bluuebunny/crossref_metadata_2025 huggingface dataset:
Two modes of mid-frontal theta suggest a role in conflict and error processing
Authors: Muralidharan Vignesh , Aron Adam R , Cohen Michael X , Schmidt Robert
Published: March 2022

Mid-frontal theta increases during scenarios when conflicts are successfully resolved. Often considered as a generic signal of cognitive control, its temporal nature has hardly been investigated. Using advanced spatiotemporal techniques, we uncover that mid-frontal theta occurs as a transient oscillation or “event” at single trials with their timing reflecting computationally distinct modes. Single-trial analyses of electrophysiological data from participants performing the Flanker (N = 28) and Simon task (N = 18) were used to probe the relationship between theta and metrics of response conflict. We specifically investigated “partial errors”, in which a small burst of muscle activity in the incorrect response effector occurred, quickly followed by a correction. We found that transient theta events in single trials could be categorized into two distinct theta modes based on their relative timing to different task events. Theta events from the first mode occurred briefly after the task stimulus and might reflect conflict-related processing of the stimulus. In contrast, theta events from the second mode were more likely to occur around the time partial errors were committed, suggesting they were elicited by a potential upcoming error. Importantly, in trials in which a full error was committed, this “error-related theta” occurred too late with respect to the onset of the erroneous muscle response, supporting a role of theta also in error correction. We conclude that different modes of transient mid-frontal theta can be adopted in single trials not only to process stimulus conflict, but also to correct erroneous responses.
Single Nucleus RNA Sequencing of Pre-Malignant Liver Reveals Disease-Associated Hepatocyte State with HCC Prognostic Potential
Authors: Carlessi Rodrigo , Denisenko Elena , Boslem Ebru , Koehn-Gaone Julia , Main Nathan , Abu Bakar N. Dianah B. , Shirolkar Gayatri D. , Jones Matthew , Poppe Daniel , Dwyer Benjamin J. , Jackaman Connie , Tjiam M. Christian , Lister Ryan , Karin Michael , Fallowfield Jonathan A. , Kendall Timothy J. , Forbes Stuart J. , Olynyk John K. , Yeoh George , Forrest Alistair R. R. , Ramm Grant A. , Febbraio Mark A. , Tirnitz-Parker Janina E. E.
Published: March 2022

Current approaches to stage chronic liver diseases have limited utility to directly predict liver cancer risk. Here, we employed single nucleus RNA sequencing (snRNA-seq) to characterize the cellular microenvironment of healthy and chronically injured pre-malignant livers using two distinct mouse models. Analysis of 40,748 hepatic nuclei unraveled a previously uncharacterized disease-associated hepatocyte transcriptional state (daHep). These cells were absent in healthy livers, but were increasingly prevalent as chronic liver disease progressed towards hepatocarcinogenesis. Gene expression deconvolution of 1,439 human liver transcriptomes from publicly available datasets revealed that daHep frequencies highly correlate with current histopathological liver disease staging systems. Importantly, we show that high daHep levels precede carcinogenesis in mice and humans and predict a higher risk of hepatocellular carcinoma (HCC) development. This novel transcriptional signature with diagnostic and, more importantly, prognostic significance has the potential to change the way chronic liver disease patients are staged, surveilled and risk-stratified.
The role of optogenetic stimulations of parvalbumin-positive interneurons in the prefrontal cortex and the ventral hippocampus on an acute MK801 model of schizophrenia-like cognitive inflexibility
Authors: Patrono Enrico , Hrůzova Karolina , Svoboda Jan , Stuchlík Aleš
Published: March 2022

Background and Hypothesis

Schizophrenia research arose in the last decades, focusing more on its neural basis. Executive functions such as decision making and cognitive flexibility are the main cognitive areas that are impaired and are considered schizophrenia endophenotypes. Recently, cognitive impairment has been connected with the ablation of glutamatergic NMDARs resulting in increased cortical activity. Selective NMDARs antagonists such as dizocilpine have been used to model cognitive inflexibility in schizophrenia. Moreover, a decreased GABAergic inhibitory activity has been shown elsewhere along with the enhanced cortical activity. This NMDARs/GABA unbalanced ratio may reduce the entrainment of prefrontal gamma and hippocampal theta rhythm, resulting in a prefrontal-hippocampal gamma/theta band desynchronization.

Study Design

The study addressed the role of acute administrations of dizocilpine to model schizophrenia-like cognitive inflexibility in rats. We used a new version of the attentional set-shifting task, where rats learned switching/reversing the relevant rule. Moreover, we used the new ASST after dizocilpine systemic injections to test cognitive flexibility. Finally, we used in vivo optogenetic stimulations at specific light pulses of parvalbumin-positive interneurons in the prefrontal cortex and ventral hippocampus.

Results

The first experiments showed that acute dizocilpine in rats reproduced schizophrenia-like cognitive inflexibility. The second set of experiments demonstrated that appropriate optogenetic light pulses frequencies could rescue the cognitive flexibility previously altered by acute dizocilpine.

