bluuebunny/crossref_metadata_2025 huggingface dataset:
Earthworm and Soil Microbial Communities in Flower Strips
Authors: Bednar Zita , Vaupel Anna , Blümel Simon , Herwig Nadine , Hommel Bernd , Haberlah-Korr Verena , Beule Lukas
Published: December 2022

Flower strips are a common agricultural practice to increase aboveground biodiversity and beneficial ecosystem services. Although soil communities are a key component of terrestrial biodiversity and drive important ecosystem services, their abundance, diversity, and composition in flower strips remain largely unexplored. Here, we shed light on earthworms and soil microorganisms in flower strips and aim to provide a starting point for research on belowground communities in flower strips. In 2020, we established a field margin vegetation as well as two annual and two perennial flower strip mixtures at three study sites in Germany that were previously conventional croplands or fallow. Two years following this conversion, we determined earthworm communities and investigated the soil microbiome using real-time PCR (archaea, bacteria, fungi, and soil-N-cycling genes) and amplicon sequencing (bacteria and fungi). Different plant mixtures (i.e. field margin, annual, and perennial flower strips) harbored distinct earthworm and soil microbial communities. Earthworm density and biomass declined or remained unaffected in annual flower strips but increased in perennial flower strips as compared to field margins. Arbuscular mycorrhizal fungi showed greater diversity and community share in non-tilled (i.e. field margin and perennial flower strips) as compared to tilled plant mixtures (i.e. annual flower strips). We attribute changes in earthworms and microorganisms mainly to the effect of tillage and plant diversity. Overall, we suggest that perennial flower strips serve as refugia for soil biota in agricultural landscapes. Future studies should compare soil biota in perennial flower strips to those in adjacent fields and investigate whether beneficial belowground effects are restricted to the flower strips or spatially extend into adjacent fields (‘spillover’).
Distinct myofibre domains of the human myotendinous junction revealed by single nucleus RNA-seq
Authors: Karlsen Anders , Yeung Ching-Yan Chloé , Schjerling Peter , Denz Linda , Hoegsbjerg Christian , Jakobsen Jens R. , Krogsgaard Michael R. , Koch Manuel , Schiaffino Stefano , Kjaer Michael , Mackey Abigail L.
Published: December 2022

The myotendinous junction (MTJ) is a specialized domain of the multinucleated myofibre, faced with the challenge of maintaining robust cell-matrix contact with the tendon under high mechanical stress and strain. Here, we profiled 24,161 nuclei in semitendinosus muscle-tendon samples from 3 healthy males by single nucleus RNA-sequencing (snRNA-seq), alongside spatial transcriptomics, to gain insight into the genes characterizing this specialization in humans. We identified a cluster of MTJ myonuclei, represented by 47 enriched transcripts, of which the presence ofABI3BP, ABLIM1, ADAMTSL1, BICD1, CPM, FHOD3, FRAS1andFREM2was confirmed at the MTJ at the protein level by immunofluorescence. Four distinct subclusters of MTJ myonuclei were apparent and segregated into two COL22A1-expressing subclusters and two lacking COL22A1 but with a clear fibre type profile expressingMYH7orMYH1/2. Our findings reveal distinct myonuclei profiles of the human MTJ, a weak link in the musculoskeletal system, which is selectively affected in pathological conditions, from muscle strains to muscular dystrophies.
A high-throughput COPD bronchosphere model for disease-relevant phenotypic compound screening
Authors: Beri Pranjali , Woo Young Jae , Schierenbeck Katie , Chen Kaisheng , Barnes S. Whitney , Ross Olivia , Krutil Douglas , Quackenbush Doug , Fang Bin , Walker John , Barnes William , Toyama Erin
Published: December 2022

