. Directional connectivity between frontal, temporal, and parietal lobes was analyzed. Both aging- and condition-related changes were found, particularly in theta. In younger adults, as background noise increased, there was a transition from predominantly temporal-to-frontal (bottom-up) connections, to predominantly frontal-to-temporal (top-down). In contrast, older adults showed bidirectional information flow between frontal and temporal cortices even for speech in quiet, not changing substantially with increased noise. Additionally, younger listeners did not show changes in the nature of their cortical links for different listening conditions, whereas older listeners exhibited a switch from predominantly facilitative links to predominantly sharpening, when noise increased.

Graphical Abstract

Highlights

Different bands show strong differences in directional functional connectivity patterns

Directional functional connectivity patterns altered by listening task difficulty

Aging dramatically alters directional functional connectivity patterns in during listening

Nature of functional connectivity, additive vs subtractive, depends on age and task
Recruitment status and host-fish-limitation threats to endangered freshwater pearl mussel (Margaritifera laevis) in eastern Hokkaido, northern Japan
Authors: Miura Kazuki , Ishiyama Nobuo , Negishi Junjiro N. , Kawajiri Keita , Izumi Hokuto , Ito Daisetsu , Nakamura Futoshi
Published: May 2023

Recruitment failure is a major threat to freshwater mussel (Order Unionoida) populations worldwide. Assessments of the recruitment status and determining the bottleneck factors of mussel recruitment are crucial for preventing future declines in mussel populations. In this study, we investigated the recruitment status (i.e., size structure and the proportion of juveniles within a population) of the endangered freshwater pearl musselMargaritifera laevisin 22 rivers in eastern Hokkaido, northern Japan. We also quantified the density of the host fishOncorhynchus masou masouand examined the relationship between the proportion of juveniles and host-fish density to assess host-limitation threats toM. laevisrecruitment. Our assessments showed that 13 (59.1%) rivers had no signs of recent recruitment within 10 years, with a low mean proportion of juveniles (mean:0.02 [range:0.00–0.09] fraction), indicating that these populations are threatened by sustained recruitment failure. The proportion of juveniles was positively associated with host-fish density, suggesting that host-fish limitation could be a bottleneck factor forM. laevisrecruitment. These results highlight the urgent need for prompt conservation measures, including the enhancement of host-fish availability, to sustainM. laevispopulations in the study region.
Parsing the effect of co-culture with brain organoids on Diffuse Intrinsic Pontine Glioma (DIPG) using quantitative proteomics
Authors: Prior Victoria G , Maksour Simon , Miellet Sara , Hulme Amy J , Chen Yuyan , Mirzaei Mehdi , Wu Yunqi , Dottori Mirella , O’Neill Geraldine M
Published: May 2023

Diffuse Intrinsic Pontine Gliomas (DIPGs) are deadly brain cancers in children for which there is currently no effective treatment. This can partly be attributed to preclinical models that lack essential elements of thein vivotissue environment, resulting in treatments that appear promising preclinically, but fail to result in effective cures. Recently developed co-culture models combining stem cell-derived brain organoids with brain cancer cells provide tissue dimensionality and a human-relevant tissue-like microenvironment. As these models are technically challenging and time consuming it is imperative to establish whether interaction with the organoid influences DIPG biology and thus warrants their use. To address this question, we cultured DIPG cells with cortical organoids. We created “mosaic” co-cultures enriched for tumour cell-neuronal cell interactions versus “assembloid” co-cultures enriched for tumour cell-tumour cell interactions. Sequential window acquisition of all theoretical mass spectra (SWATH-MS) was used to analyse the proteomes of DIPG fractions isolated by flow-assisted cell sorting. Control proteomes from DIPG spheroids were compared with DIPG cells isolated from mosaic and assembloid co-cultures. This revealed that tumour cell adhesion was reduced, and DNA synthesis and replication were increased, in DIPG cells under either co-culture condition. By contrast, the mosaic co-culture was associated with pathways implicated in dendrite growth. We propose that co-culture with brain organoids is a valuable tool to parse the contribution of the brain microenvironment to DIPG tumour biology.
Laboratory biomarkers associated with mortality in COVID-19 patients in Addis Ababa, Ethiopia
Authors: Sisay Abay , Woldesenbet Zerihun , Yalew Anteneh , Shamenna Aklilu Toma , Worku Asnake , Tesfaye Abraham , Getnet Fentabil , Tesfaye Latera , Hassen Mohammed B. , Geleso Mulugeta , Ngah Veranyuy D , Savieri Perseverence , Mirkuze Alemnesh H. , Sigwadhi Lovemore , Desta Adey Feleke , Nyasulu Peter S
Published: May 2023

Background

Laboratory biomarkers are amongst the best imperative predictors of disease outcomes in hospital-admitted COVID-19 patients. Although data is available in this regard at a global level, there is a paucity of information in Ethiopia. Thus, this study aimed to assess the laboratory biomarkers association with death among COVID-19 patients in Ethiopia.