Conclusions

These findings advance our knowledge on the pivotal role of parvalbumin interneurons in schizophrenia-like cognitive impairment and may serve as a standpoint for further research of this severe psychiatric disorder.
Telomerase-independent maintenance of telomere length in a vertebrate
Authors: Yu Qinghao , Gates Phillip B. , Rogers Samuel , Mikicic Ivan , Elewa Ahmed , Salomon Florian , Lachnit Martina , Caldarelli Antonio , Flores-Rodriguez Neftali , Cesare Anthony J , Simon Andras , Yun Maximina H.
Published: March 2022

Telomere shortening places a key limitation on cell proliferation1. In all vertebrates explored to date, this limitation is overcome by telomerase-dependent telomere extension. Failure to maintain telomere length results in premature ageing and functional impairments in highly replicative cell populations as telomeres erode2. Alternative lengthening of telomeres (ALT), a telomerase-independent mechanism, compensates for telomere loss in a subset of human cancer cell lines 2. Here, we demonstrate that the highly regenerative newt Pleurodeles waltl lacks telomerase activity, contains telomeres distinct from all known vertebrates in both sequence and structure, and deploys ALT for physiological telomere maintenance. This constitutes the first report of telomerase-independent resolution of the end-replication problem at the whole-organism level within Chordata.

One-Sentence Summary

P. waltl telomere biology is distinct amongst vertebrates and uses ALT at the whole-organism level.
Adversary of DNA integrity: a long non-coding RNA stimulates driver oncogenic chromosomal rearrangement in human thyroid cells
Authors: Demin Denis Eriksonovich , Murashko Matvey Mikhailovich , Uvarova Aksinya Nicolaevna , Stasevich Ekaterina Mikhailovna , Shyrokova Elena Yurievna , Gorlachev Gennady Efimovich , Korneev Kirill Viktorovich , Ustiugova Alina Sergeevna , Tkachenko Elena Andreevna , Kostenko Valentina Vitalevna , Tatosyan Karina Aleksandrovna , Sheetikov Saveliy Andreevich , Spirin Pavel Vladimirovich , Kuprash Dmitriy Vladimirovich , Schwartz Anton Markovich
Published: March 2022

The flurry of publications devoted to the functions of long non-coding RNAs (lncRNAs) published in the last decade leaves no doubt about the exceptional importance of lncRNAs in various areas including tumor biology. Contribution of lncRNAs to the early stages of oncogenesis remains poorly understood. In this study we explored a new role for lncRNAs: stimulation of driver oncogenic mutations that result from specific chromosomal rearrangements. We demonstrated that lncRNA CASTL1 (ENSG00000269945) stimulates the formation of the CCDC6-RET inversion (RET/PTC1) in human thyroid cells subjected to radiation or chemical DNA damage. Facilitation of chromosomal rearrangement requires lncRNA to contain regions complementary to the introns of both CCDC6 and RET genes as deletion of these regions deprives CASTL1 of the ability to stimulate the gene fusion. We found that CASTL1 expression is elevated in tumors with CCDC6-RET fusion which is the most frequent rearrangement in papillary thyroid carcinoma. Our results open a new venue for the studies of early oncogenesis in various tumor types, especially those associated with physical or chemical DNA damage.

Graphical abstract
Mark loss can strongly bias demographic rates in multi-state models: a case study with simulated and empirical datasets
Authors: Touzalin Frédéric , Petit Eric J. , Cam Emmanuelle , Stagier Claire , Teeling Emma C. , Puechmaille Sébastien J.
Published: March 2022

The development of methods for individual identification in wild species and the refinement of Capture-Mark-Recapture (CMR) models over the past few decades have greatly improved the assessment of population demographic rates to address ecological and conservation questions. In particular, multi-state models, which offer flexibility in analysing complex study systems, have gained popularity within the ecological community. In this study, we focus on the issue of mark loss and the associated recycling of remarked individuals, which requires further exploration given the increasing use of these models.

To fill this knowledge gap, we employed a wide range of simulation scenarios that reflect commonly encountered real case studies, drawing inspiration from the survival rates of 700 vertebrate species. Using a multi-state, Arnason-Schwartz (AS) modelling framework, we estimated the effects of mark loss and recycled individuals on parameter estimates. We assessed parameter bias by simulating a metapopulation system with varying capture and survival rates. Additionally, we demonstrated how mark loss can be easily estimated and accounted for using a 10-year empirical CMR dataset of bats. The bats were individually identified using Passive Integrated Transponder (PIT) tag technology as potentially lost marks and multi-locus genotypes as ’permanent marks’.

Our simulation results revealed that the occurrence of bias and the affected parameters were highly dependent on the study system, making it difficult to establish general rules to predict biasa priori. The model structure and the interdependency among parameters pose challenges in predicting the impact of bias on estimates.

Our findings underscore the importance of assessing the effect of mark loss when using AS models. Ignoring such violations of model assumptions can have significant implications for ecological inferences and conservation policies. In general, the use of permanent marks, such as genotypes, should always be preferred when modelling population dynamics. If that is not feasible, an alternative is to combine two independent types of temporary marks, such as PIT tags and bands.

Analysis of our empirical dataset onMyotis myotisbats revealed that tag loss is higher in juveniles than in adults during the first year after tagging. The use of surgical glue to close the injection hole reduces tag loss rate from 28% to 19% in juveniles, while it has no effect on the tag loss rate in adults (∼10%). The main bias observed in our metapopulation system appears in the survival rate, with up to a 20% underestimation if tag loss is not accounted for.
Peripheral blood T-cell deficiency and hyperinflammatory monocyte responses associate with MAC lung disease
Authors: Lindestam Arlehamn Cecilia S. , Benson Basilin , Kuan Rebecca , Dill-McFarland Kimberley A. , Peterson Glenna J. , Paul Sinu , Nguyen Felicia K. , Gilman Robert H. , Saito Mayuko , Taplitz Randy , Arentz Matthew , Goss Christopher H. , Aitken Moira L. , Horne David J. , Shah Javeed A. , Sette Alessandro , Hawn Thomas R.
Published: March 2022

Rationale

Although nontuberculous mycobacterial (NTM) disease is a growing problem, available treatments are suboptimal and diagnostic tools are inadequate. Immunological mechanisms of susceptibility to NTM disease are poorly understood.