COPD is the third leading cause of death worldwide, but current therapies for COPD are only effective at treating the symptoms of the disease rather than targeting the underlying pathways that are driving the pathogenic changes. The lack of targeted therapies for COPD is in part due to a lack of knowledge about drivers of disease progression and the difficulty in building relevant and high throughput models that can recapitulate the phenotypic and transcriptomic changes associated with pathogenesis of COPD. To identify these drivers, we have developed a cigarette smoke extract (CSE)-treated bronchosphere assay in 384-well plate format that exhibits CSE-induced decreases in size and increase in luminal secretion of MUC5AC. Transcriptomic changes in CSE-treated bronchospheres resemble changes that occur in human smokers both with and without COPD compared to healthy groups, indicating that this model can capture human smoking signature. To identify new targets, we ran a small molecule compound deck screening with diversity in target mechanisms of action and identified hit compounds that attenuated CSE induced changes, either decreasing spheroid size or increasing secreted mucus. This work provides insight into the utility of this bronchosphere model in examining human respiratory diseases, the pathways implicated by CSE, and compounds with known mechanisms of action for therapeutic development.
Gestational stress decreases postpartum mitochondrial respiration in the prefrontal cortex of female rats
Authors: Gorman-Sandler Erin , Robertson Breanna , Crawford Jesseca , Arishe Olufunke O. , Clinton Webb R. , Hollis Fiona
Published: December 2022

Postpartum depression (PPD) is a major psychiatric complication of childbirth, affecting up to 20% of mothers, yet remains understudied. Mitochondria, dynamic organelles crucial for cell homeostasis and energy production, share links with many of the proposed mechanisms underlying PPD pathology. Brain mitochondrial function is affected by stress, a major risk factor for development of PPD, and is linked to anxiety-like and social behaviors. Considering the importance of mitochondria in regulating brain function and behavior, we hypothesized that mitochondrial dysfunction is associated with behavioral alterations in a chronic stress-induced rat model of PPD. Using a validated and translationally relevant chronic mild unpredictable stress paradigm during late gestation, we induced PPD-relevant behaviors in adult postpartum Wistar rats. In the mid-postpartum, we measured mitochondrial function in the prefrontal cortex (PFC) and nucleus accumbens (NAc) using high-resolution respirometry. We then measured protein expression of mitochondrial complex proteins and 4-hydroxynonenal (a marker of oxidative stress), and Th1/Th2 cytokine levels in PFC and plasma. We report novel findings that gestational stress decreased mitochondrial function in the PFC, but not the NAc of postpartum dams. However, in groups controlling for the effects of either stress or parity alone, no differences in mitochondrial respiration measured in either brain regions were observed compared to nulliparous controls. This decrease in PFC mitochondrial function in stressed dams was accompanied by negative behavioral consequences in the postpartum, complex-I specific deficits in protein expression, and increased Tumor Necrosis Factor alpha cytokine levels in plasma and PFC. Overall, we report an association between PFC mitochondrial respiration, PPD-relevant behaviors, and inflammation following gestational stress, highlighting a potential role for mitochondrial function in postpartum health.
Nodavirus RNA Replication Crown Architecture Reveals Proto-Crown Precursor and Viral Protein A Conformational Switching
Authors: Zhan Hong , Unchwaniwala Nuruddin , Rebolledo-Viveros Andrea , Pennington Janice , Horswill Mark , Broadberry Roma , Myers Jonathan , den Boon Johan A. , Grant Timothy , Ahlquist Paul
Published: December 2022

Positive-strand RNA viruses replicate their genomes in virus-induced membrane vesicles, and the resulting RNA replication complexes are a major target for virus control. Nodavirus studies first revealed viral RNA replication proteins forming a 12-fold symmetric “crown” at the vesicle opening to the cytosol, an arrangement recently confirmed to extend to distantly related alphaviruses. Using cryo-electron microscopy (cryo-EM), we show that mature nodavirus crowns comprise two stacked 12-mer rings of multi-domain viral RNA replication protein A. Each ring contains an~19 nm circle of C-proximal polymerase domains, differentiated by strikingly diverged positions of N-proximal RNA capping/membrane binding domains. The lower ring is a “proto-crown” precursor that assembles prior to RNA template recruitment, RNA synthesis and replication vesicle formation. In this proto-crown, the N-proximal segments interact to form a toroidal central floor, whose 3.1 Å resolution structure reveals many mechanistic details of the RNA capping/membrane binding domains. In the upper ring, cryo-EM fitting indicates that the N-proximal domains extend radially outside the polymerases, forming separated, membrane-binding “legs.” The polymerase and N-proximal domains are connected by a long linker accommodating the conformational switch between the two rings and possibly also polymerase movements associated with RNA synthesis and non-symmetric electron density in the lower center of mature crowns. The results reveal remarkable viral protein multifunctionality, conformational flexibility and evolutionary plasticity and new insights into (+)RNA virus replication and control.