Methods

A health facility-based longitudinal study was conducted from 2020 to 2022 among RT-PCR-confirmed COVID-19 patients admitted and on treatment follow-up at COVID-19 treatment hospitals in Addis Ababa. A robust Poisson regression model was fitted to assess the association between demographic, clinical, and laboratory factors and death. Significance was determined at p<0.05, and variables with p□<□ 0.15 in bivariate analyses were included in the final multivariable models. Incidence rate ratio (IRR) with a 95% confidence interval (CI) was used to describe associations.

Results

Of the 2357 COVID-19 patients, 248 (10.5%) died. The median age of participants was 59 (IQR= 45-70) years, and the majority (64.9%) of them were male. Lower median RBC was observed among those who died at 4.58 (4.06-5.07) as compared to those who survived at 4.69 (4.23-5.12) whereas high median (IQR) WBC was a predictor of mortality with 11.2 (7.7-15.9). After adjusting for confounders, death was associated with age >74 years having adjusted incidence rate ratio [aIRR (95%CI): 2.46 (1.40-4.34)], and critical clinical situations [aIRR (95% CI): 4.04 (2.18-7.52)].

Conclusion

Our results demonstrate that abnormal liver function tests, abnormal white blood cells, age of the patients, and clinical status of the patients during admission are associated with unfavorable outcomes of COVID-19. Hence, timely monitoring of these laboratory results at the earliest phase of the disease was highly commendable.
Evaluating the Association of Depressive Disorder Symptoms and Moral Injuries in Healthcare Workers during COVID-19 Pandemic
Authors: Behnampour Amirhossein , Ebrahimi Sedigheh , Bazrafshan Amir , Kamyab Amirhossein , Pakdin Majid , Ebrahimi Alireza
Published: May 2023

Background

Moral injury occurs when negative distressing emotions appear and are suppressed. This could lead to several mental health problems such as depression and post-traumatic stress disorder, and result in long-lasting emotional, behavioral, and social problems. Moral injury, a term more commonly used in war contexts, has come into the spotlight during COVID-19 pandemic. We aimed to evaluate the rate of moral injury and its association with psychological injuries during this healthcare crisis.

Methods

We assessed the rates of depression, anxiety, stress, and their association with moral injury among 333 nurses, medical interns, and residents between December 2020 and January 2021. This study was done using validated versions of Depression Anxiety Stress Scales (DASS- and Moral Injury Symptom Scale-Healthcare Professionals (MISS-HP) scores.

Results

Totally 333 healthcare professionals participated in this study, mostly aged between 26 to 30 years old. Nearly half of the participants had a clinically significant moral injury. The average scores of anxiety and stress were significantly higher in women. The participants who were single showed higher rates of depression and moral injury than married ones. Moreover, anxiety, stress, depression, and moral injury were higher in nurses than other healthcare professionals. The scarcity of personal protective equipment at the workplace and giving care to patients with end-stage COVID-19 diagnosis were among the factors associated with a higher risk of developing mental health problems.

Conclusion

The results of this study showed that anxiety, stress, depression, and moral injury were prevalent among healthcare professionals during COVID-19 pandemic. Also, the rates of anxiety, stress, and depression were associated with moral injuries.
WITHDRAWN: Reversing olorofim activity reveals multifactorial drug tolerance in Aspergillus fumigatus
Authors: Valero Clara , Mcmanus Myles , Ram Anna Durai Ashva , Tindale James , Gago Sara , Bromley Michael J
Published: May 2023

Withdrawal Statement

The authors have withdrawn their manuscript owing to finding new evidence contradictory to the results and conclusions presented in this work. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.
Rapid classification vs. refined learning about harm intent: Roles of serotonin and racial bias
Authors: Moutoussis Michael , Barnby Joe , Durand Anais , Croal Megan , Dilley Laura , Rutledge Robb B. , Mason Liam
Published: May 2023

Attributing motives to others is a crucial aspect of mentalizing, can be biased by prejudice, and also is affected by common psychiatric disorders. It is therefore important to understand in depth the mechanisms underpinning it. Toward improving models of mentalizing motives, we hypothesized that people quickly infer whether other’s motives are likely beneficial or detrimental, then refine their judgment (‘Classify-refine’). To test this, we used a modified Dictator game, a game theoretic task, where participants judged the likelihood of intent to harm vs. self-interest in economic decisions. Toward testing the role of serotonin in judgments of intent to harm, we delivered the task in a week-long, placebo vs. Citalopram study.