Objective

To understand NTM pathogenesis, we evaluated innate and antigen-specific adaptive immune responses to Mycobacterium avium complex (MAC) in individuals with MAC lung disease (MACDZ).

Methods

We synthesized 15mer MAC-, NTM-, or MAC/Mtb-specific peptides and stimulated peripheral blood mononuclear cells (PBMC) with pools of these peptides. We measured T-cell responses by cytokine production, expression of surface markers, and analysis of global gene expression in 27 MACDZ individuals and 32 healthy controls. We also analyzed global gene expression in Mav-infected and uninfected peripheral blood monocytes from 17 MACDZ and 17 healthy controls.

Measurements and Main Results

We were unable to detect T-cell responses against the peptide libraries or Mav lysate that has increased reactivity in MACDZ subjects compared to controls. T-cell responses to non-mycobacteria derived antigens were preserved.

MACDZ individuals had a lower frequency of Th1 and Th1* T-cell populations. By contrast, global gene expression analysis demonstrated upregulation of proinflammatory pathways in uninfected and Mav-infected monocytes derived from MACDZ subjects compared to controls.

Conclusions

Peripheral blood T-cell responses to Mycobacterial antigens and the frequency of Th1 and Th1* cell populations are diminished in individuals with MAC disease. In contrast, MACDZ subjects had hyperinflammatory monocyte responses. Together, these data suggest a novel immunologic defect which underlies MAC pathogenesis and includes concurrent innate and adaptive dysregulation.
A Conformation-specific Nanobody Targeting the NMN-activated State of SARM1
Authors: Hou Yun Nan , Cai Yang , Li Wan Hua , He Wei Ming , Zhao Zhi Ying , Zhu Wen Jie , Wang Qiang , Liu Jun , Lee Hon Cheung , Goran Stjepanovic , Zhang Hongmin , Zhao Yong Juan
Published: March 2022

Upon axonal injury, Sterile alpha (SAM) and Toll/interleukin-1 receptor (TIR) motif containing 1 (SARM1) is activated by nicotinamide mononucleotide (NMN) to deplete NAD and consequently promote the process of axon degeneration (AxD). Currently, only the inactive form of SARM1 in its auto-inhibitory conformation has been resolved. The flexibility of the enzymatically active form of SARM1 has so far precluded its structural determination. To solve the problem, we generated a stabilizing nanobody, Nb-C6, that specifically recognized only the NMN-activated form of SARM1. The conformation specificity was verified by immunoprecipitation and surface plasmon resonance. Fluorescently labeled Nb-C6 could immunostain only the activated SARM1 in cells stimulated with CZ-48, a permeant mimetic of NMN. Expression of Nb-C6 in live cells resulted in stabilization of the active form of the endogenous and exogenous SARM1, producing and elevating cellular levels of cyclic ADP-ribose, a calcium messenger. Cryo-EM of the NMN-activated SARM1 complexed by Nb-C6 showed an octameric structure resembling a “blooming lotus” with the ARM domains bending significantly inward and swinging out together with the TIR domains to form the “petals of the lotus”. Nb-C6 bound to the SAM domain of the activated SARM1 and stabilized its Armadillo repeat motif domain. Analyses using hydrogen-deuterium exchange mass spectrometry (HDX-MS), and cross-linking MS (XL-MS) indicate that the activated SARM1 is highly dynamic and flexible and the neighboring TIRs form dimers via the surface close to one BB loop. The Nanobody is thus a valuable tool for delineating the mechanism of activation of SARM1 in AxD and other cellular processes.
Topographic axonal projection at single-cell precision supports local retinotopy in the mouse superior colliculus
Authors: Molotkov Dmitry , Ferrarese Leiron , Boissonnet Tom , Asari Hiroki
Published: March 2022

Retinotopy, like all long-range projections, can arise from the axons themselves or their targets. The underlying connectivity pattern, however, remains elusive at the fine scale in the mammalian brain. To address this question, we functionally mapped the spatial organization of the input axons and target neurons in the mouse retinocollicular pathway at single-cell resolution usingin vivotwo-photon calcium imaging. We found a near-perfect retinotopic tiling of retinal ganglion cell axon terminals, with an average error below 30 μm or 2 degrees of visual angle. The precision of retinotopy was relatively lower for local neurons in the superior colliculus. Subsequent data-driven modelling ascribed it to a low input convergence, on average 5.5 retinal ganglion cell inputs to a postsynaptic cell in the superior colliculus. These results indicate that retinotopy arises largely from topographically precise input from presynaptic cells, rather than elaborating local circuitry to reconstruct the topography by postsynaptic cells.
Model biomolecular condensates have heterogeneous structure quantitatively dependent on the interaction profile of their constituent macromolecules
Authors: Shillcock Julian C. , Lagisquet Clément , Alexandre Jérémy , Vuillon Laurent , Ipsen John H.
Published: March 2022

Biomolecular condensates play numerous roles in cells by selectively concentrating client proteins while excluding others. These functions are likely to be sensitive to the spatial organization of the scaffold proteins forming the condensate. We use coarse-grained molecular simulations to show that model intrinsically-disordered proteins phase separate into a heterogeneous, structured fluid characterized by a well-defined length scale. The proteins are modelled as semi-flexible polymers with punctate, multifunctional binding sites in good solvent conditions. Their dense phase is highly solvated with a spatial structure that is more sensitive to the separation of the binding sites than their affinity. We introduce graph theoretic measures to show that the proteins are heterogeneously distributed throughout the dense phase, an effect that increases with increasing binding site number, and exhibit multi-timescale dynamics. The simulations predict that the structure of the dense phase is modulated by the location and affinity of binding sites distant from the termini of the proteins, while sites near the termini more strongly affect its phase behaviour. The relations uncovered between the arrangement of weak interaction sites on disordered proteins and the material properties of their dense phase can be experimentally tested to give insight into the biophysical properties and rational design of biomolecular condensates.
Sex-specific effects of chronic paternal stress on offspring development are partially mediated via mothers
Authors: Mashoodh Rahia , Habrylo Ireneusz B. , Gudsnuk Kathryn , Champagne Frances A.
Published: March 2022

Paternal stress exposure is known to impact the development of stress-related behaviors in offspring. Previous work has highlighted the importance of sperm mediated factors, such as RNAs, in transmitting the effects of parental stress. However, a key unanswered question is whether mothers’ behavior could drive or modulate the transmission of paternal stress effects on offspring development. Here we investigate how chronic variable stress in Balb/C mice influences the sex-specific development of anxiety- and depression-like neural and behavioral development in offspring. Moreover, we examined how stressed fathers influenced mate maternal investment towards their offspring and how this may modulate the transmission of paternal stress effects on offspring. We show that paternal stress leads to sex-specific effects on offspring behavior. Males that are chronically stressed sire female offspring that show increased anxiety and depression-like behaviors. However, male offspring of stressed fathers show reductions in anxiety- and depression-behaviors and are generally more exploratory. Moreover, we show that females mated with stressed males gain less weight during pregnancy and provide less care towards their offspring which additionally influenced offspring development. These data indicate that paternal stress can influence offspring development directly and indirectly via changes in mothers, with implications for divergent development between male and female offspring.
Petascale pipeline for precise alignment of images from serial section electron microscopy
Authors: Popovych Sergiy , Macrina Thomas , Kemnitz Nico , Castro Manuel , Nehoran Barak , Jia Zhen , Bae J. Alexander , Mitchell Eric , Mu Shang , Trautman Eric T. , Saalfeld Stephan , Li Kai , Seung Sebastian
Published: March 2022

The reconstruction of neural circuits from serial section electron microscopy (ssEM) images is being accelerated by automatic image segmentation methods. Segmentation accuracy is often limited by the preceding step of aligning 2D section images to create a 3D image stack. Precise and robust alignment in the presence of image artifacts is challenging, especially as datasets are attaining the petascale. We present a computational pipeline for aligning ssEM images with several key elements. Self-supervised convolutional nets are trained via metric learning to encode and align image pairs, and they are used to initialize iterative fine-tuning of alignment. A procedure called vector voting increases robustness to image artifacts or missing image data. For speedup the series is divided into blocks that are distributed to computational workers for alignment. The blocks are aligned to each other by composing transformations with decay, which achieves a global alignment without resorting to a time-consuming global optimization. We apply our pipeline to a whole fly brain dataset, and show improved accuracy relative to prior state of the art. We also demonstrate that our pipeline scales to a cubic millimeter of mouse visual cortex. Our pipeline is publicly available through two open source Python packages.
Exosomally targeting microRNA23a ameliorates microvascular endothelial barrier dysfunction following rickettsial infection
Authors: Zhou Changcheng , Bei Jiani , Qiu Yuan , Chang Qing , Nyong Emmanuel , Yang Jun , Krishnan Balaji , Khanipov Kamil , Jin Yang , Fang Xiang , Gaitas Angelo , Gong Bin
Published: March 2022

Spotted fever group rickettsioses caused by Rickettsia (R) are devastating human infections, which mainly target microvascular endothelial cells (EC) and can induce lethal EC barrier dysfunction in the brain and lungs. Our previous evidence reveals that exosomes (Exos) derived from rickettsial-infected ECs, namely R-ECExos, can induce disruption of the tight junctional (TJ) protein ZO-1 and barrier dysfunction of human normal recipient brain microvascular endothelial cells (BMECs). However, the underlying mechanism remains elusive. Given that we have observed that microRNA23a (miR23a), a negative regulator of endothelial ZO-1 mRNA, is selectively sorted into R-ECExos, the aim of the present study was to characterize the potential functional role of exosomal miR23a delivered by R-ECExos in normal recipient BMECs. We demonstrated that EC-derived Exos (ECExos) have the capacity to deliver oligonucleotide RNAs to normal recipient BMECs in an RNase-abundant environment. miR23a in ECExos impairs normal recipient BMEC barrier function, directly targeting TJ protein ZO-1 mRNAs. In separate studies using a traditional in vitro model and a novel single living-cell biomechanical assay, our group demonstrated that miR23a anti-sense oligonucleotide-enriched ECExos ameliorate R-ECExo-provoked recipient BMEC dysfunction in association with stabilization of ZO-1 in a dose-dependent manner. These results suggest that Exo-based therapy could potentially prove to be a promising strategy to improve vascular barrier function during bacterial infection and concomitant inflammation.
Targeting tumor stemness switch phenotype by activating pathogen induced stem cell niche defense
Authors: Bhuyan Seema , Pal Bidisha , Pathak Lekhika , Saikia Partha Jyoti , Mitra Shirsajit , Gayan Sukanya , Mokhtari Reza Bayat , Li Hong , Ramana Chilakamarti V , Baishya Debabrat , Das Bikul
Published: March 2022