Computational model comparison provided clear evidence for the superiority of Classify-refine models over traditional ones, strongly supporting the central hypothesis. Further, while Citalopram helped refine attributions about motives through learning, it did not induce more positive initial inferences about others’ motives. Finally, model comparison indicated a minimal role for racial bias within economic decisions for the large majority of our sample. Overall, these results support a proposal that classify-refine social cognition is adaptive, although relevant mechanisms of Serotonergic antidepressant action will need to be studied over longer time spans.
Optimized design andin vivoapplication of optogenetically functionalizedDrosophiladopamine receptors
Authors: Zhou Fangmin , Tichy Alexandra-Madelaine , Imambocus Bibi Nusreen , Rodriguez Jimenez Francisco J. , Martínez Marco González , Jahan Ishrat , Habib Margarita , Wilhelmy Nina , Bräuler Vanessa , Lömker Tatjana , Sauter Kathrin , Helfrich-Förster Charlotte , Pielage Jan , Grunwald Kadow Ilona C. , Janovjak Harald , Soba Peter
Published: May 2023

Neuromodulatory signalingviaG protein-coupled receptor (GPCRs) plays a pivotal role in regulating neural network function and animal behavior. Recent efforts have led to the development of optogenetic tools to induce G protein-mediated signaling, with the promise of acute and cell type-specific manipulation of neuromodulatory signals. However, designing and deploying optogenetically functionalized GPCRs (optoXRs) with accurate specificity and activity to mimic endogenous signalingin vivoremains challenging. Here we optimized the design of optoXRs by considering evolutionary conserved GPCR-G protein interactions and demonstrate the feasibility of this approach using twoDrosophilaDopamine receptors (optoDopRs). We validated these optoDopRs showing that they exhibit high signaling specificity and light sensitivityin vitro.In vivowe detected receptor and cell type-specific effects of dopaminergic signaling in various behaviors including the ability of optoDopRs to rescue loss of the endogenous receptors. This work demonstrates that OptoXRs can enable optical control of neuromodulatory receptor specific signaling in functional and behavioral studies.
GlycoDraw: A Python Implementation for Generating High-Quality Glycan Figures
Authors: Lundstrøm Jon , Urban James , Thomès Luc , Bojar Daniel
Published: May 2023

Glycans are essential to all scales of biology, with their intricate structures being crucial for their biological functions. The structural complexity of glycans is communicated through simplified and unified visual representations according to the Symbol Nomenclature for Glycans (SNFG) guidelines adopted by the community. Here, we introduce GlycoDraw, a Python-native implementation for high-throughput generation of high-quality, SNFG-compliant glycan figures with flexible display options. GlycoDraw is released as part of our glycan analysis ecosystem, glycowork, facilitating integration into existing workflows by enabling fully automated annotation of glycan-related figures and thus assisting the analysis of e.g., differential abundance data or glycomics mass spectra.
Tailoring evidence into action: using a codesign approach for biodiversity information in the Tropical Andes
Authors: Valdez Jose W. , Pereira Henrique M. , Morejón Gustavo Francisco , Acosta-Muñoz Cristina , Bonet Garcia Francisco Javier , Castro Vergara Lucía , Xavier Claros R. , Gill Michael J. , Josse Carmen , Lafuente-Cartagena Indyra , Langstroth Robert , Sheppard Sidney Novoa , Orihuela Gabriela , Prieto-Albuja Francisco J. , Quillahuaman Natividad , Terán Marcos F. , Zambrana-Torrelio Carlos M. , Navarro Laetitia M. , Fernandez Miguel
Published: May 2023

Biodiversity conservation is a complex and transdisciplinary problem that requires engagement and cooperation among scientific, societal, economic, and political institutions. However, historical approaches have often failed to bring together and address the needs of relevant stakeholders in decision-making processes. The Tropical Andes, a biodiversity hotspot where conservation efforts often conflict with socioeconomic issues and policies that prioritize economic development, provides an ideal model to develop and implement more effective approaches. In this study, we present a codesign approach that mainstreams and improves the flow of biodiversity information in the Tropical Andes, while creating tailored outputs that meet the needs of economic and societal stakeholders. We employed a consultative process that brought together biodiversity information users and producers at the local, national, and regional levels through a combination of surveys and workshops. This approach identified priority needs and limitations of the flow of biodiversity information in the region, which led to the co-design of user-relevant biodiversity indicators. By leveraging the existing capacities of biodiversity information users and producers, we were able to co-design multiple biodiversity indicators and prioritize two for full implementation ensuring that the data was findable, accessible, interoperable, and reusable based on the FAIR principles. This approach helped address limitations that were identified in the stakeholder engagement process, including gaps in data availability and the need for more accessible biodiversity information. Additionally, capacity-building workshops were incorporated for all stakeholders involved, which aimed to not only improve the current flow of biodiversity information in the region but also facilitate its future sustainability. Our approach can serve as a valuable blueprint for mainstreaming biodiversity information and making it more inclusive in the future, especially considering the diverse worldviews, values, and knowledge systems between science, policy, and practice.
Protein-Protein interaction and quantitative phosphoproteomic studies reveal potential mitochondrial substrates of protein phosphatase 2A-B’ζ holoenzyme
Authors: Elshobaky Ahmed , Lillo Cathrine , Hodén Kristian Persson , Kataya Amr R.A.
Published: May 2023

Protein phosphatase 2A (PP2A) is a heterotrimeric conserved serine/threonine phosphatase complex including a catalytic, scaffolding, and regulatory subunit. Three A Subunits, 17 B subunits, and five C subunits are encoded by theArabidopsisgenome, allowing 255 possible PP2A holoenzyme combinations. The regulatory subunits are crucial for substrate specificity and PP2A complex localization and are classified into B, B’, and B’’ non-related families in land plants. InArabidopsis, the close homologs B’η, B’θ, B’γ, and B’ζ are further classified into a subfamily of B’ called B’η. Previous studies suggest a role of mitochondrial targeted PP2A subunit (B’ζ) in energy metabolism and plant innate immunity. Potentially, the PP2A-B’ζ holoenzyme is involved in the regulation of the mitochondrial succinate/fumarate translocator or affects enzymes involved in energy metabolism. To investigate this hypothesis, the interaction between PP2A-B’ζ and enzymes involved in the mitochondrial energy flow was investigated using bimolecular fluorescence complementation in tobacco and onion cells. Interaction of B’ζ subunit was confirmed with the Krebs cycle proteins Succinate/fumarate translocator (mSFC1), Malate dehydrogenase (mMDH2), and Aconitase (ACO3). Additional putative interacting candidates were deduced from comparing the enriched phosphoproteomes of wild type and B’ζ mutants: the mitochondrial regulatorArabidopsispentatricopeptide repeat 6 (PPR6) and the two metabolic enzymes Phosphoenolpyruvate carboxylase (PPC3) and Phosphoenolpyruvate carboxykinase (PCK1). Overall, this study identifies potential PP2A substrates and highlights the role of PP2A in regulating energy metabolism in mitochondria.
Saturating the eQTL map inDrosophila melanogaster: genome-wide patterns of cis and trans regulation of transcriptional variation in outbred populations
Authors: Pallares Luisa F. , Melo Diogo , Wolf Scott , Cofer Evan M. , Abhyankar Varada , Peng Julie , Ayroles Julien F.
Published: May 2023

Decades of genome-wide mapping have shown that most genetic polymorphisms associated with complex traits are found in non-coding regions of the genome. Characterizing the effect of such genetic variation presents a formidable challenge, and eQTL mapping has been a key approach to understand the non-coding genome. However, comprehensive eQTL maps are available only for a few species like yeast and humans. With the aim of understanding the genetic landscape that regulates transcriptional variation inDrosophila melanogaster, we developed an outbred mapping panel in this species, theDrosophilaOutbred Synthetic Panel (Dros-OSP). Using this community resource, we collected transcriptomic and genomic data for 1800 individual flies and were able to mapcisandtranseQTLs for 98% of the genes expressed inD. melanogaster, increasing by thousands the number of genes for which regulatory loci are known in this species. We described, for the first time in the context of an outbred population, the properties of local and distal regulation of gene expression in terms of genetic diversity, heritability, connectivity, and pleiotropy. We uncovered that, contrary to long-standing assumptions, a significant part of gene co-expression networks is organized in a non-modular fashion. These results bring the fruit fly to the level of understanding that was only available for a few other organisms, and offer a new mapping resource that will expand the possibilities currently available to theDrosophilacommunity. This data is available atDrosophilaeqtl.org.
Noradrenergiclocus coeruleusactivity functionally partitions NREM sleep to gatekeep the NREM-REM sleep cycle
Authors: Osorio-Forero Alejandro , Foustoukos Georgios , Cardis Romain , Cherrad Najma , Devenoges Christiane , Fernandez Laura M.J. , Lüthi Anita
Published: May 2023