Cancer stem cells (CSCs) reside in their tumor microenvironment (TME) niches, which are often hypoxic. Previously, we found that hypoxia and oxidative stress prevalent in TME may reprogram CSCs to a highly aggressive and inflammatory phenotype, the tumor stemness switch (TSS) phenotype. We previously reported a “stem cell niche defense” mechanism in bone marrow and lung mesenchymal stem cell niche against pathogen. Pathogen induced bystander apoptosis (PIBA) of stem cells harboring intracellular pathogen may be part of this defense mechanism. We speculate that the TSS phenotype may also activate this niche defense mechanism to defend their TME niche against pathogen and therefore could be exploited to target CSCs. Here we report that CSCs of TSS phenotype enriched in post-hypoxia ABCG2+ CSCs of several cell lines of diverse tumors including oral squamous cell carcinoma cell line SCC-25 exhibited bystander apoptosis when infected with either Bacillus Calmette Guerin (BCG) or mutantMycobacterium tuberculosis (Mtb) strain 18b. The conditioned media (CM) of the infected cells not only exhibited marked anti-tumor activity in vivo, but also showed significant anti-microbial activity. A detailed mechanisms study revealed that some of the infected ABCG2+ CSCs underwent pyroptosis and released a high mobility group box protein 1 (HMGB1)/p53 death signal that can induce a toll like receptor (TLR) 2/4 mediated bystander apoptosis. Thus, our findings suggest that PIBA can be utilized to activate the “niche defense” mechanism in TSS phenotype, which not only target the TSS, but also exhibit marked anti-tumor activity in vivo.
Structure-guided synthesis of FK506 and FK520 analogs with increased selectivity exhibit in vivo therapeutic efficacy against Cryptococcus
Authors: Hoy Michael J. , Park Eunchong , Lee Hyunji , Lim Won Young , Cole D. Christopher , DeBouver Nicholas D. , Bobay Benjamin G. , Pierce Phillip G. , Fox David , Ciofani Maria , Juvvadi Praveen R. , Steinbach William , Hong Jiyong , Heitman Joseph
Published: March 2022

Calcineurin is an essential virulence factor that is conserved across human fungal pathogens includingCryptococcus neoformans,Aspergillus fumigatus, andCandida albicans. Although an excellent target for antifungal drug development, the serine-threonine phosphatase activity of calcineurin is conserved in mammals and inhibition of this activity results in immunosuppression. FK506 (tacrolimus) is a naturally produced macrocyclic compound that inhibits calcineurin by binding to the immunophilin FKBP12. Previously, our fungal calcineurin-FK506-FKBP12 structure-based approaches identified a non-conserved region of FKBP12 that can be exploited for fungal-specific targeting. These studies led to the design of an FK506 analog, APX879, modified at the C22 position that was less immunosuppressive yet maintained antifungal activity. We now report high resolution protein crystal structures of fungal FKBP12 and a human truncated calcineurin-FKBP12 bound to a natural FK506 analog, FK520 (ascomycin). Based on information from these structures and the success of APX879, we synthesized and screened a novel panel of C22-modified compounds derived from both FK506 and FK520. One compound, JH-FK-05 demonstrates broad-spectrum antifungal activityin vitroand is non-immunosuppressivein vivo. In murine models of pulmonary and disseminatedC. neoformansinfection, JH-FK-05 treatment significantly reduced fungal burden and extended animal survival alone and in combination with fluconazole. Furthermore, molecular dynamic simulations performed with JH-FK-05 binding to fungal and human FKBP12 identified additional residues outside of the C22 and C21 positions that could be modified to generate novel FK506 analogs with improved antifungal activity.

Significance

Due to rising rates of antifungal drug resistance and a limited armamentarium of antifungal treatments, there is a paramount need for novel antifungal drugs to treat systemic fungal infections. Calcineurin has been established as an essential and conserved virulence factor in several fungi, making it an attractive antifungal target. However, due to the immunosuppressive action of calcineurin inhibitors, they have not been successfully utilized clinically for antifungal treatment in humans. Recent availability of crystal structures of fungal calcineurin bound inhibitor complexes have enabled the structure-guided design of FK506 analogs and led to a breakthrough in the development of a compound increased for fungal specificity. The development of a calcineurin inhibitor with reduced immunosuppressive activity and therapeutic antifungal activity would add a significant tool to the treatment options for these invasive fungal infections with exceedingly high rates of mortality.
Facultative parthenogenesis: a transient state in transitions between sex and obligate asexuality in stick insects?
Authors: Larose Chloé , Lavanchy Guillaume , Freitas Susana , Parker Darren J. , Schwander Tanja
Published: March 2022