The noradrenergiclocus coeruleus(LC) is vital for brain states underlying wakefulness, whereas its roles for sleep remain uncertain. Combining mouse sleep-wake monitoring, behavioral manipulations, LC fiber photometry and closed-loop optogenetics, we found that LC neuronal activity partitioned non-rapid-eye-movement sleep (NREMS) into alternating brain and autonomic states that rule the NREMS-REMS cycle. High LC activity levels generated an autonomic-subcortical arousal state that facilitated cortical microarousals, while low levels were obligatory for REMS entries. Timed optogenetic LC inhibition revealed that this functional alternation set the duration of the NREMS-REMS cycle by ruling REMS entries during undisturbed sleep and when pressure for REMS was high. A stimulus-enriched, stress-promoting wakefulness increased high LC activity levels at the expense of low ones in subsequent NREMS, fragmenting NREMS through microarousals and delaying REMS onset. We conclude that LC activity fluctuations gatekeep the NREM-REMS cycle over recurrent infraslow intervals, but they also convey vulnerability to adverse wake experiences.
Binding of cortical functional modules by synchronous high frequency oscillations
Authors: Garrett Jacob C. , Verzhbinsky Ilya A. , Kaestner Erik , Carlson Chad , Doyle Werner K. , Devinsky Orrin , Thesen Thomas , Halgren Eric
Published: May 2023

Whether high-frequency phase-locked oscillations facilitate integration (‘binding’) of information across widespread cortical areas is controversial. Here we show with intracranial EEG that cortico-cortical co-ripples (∼100ms long ∼90Hz oscillations) increase during reading and semantic decisions, at the times and co-locations when and where binding should occur. Fusiform wordform areas co-ripple with virtually all language areas, maximally from 200-400ms post-word-onset. Semantically-specified target words evoke strong co-rippling between wordform, semantic, executive and response areas from 400-800ms, with increased co-rippling between semantic, executive and response areas prior to correct responses. Co-ripples were phase-locked at zero-lag over long distances (>12cm), especially when many areas were co-rippling. General co-activation, indexed by non-oscillatory high gamma, was mainly confined to early latencies in fusiform and earlier visual areas, preceding co-ripples. These findings suggest that widespread synchronous co-ripples may assist the integration of multiple cortical areas for sustained periods during cognition.

One Sentence Summary

Widespread visual, wordform, semantic, executive and response areas phase-lock at 90Hz during integrative semantic processing.
Components of Isolated Skeletal Muscle Differentiated Through Antibody Validation
Authors: Stephens Dominique C. , Mungai Margaret , Crabtree Amber , Beasley Heather K. , Garza-Lopez Edgar , Vang Larry , Neikirk Kit , Vue Zer , Vue Neng , Marshall Andrea G. , Turner Kyrin , Shao Jian-qiang , Sarker Bishnu , Murray Sandra , Gaddy Jennifer A. , Davis Jamaine , Damo Steven M. , Hinton Antentor O.
Published: May 2023

Isolation of skeletal muscles allows for the exploration of many complex diseases. Fibroblasts and myoblast play important roles in skeletal muscle morphology and function. However, skeletal muscles are complex and made up of many cellular populations and validation of these populations is highly important. Therefore, in this article, we discuss a comprehensive method to isolate mice skeletal muscle, create satellite cells for tissue culture, and use immunofluorescence to validate our approach.

Graphical Abstract
Creating Optimal Conditions for OPA1 Isoforms by Western Blot in Skeletal Muscle Cells and Tissue
Authors: Stephens Dominique C. , Mungai Margaret , Crabtree Amber , Beasley Heather K. , Garza-Lopez Edgar , Neikirk Kit , Bacevac Serif , Vang Larry , Vue Zer , Vue Neng , Marshall Andrea G. , Turner Kyrin , Shao Jianqiang , Murray Sandra , Gaddy Jennifer A. , Wanjalla Celestine , Davis Jamaine , Damo Steven , Hinton Antentor O.
Published: May 2023

OPA1 is a dynamin-related GTPase that modulates various mitochondrial functions and is involved in mitochondrial morphology. There are eight different isoforms of OPA1 in humans and five different isoforms in mice that are expressed as short or long-form isoforms. These isoforms contribute to OPA1’s ability to control mitochondrial functions. However, isolating OPA1 all long and short isoforms through western blot has been a difficult task. To address this issue, we outline an optimized western blot protocol to isolate 5 different isoforms of OPA1 on the basis of different antibodies. This protocol can be used to study changes in mitochondrial structure and function.

Tweetable Abstract

Western blot protocol optimization to visualize OPA1 isoforms.

Highlights

Protocol for isolating OPA1 isoforms in primary skeletal muscle myoblast and myotubes

Steps for running isolated skeletal muscle cells from muscle tissue on a gel

How to collect samples in preparation for western blotting

Detection of OPA1 isoforms

Graphical Abstract

Method Summary

Samples for western blot analysis are isolated from lysed cells, loaded onto a gel, and ran using optimized conditions to better isolate OPA1 isoforms. Samples are transferred to a membrane for incubation and protein detection using OPA1 antibodies.
Ablation ofSam50is associated with fragmentation and alterations in metabolism in murine and human myotubes
Authors: Shao Bryanna , Killion Mason , Oliver Ashton , Vang Chia , Zeleke Faben , Neikirk Kit , Vue Zer , Garza-Lopez Edgar , Shao Jian-qiang , Mungai Margaret , Lam Jacob , Williams Qiana , Altamura Christopher T. , Whiteside Aaron , Kabugi Kinuthia , McKenzie Jessica , Koh Alice , Scudese Estevão , Vang Larry , Marshall Andrea G. , Crabtree Amber , Tanghal Janelle I. , Stephens Dominique , Koh Ho-Jin , Jenkins Brenita C. , Murray Sandra A. , Cooper Anthonya T. , Williams Clintoria , Damo Steven M. , McReynolds Melanie R. , Gaddy Jennifer A. , Wanjalla Celestine N. , Beasley Heather K. , Hinton Antentor
Published: May 2023

The Sorting and Assembly Machinery (SAM) Complex is responsible for assembling β-barrel proteins in the mitochondrial membrane. Comprising three subunits, Sam35, Sam37, and Sam50, the SAM complex connects the inner and outer mitochondrial membranes by interacting with the mitochondrial contact site and cristae organizing system (MICOS) complex. Sam50, in particular, stabilizes the mitochondrial intermembrane space bridging (MIB) complex, which is crucial for protein transport, respiratory chain complex assembly, and regulation of cristae integrity. While the role of Sam50 in mitochondrial structure and metabolism in skeletal muscle remains unclear, this study aims to investigate its impact. Serial block-face-scanning electron microscopy (SBF-SEM) and computer-assisted 3D renderings were employed to compare mitochondrial structure and networking inSam50-deficient myotubes from mice and humans with wild-type (WT) myotubes. Furthermore, autophagosome 3D structure was assessed in human myotubes. Mitochondrial metabolic phenotypes were assessed using Gas Chromatography-Mass Spectrometry-based metabolomics to explore differential changes in WT andSam50-deficient myotubes. The results revealed increased mitochondrial fragmentation and autophagosome formation inSam50-deficient myotubes compared to controls. Metabolomic analysis indicated elevated metabolism of propanoate and several amino acids, including ß-Alanine, phenylalanine, and tyrosine, along with increased amino acid and fatty acid metabolism inSam50-deficient myotubes. Furthermore, impairment of oxidative capacity was observed uponSam50ablation in both murine and human myotubes, as measured with the XF24 Seahorse Analyzer. Collectively, these findings support the critical role of Sam50 in establishing and maintaining mitochondrial integrity, cristae structure, and mitochondrial metabolism. By elucidating the impact ofSam50-deficiency, this study enhances our understanding of mitochondrial function in skeletal muscle.
A method for extracting an approximated connectome from libraries of single anatomically co-registered cell reconstructions
Authors: Manjunatha K.K.H. , Bruzzone M. , Nicoletti G. , Suweis S. , dal Maschio M.
Published: May 2023