Transitions from obligate sex to obligate parthenogenesis have occurred repeatedly across the tree of life. Whether these transitions occur abruptly or via a transient phase of facultative parthenogenesis is rarely known. We discovered and characterised facultatively parthenogenetic populations of the North American stick insectTimema douglasi, a species in which only obligately parthenogenetic populations were known so far. These populations comprised three genetic lineages. Females from all lineages were capable of parthenogenesis (with variable efficiency) but their propensity to reproduce sexually after mating varied extensively. In all three lineages, parthenogenesis resulted in the complete loss of heterozygosity in a single generation. Obligately parthenogeneticTimemahave also lost all heterozygosity, suggesting that the transition to obligate parthenogenesis did not require a modification of the proximate mechanism, but rather involved a gradual increase in frequency. We speculate that facultative parthenogenesis may often be transient and be replaced by obligate strategies (either sex or parthenogenesis) because of a trade-off between the efficiency of the two reproductive modes. Such a trade-off could help explain why facultative parthenogenesis is rare among animals, despite its potential to combine the known benefits of sex and parthenogenesis.
Treatment with an Antigen-Specific Dual Microparticle System Reverses Advanced Multiple Sclerosis in Mice
Authors: Kwiatkowski Alexander J , Helm Eric Y , Stewart Joshua M , Drashansky Theodore T , Cho Jonathan J , Avram Dorina , Keselowsky Benjamin G
Published: March 2022

Antigen-specific therapies hold promise for treating autoimmune diseases such as multiple sclerosis while avoiding the deleterious side effects of systemic immune suppression. In this study, an antigen-specific dual-sized microparticle (dMP) immunotherapy reversed hind limb paralysis when administered in mice with advanced experimental autoimmune encephalomyelitis (EAE). Treatment reduced central nervous system (CNS) immune cell infiltration, demyelination and inflammatory cytokine levels. Mechanistic insights using single-cell RNA sequencing showed that treatment impacted the MHC II antigen presentation pathway in dendritic cells, macrophages, B cells and microglia, not only in the draining lymph nodes, but strikingly also in the spinal cord. CD74 and cathepsin S were among the common genes downregulated in most antigen presenting cell (APC) clusters, with B cells also having numerous MHC II genes reduced. Efficacy of the treatment diminished when B cells were absent, suggesting their impact in this therapy, in concert with other immune populations. Activation and inflammation were reduced in both APCs and T cells. This promising antigen-specific therapeutic approach advantageously engaged essential components of both innate and adaptive autoimmune responses, and capably reversed paralysis in advanced EAE without the use of broad immunosuppressant.

Significance Statement

Multiple sclerosis (MS) is a debilitating autoimmune disease that can lead to paralysis. We demonstrate an antigen-specific microparticle treatment can reverse hind limb paralysis when administered in advanced EAE. Single-cell RNA-sequencing and flow cytometry analysis provide evidence the treatment acts by diminishing Ag presentation in APCs, including B cells in the CNS and the draining lymph nodes. Thus, the antigen-specific dual-sized microparticle treatment is a promising therapy even in advanced EAE, and potentially MS.
Single-cell transcriptomic atlas of the human substantia nigra in Parkinson’s disease
Authors: Wang Qian , Wang Minghui , Choi Insup , Ho Lap , Farrell Kurt , Beaumont Kristin G. , Sebra Robert , Crary John F , Davis David A. , Sun Xiaoyan , Zhang Bin , Yue Zhenyu
Published: March 2022

Parkinson’s disease (PD) is a common and complex neurodegenerative disorder. Loss of neuromelanin-containing dopaminergic (DA) neurons in the substantia nigra (SN) is a hallmark of PD neuropathology; the etiology of PD remains unclear. Single-cell (-nucleus) RNA sequencing (sc or snRNAseq) has significantly advanced our understanding of neurodegenerative diseases including Alzheimer’s, but limited progress has been made in PD. Here we generated by far the largest snRNAseq data of high-quality 315,867 nuclei from the human SN including 9 healthy controls and 23 idiopathic PD cases across different Braak stages. Clustering analysis identified major brain cell types including DA neurons, excitatory neurons, inhibitory neurons, glial cells, endothelial, pericytes, fibroblast and T-cells in the human SN. By combining immunostaining and validating against the datasets from independent cohorts, we identified three molecularly distinct subtypes of DA-related neurons, including a RIT2-enriched population, in human aged SN. All DA neuron subtypes degenerated in PD, whereas the composition of non-neuronal cell clusters including major glial types showed little change. Our study delineated cell-type-specific PD-linked gene expression in the SN and their alterations in PD. Examination of cell-type-based transcriptomic changes suggests the complexity and diversity of molecular mechanisms of PD. Analysis of the remaining DA neurons of the three subtypes from PD demonstrated alterations of common gene sets associated with neuroprotection. Our findings highlight the heterogeneity of DA neurons in the human SN and suggest molecular basis for vulnerability and resilience of human DA neurons in PD. Our cohort thus provides a valuable resource for dissecting detailed mechanisms of DA neuron degeneration and identifying new neuroprotective strategies for PD.
Chemical inhibition of prolyl hydroxylases impairs angiogenic competence of human vascular endothelium through metabolic reprogramming
Authors: Tiwari Ratnakar , Bommi Prashant V. , Gao Peng , Schipma Matthew J. , Zhou Yalu , Quaggin Susan E. , Chandel Navdeep S. , Kapitsinou Pinelopi P.
Published: March 2022