Mapping functional brain information onto a circuit wiring diagram supporting possible brain circuit working mechanisms remains challenging. When available, the analysis of electron microscopy based high-resolution connectomes, leveraging synapse annotation, allows for unraveling portions of the nervous system’s wiring diagram or specific circuit motifs. However, obtaining a complete brain reconstruction with annotation of the synapses and extracting the general organization principles of the neuronal networks across the brain remains a demanding effort. To extend the available methods, we present here an approach to obtain an approximate brain connectome starting from libraries of single cell reconstructions co-registered in a reference anatomical space. Our approach relies on both the Strahler numbering of the nodes characterizing the cell morphological reconstructions and a proximity range criterion, to infer the general connectivity structures between the different cells, in the absence of synapse annotation. First, we confronted this approach on a ground truth derived from a fully annotated EM dataset ofDrosophila melanogaster. The connectivity modules identified with our approach show generally a similar organization compared to those resulting from the EM annotated dataset, in terms of Normalized Mutual Information and Adjusted Rand Index, always with high significance with respect to random null models. Building on this, we then applied this approach to extract an approximate connectome of the zebrafish larvae brain starting from a light microscopy-based dataset of about 3000 co-registered neuronal skeletonizations. Modularity analysis of the retrieved directed and undirected connectomes revealed a hierarchical organization of the resulting graph. The identified architecture encompasses a series of high-order brain-wide neuronal modules composed of a number of lower-order submodules. Importantly, we show that this approximated representation is sufficient for capturing precise and topographically organized connection patterns as well as known functional circuit architectures. In conclusion, we present a scalable, from-circuit-to-brain range, approach to reveal neuronal architectures supporting brain mechanisms, potentially suitable for hypothesis drawing and as for guiding the exploration of and integration with EM connectomes.
SNAP25 disease mutations change the energy landscape for synaptic exocytosis due to aberrant SNARE interactions
Authors: Kádková Anna , Murach Jacqueline , Østergaard Maiken , Malsam Andrea , Malsam Jörg , Lolicato Fabio , Nickel Walter , Söllner Thomas H. , Sørensen Jakob B.
Published: May 2023

SNAP25 is one of three neuronal SNAREs driving synaptic vesicle exocytosis. We studied three mutations in SNAP25 that cause epileptic encephalopathy: V48F, and D166Y in the Synaptotagmin-1 (Syt1) binding interface, and I67N, which destabilizes the SNARE-complex. All three mutations reduced Syt1-dependent vesicle docking to SNARE-carrying liposomes and Ca2+-stimulated membrane fusionin vitroand in neurons. The V48F and D166Y mutants (with potency D166Y > V48F) led to reduced Readily Releasable Pool (RRP) size, due to increased spontaneous (mEPSC) release and decreased priming rates. These mutations lowered the energy barrier for fusion and increased the release probability, which are gain-of-function features not found inSyt1knockout (KO) neurons; normalized mEPSC release rates were higher (potency D166Y>V48F) than in theSyt1KO. These mutations (potency D166Y > V48F) increased spontaneous association to partner SNAREs, resulting in unregulated membrane fusion. In contrast, the I67N mutant decreased mEPSC frequency and evoked EPSC amplitudes due to an increase in the apparent height of the energy barrier for fusion, whereas the RRP size was unaffected. This could be partly compensated by positive charges lowering the energy barrier. Overall, pathogenic mutations in SNAP25 cause complex changes in the energy landscape for priming and fusion.
Age- and caste-independent piRNAs in the germline and miRNA profiles linked to caste and fecundity in the antTemnothorax rugatulus
Authors: Seistrup Ann-Sophie , Choppin Marina , Govind Shamitha , Feldmeyer Barbara , Kever Marion , Karaulanov Emil , Séguret Alice , Karunanithi Sivarajan , Almeida Miguel V. , Ketting René F. , Foitzik Susanne
Published: May 2023

Social insects are models for phenotypic plasticity: the generation of different phenotypes from the same genotype. Ant queens and workers differ not only in their morphology and behaviour, but also in their fecundity and lifespan, which is often several times higher in queens. However, the gene regulatory mechanisms underlying these differences are not yet well understood. Since ant queens can live and reproduce for more than two decades, they need to protect their germline from the activity of transposable elements (TEs). This protection may be redundant in short-lived, often sterile workers. We have analysed the expression of two protective classes of smallRNAs, microRNAs (miRNAs) and Piwi-interacting RNAs (piRNAs), in different tissues, castes, and age classes of the ant speciesTemnothorax rugatulus. We show that piRNAs are particularly active in the ovaries of queens. TEs are clear targets of the piRNAs in this ant species, and piRNA-specific sequence signatures in the ovaries of all queens regardless of age indicate that young and old queens have similarly active piRNA pathways. Interestingly, the reduced ovaries of the workers also showed the same level of piRNA activity. This was not only the case in young, fertile workers from queenless nests, but also in the presumably older foragers, which have almost completely regressed ovaries. These findings suggest that the germline in these ants is invariably protected by piRNA activity, irrespective of ovarian development. The brain and thorax of queens also contained piRNAs, but at lower levels, and the piRNA-specific ping-pong signatures were strongly reduced in these tissues. We also annotated and analysed miRNAs in different tissues. We confidently detected the expression of 304 miRNAs. Of these, 10 were enriched in the brain and three to the thorax, whereas 83 were specific to the ovaries. 105 miRNAs were found to be expressed in all three tissues. We also identified miRNAs whose expression potentially is related to ant caste, fecundity, and age, suggesting that caste-specific gene activity may be regulated in part by miRNAs. In contrast, our studies of piRNA activity indicate similar profiles across caste, fecundity and age groups, but strong tissue specificity with the highest piRNA mediated TE protection in the germline.
Time-gated Fluorescence Lifetime Imaging in the Near Infrared Regime; A Comprehensive Study Toward In Vivo Imaging
Authors: Harel Meital , Arbiv Uri , Ankri Rinat
Published: May 2023

Fluorescence lifetime imaging has an enormous impact on our understanding of biological systems, both in vitro and in vivo. It is a powerful tool for the non-invasive in vitro and in vivo biomolecular and cellular investigations. In particular, it has the potential to target and multiplex different species with high sensitivity and specificity, providing a fast and noninvasive readout at low cost. In this work, we present a time-saving Monte Carlo (MC) simulation of fluorescent photons scattering within a turbid medium, followed by phasor analyzes which enabled the simple multiplexing of different targets in one frame. We then demonstrate a simple and fast method for wide-field FLI in the near-infrared (NIR) region, where tissue scattering and autofluorescence are significantly lower, to enable imaging of deep tissue, using the state-of-the-art timed single-photon avalanche diode array camera (SPAD), SPAD512S. In particular, we show how phasor scattering increases with depth. However, using appropriate background correction, a simple “cut-off” method, and averaging, we can multiplex two targets in one image to a depth of 1 cm in tissue. Our results show that it is possible to perform in vivo FLI under challenging conditions, using standard NIR fluorophores with short lifetimes.
Mechanistic explanation of neuroplasticity using equivalent circuits
Authors: Nilsson Martin N P
Published: May 2023

This paper presents a comprehensive mechanistic model of a neuron with plasticity that explains how information input as time-varying signals is processed and stored. Additionally, the model addresses two long-standing, specific biological challenges: Integrating Hebbian and homeostatic plasticity, and identifying a concise synaptic learning rule. A biologically accurate electric-circuit equivalent is derived through a one-to-one mapping from the known properties of ion channels. The often-overlooked dynamics of the synaptic cleft is essential in this process. Analysis of the model reveals a simple and succinct learning rule, indicating that the neuron functions as an internal-feedback adaptive filter, a common concept in signal processing. Simulations confirm the model's functionality, stability, and convergence, demonstrating that even a single neuron without external feedback can act as a potent signal processor. The model replicates several key characteristics typical of biological neurons, which are seldom captured in other neuron models. It can encode time-varying functions, learn without risking instability, and bootstrap from a state where all synaptic weights are zero. This paper explores the function of neurons with a focus on biological accuracy, not computational efficiency. Unlike neuromorphic models, it does not aim to design devices. The electronic circuit analogy aids understanding by leveraging decades of electronics expertise but is not intended for physical implementation. This interdisciplinary work spans a broad range of subjects within the realm of neurobiophysics, including neurobiology, electronics, and signal processing.
Genomic, effector protein and culture-based analysis ofCyclaneusma minusin New Zealand provides evidence for multiple morphotypes
Authors: Tarallo M. , Dobbie K. , Leite L. Nunes , Waters T. , Gillard K. , Sen D. , McDougal R.L. , Mesarich C.H. , Bradshaw R.E.
Published: May 2023

Cyclaneusma needle cast, caused byCyclaneusma minus, affectsPinusspecies around the world. Previous studies suggested the presence of two distinct morphotypes in New Zealand, ‘verum’ and ‘simile’. Traditional mycological analyses revealed a third morphotype with clear differences in colony morphology and cardinal growth rates at varying temperatures. Genome sequencing of eightC. minusisolates provided further evidence of the existence of a third morphotype, named ‘novus’ in this study. To further analyse these morphotypes, we predicted candidate effector proteins for all eight isolates, and also characterized a cell-death eliciting effector family, Ecp32, which is present in other pine phytopathogens. In concordance with their distinct classification into three different morphotypes, the number of Ecp32 family members differed, with patterns of pseudogenization and some family members being found exclusively in some morphotypes. We also showed that proteins belonging to the Ecp32 family trigger cell death responses in non-hostNicotianaspecies, and, as previously demonstrated for other plant pathogens, theC. minusproteins belonging to the Ecp32 family adopt a β-trefoil fold. Understanding the geographical range and variations in virulence and pathogenicity of these morphotypes will provide a better understanding of pine needle diseases as well as enable the development of more durable methods to control this disease.