Endothelial cell (EC) metabolism has emerged as a driver of angiogenesis. While hypoxia inactivates prolyl-4 hydroxylase domain containing proteins 1-3 (PHD1-3) and stabilizes hypoxia inducible factors (HIFs) stimulating angiogenesis, the effects of PHDs on EC functions remain unclear. Here, we investigated the impact of PHD inhibition by dimethyloxalylglycine (DMOG) on angiogenic competence and metabolism of human vascular ECs. PHD inhibition reduced EC proliferation, migration, and tube formation capacities. Furthermore, transcriptomic and metabolomic analyses revealed an unfavorable metabolic reprogramming for angiogenesis following treatment with DMOG. Despite the induction of glycolytic genes and high levels of lactate, multiple genes encoding sub-units of mitochondrial complex I were suppressed with concurrent decline in nicotinamide adenine dinucleotide (NAD+) levels. Importantly, defective EC migration due to DMOG could be partially restored by augmenting NAD+ levels. Combined, our data provide metabolic insights into the mechanism by which chemical PHD inhibition impairs angiogenic competence of human vascular ECs.
Pharmacological manipulations of physiological arousal and sleep-like slow waves modulate sustained attention
Authors: Pinggal Elaine , Dockree Paul M. , O’Connell Redmond G. , Bellgrove Mark A. , Andrillon Thomas
Published: March 2022

Sustained attention describes our ability to keep a constant focus on a given task. This ability is modulated by our physiological state of arousal. Although lapses of sustained attention have been linked with dysregulations of arousal, the underlying physiological mechanisms remain unclear. An emerging body of work proposes that the intrusion during wakefulness of sleep-like slow waves, a marker of the transition toward sleep, could mechanistically account for attentional lapses. This study aimed to expose, via pharmacological manipulations of the monoamine system, the relationship between the occurrence of sleep-like slow waves and the behavioural consequences of sustained attention failures. In a double-blind, randomised-control trial, 32 healthy human male participants received methylphenidate, atomoxetine, citalopram or placebo during four separate experimental sessions. During each session, electroencephalography (EEG) was used to measure neural activity whilst participants completed a visual task requiring sustained attention. Methylphenidate, which increases wake-promoting dopamine and noradrenaline across cortical and subcortical areas, improved behavioural performance whereas atomoxetine, which increases dopamine and noradrenaline predominantly over frontal cortices, led to more impulsive responses. Additionally, citalopram, which increases sleep-promoting serotonin, led to more missed trials. Based on EEG recording, citalopram was also associated with an increase in sleep-like slow waves. Importantly, compared to a classical marker of arousal such as alpha power, only slow waves differentially predicted both misses and faster responses in a region-specific fashion. These results suggest that a decrease in arousal can lead to local sleep intrusions during wakefulness which could be mechanistically linked to impulsivity and sluggishness.

Significance Statement

We investigated whether the modulation of attention and arousal could not only share the same neuromodulatory pathways but also rely on similar neuronal mechanisms; for example, the intrusion of sleep-like activity within wakefulness. To do so, we pharmacologically manipulated noradrenaline, dopamine, and serotonin in a four-arm, randomised, placebo-controlled trial and examined the consequences on behavioural and EEG indices of attention and arousal. We showed that sleep-like slow waves can predict opposite behavioural signatures: impulsivity and sluggishness. Slow waves may be a candidate mechanism for the occurrence of attentional lapses since the relationship between slow-wave occurrence and performance is region-specific and the consequences of these local sleep intrusions are in line with the cognitive functions carried by the underlying brain regions.
SpyChIP identifies cell type-specific transcription factor occupancy from complex tissues
Authors: Feng Siqian , Mann Richard S.
Published: March 2022

Chromatin immunoprecipitation (ChIP) is an important technique for characterizing protein-DNA binding in vivo. One drawback of ChIP based techniques is the lack of cell type-specificity when profiling complex tissues. To overcome this limitation, we developed SpyChIP to identify cell type-specific transcription factor (TF) binding sites in native physiological contexts without tissue dissociation or nuclei sorting. SpyChIP takes advantage of a specific covalent isopeptide bond that rapidly forms between the 15 amino acid SpyTag and the 17 kD protein SpyCatcher. In SpyChIP, the target TF is fused with SpyTag by genome engineering, and an epitope tagged SpyCatcher is expressed in cell populations of interest, where it covalently binds to SpyTag-TF. Cell type-specific ChIP is obtained by immunoprecipitating chromatin prepared from whole tissues using antibodies directed against the epitope-tagged SpyCatcher. Using SpyChIP, we identified the genome-wide binding profiles of the Hox protein Ubx in two distinct cell types of the Drosophila haltere disc. Our results revealed extensive region-specific Ubx-DNA binding events, highlighting the significance of cell type-specific ChIP and the limitations of whole tissue ChIP approaches. Analysis of Ubx::SpyChIP results provided novel insights into the relationship between chromatin accessibility and Ubx-DNA binding, as well as different mechanisms Ubx employs to regulate its downstream cis-regulatory modules (CRMs). In addition to SpyChIP, we suggest that SpyTag-SpyCatcher technology, as well as other covalent interaction peptide pairs, has many potential in vivo applications that were previously unachievable.
Charge transfer of unconfined regions in quasi-periodic quantum wells : DNA nanostructure study
Authors: Tai Alan
Published: March 2022

This study presents a quantum well model using transfer matrix to analyse the charge transfer characteristic in DNA nanostructure sequences. The unconfined state or unbound state above the quantum well are used to investigate the carrier behaviours in nanostructure of semiconductor. Similar analysis and simulations can be applied to the understanding of charge transfer in DNA nanostructure with sequence of periodic and quasi-periodic sequences. The model was validated with experiments using photoreflectance spectroscopy on nanostructure in semiconductor superlattice. In addition, the results published by the study of Thermoelectric effect and its dependence on molecular length and sequence in single DNA molecules[1] were used to compare with the simulations by the quantum well model. The results show agreement and can provide new insight into charge transfer and transport in DNA nanostructure with different types of sequences.
A gut commensal protozoa remotely shapes a lung niche for asthma-promoting eosinophils
Authors: Burrows Kyle , Ngai Louis , Chiaranunt Pailin , Cao Eric , Streutker Catherine J. , Forde Brian , O’Mahony Liam , Mortha Arthur
Published: March 2022

The gut microbiome influences chronic inflammation of the airways via the gut-lung axis. However, causal connections between microbes and their host, including the underlying mechanisms for this phenomenon remain largely unknown. Here, we show that colonization with the gut commensal protozoa,Tritrichomonas musculis(T.mu), remotely shapes the lung immune landscape and exacerbates allergic airway inflammation. We demonstrate that colonization withT.mu mediates the T and B cell-dependent accumulation and activation of inflammatory group 2 innate lymphoid cells in the lungs to constitute a tripartite immune network that serves as a niche for lung eosinophils. Animals colonized withT.mushow severely exacerbated allergic inflammation in the airways and reveal a new protozoan-driven gut-lung axis that remotely shapes the lung immune network to potentiate chronic pulmonary inflammation.

One-Sentence Summary

A gut microbe exacerbates asthma severity by promoting lung eosinophilia through a tripartite lymphocyte immune network.
Candida albicans filamentation does not require the cAMP-PKA pathway in vivo
Authors: Wakade Rohan S. , Kamara Juraj , Wellington Melanie , Krysan Damian J.
Published: March 2022

Candida albicans is one of the most prevalent human fungal pathogens. Its ability to transition between budding yeast and filamentous morphological forms (pseudohyphae and hyphae) is tightly associated with its pathogenesis. Based on in vitro studies, the cAMP-Protein Kinase A (PKA) pathway is a key regulator of C. albicans morphogenesis. Using an intravital imaging approach, we investigated the role of the cAMP-PKA pathway during infection. Consistent with their roles in vitro, the downstream effectors of the cAMP-PKA pathway Efg1 and Nrg1 function, respectively, as an activator and a repressor of in vivo filamentation. Surprisingly, strains lacking the adenylyl cyclase, CYR1, showed only slightly reduced filamentation in vivo despite being completely unable to filament in RPMI+10% serum at 37°C. Consistent with these findings, deletion of the catalytic subunits of PKA (Tpk1 and Tpk2), either singly or in combination, generated strains that also filamented in vivo but not in vitro. In vivo transcription profiling of C. albicans isolated from both ear and kidney tissue showed that the expression of a set of 184 environmentally responsive correlated well with in vitro filamentation (R2 0.62-0.68) genes. This concordance suggests that the in vivo and in vitro transcriptional responses are similar but that the upstream regulatory mechanisms are distinct. As such, these data emphatically emphasize that C. albicans filamentation is a complex phenotype that occurs in different environments through an intricate network of distinct regulatory mechanisms.

Importance

The fungus Candida albicans causes a wide range of disease in humans from common diaper rash to life-threatening infections in patients with compromised immune systems. As such, the mechanisms for its ability to cause disease are of wide interest. An intensely studied virulence property of C. albicans is its ability to switch from a round yeast form to filament-like forms (hyphae and pseudohyphae). Surprisingly, we have found that a key signaling pathway that regulates this transition in vitro, the protein kinase A pathway, is not required for filamentation during infection of the host. Our work not only demonstrates that the regulation of filamentation depends upon the specific environment C. albicans inhabits but also underscores the importance of studying these mechanisms during infection.
Drug design and repurposing with a sequence-to-drug paradigm
Authors: Chen Lifan , Fan Zisheng , Chang Jie , Yang Ruirui , Guo Hao , Zhang Yinghui , Yang Tianbiao , Zhou Chenmao , Chen Zhengyang , Zheng Chen , Hao Xinyue , Zhang Keke , Cui Rongrong , Ding Yiluan , Zhang Naixia , Luo Xiaomin , Jiang Hualiang , Zhang Sulin , Zheng Mingyue
Published: March 2022

Drug development based on target proteins has been a successful approach in recent decades. A conventional structure-based drug design pipeline is a complex, human-engineered pipeline with multiple independently optimized steps. Advances in end-to-end differentiable learning suggest the potential benefits of similarly reformulating drug design. Here, we proposed a new sequence-to-drug paradigm that discovers drug-like small-molecule modulators directly from protein sequences and validated this concept for the first time in three stages. First, we designed TransformerCPI2.0 as a core tool for the sequence-to-drug paradigm, which exhibited competitive performance with conventional structure-based drug design approaches. Second, we validated the binding knowledge that TransformerCPI2.0 has learned. Third, we applied a sequence-to-drug paradigm to discover new hits for E3 ubiquitin-protein ligases: speckle-type POZ protein (SPOP), ring finger protein 130 (RNF130) which does not have a 3D structure, and repurposed proton pump inhibitors (PPIs) for ADP-ribosylation factor 1 (ARF1). This first proof of concept shows that the sequence-to-drug paradigm is a promising direction for drug development.