Significance

Positive-strand RNA viruses - including coronaviruses, alphaviruses, flaviviruses and many other medically and economically important pathogens - replicate their RNA genomes by virus-encoded machinery that has been poorly characterized. Using an advanced nodavirus model, we identify a major precursor in RNA replication complex assembly and show it to be a 12-mer ring of viral RNA replication protein A, whose single particle cryo-EM structure reveals functional features of its membrane interaction, assembly, polymerase and RNA capping domains. We further show that fully functional RNA replication complexes acquire a second 12-mer ring of protein A in alternate conformation atop the first, and a central density likely to represent another polymerase conformation. These findings provide strong foundations for understanding, controlling and beneficially using such viruses.
UBXN1 maintains ER proteostasis and represses UPR activation by modulating translation independently of the p97 ATPase
Authors: Ahlstedt Brittany A. , Ganji Rakesh , Mukkavalli Sirisha , Paulo Joao A. , Gygi Steve P. , Raman Malavika
Published: December 2022

Endoplasmic reticulum (ER) protein homeostasis (proteostasis) is essential to facilitate proper folding and maturation of proteins in the secretory pathway. Loss of ER proteostasis due to cell stress or mutations in ER proteins can lead to the accumulation of misfolded or aberrant proteins in the ER and triggers the unfolded protein response (UPR). In this study we find that the p97 adaptor UBXN1 is an important negative regulator of the UPR. Loss of UBXN1 significantly sensitizes cells to ER stress and activates canonical UPR signaling pathways. This in turn leads to widespread upregulation of the ER stress transcriptional program. Using comparative, quantitative proteomics we show that deletion of UBXN1 results in a significant enrichment of proteins belonging to ER-quality control processes including those involved in protein folding and import. Notably, we find that loss of UBXN1 does not perturb p97-dependent ER associated degradation (ERAD). Our studies indicate that loss of UBXN1 increases translation in both resting and ER-stressed cells. Surprisingly, this process is independent of p97 function. Taken together, our studies have identified a new role for UBXN1 in repressing translation and maintaining ER proteostasis in a p97 independent manner.
Neuron secreted chemokine-like Orion is involved in the transformation of glial cells into phagocytes in different neuronal remodeling paradigms
Authors: Perron Clarisse , Carme Pascal , Rosell Arnau Llobet , Minnaert Eva , Demoulin Salomé Ruiz , Szczkowski Héloïse , Neukomm Lukas Jakob , Dura Jean-Maurice , Boulanger Ana
Published: December 2022

During animal development, neurons often form exuberant or incorrect axons and dendrites at early stages, followed by the refinement of neuronal circuits at late stages. Neural circuit refinement leads to the production of large amounts of neuronal debris in the form of neuronal cell corpses, fragmented axons and dendrites, and pruned synapses requiring disposal. In particular, the predominant phagocytes acting during the neuronal remodeling and degeneration are glial cells and critical signaling pathways between neurons and glia leading to phagocytosis are required. Chemokine-like mushroom body neuron secreted Orion ligand was shown to be essential to the astrocyte infiltration into the γ axon bundle leading to γ axon pruning and clearance of debris left from axon fragmentation. Here we show a role oforionalso in debris engulfment and phagocytosis. Interestingly, we show thatorionis also involved in the overall transformation of astrocytes into phagocytes. In addition, analysis of several neuronal paradigms demonstrates the role oforionin the elimination of both peptidergic vCrz+and PDF-Tri neuronsviaadditional phagocytic glial cells as cortex and/or ensheathing glia. Our results suggest that Orion is essential for phagocytic activation of three different types of glial cells: astrocytes, cortex and ensheathing glia and point to Orion as a trigger not only of glial infiltration but also engulfment and phagocytosis.
Improved humoral immunity and protection against influenza virus infection with a 3D porous biomaterial vaccine
Authors: Miwa Hiromi , Antao Olivia Q , Kelly-Scumpia Kindra M. , Baghdasarian Sevana , Mayer Daniel P. , Shang Lily , Sanchez Gina M. , Archang Maani M , Scumpia Philip O. , Weinstein Jason S , Carlo Dino Di
Published: December 2022

New vaccine platforms that properly activate humoral immunity and generate neutralizing antibodies are required to combat emerging and re-emerging pathogens, including influenza virus. Biomaterial scaffolds with macroscale porosity have demonstrated tremendous promise in regenerative medicine where they have been shown to allow immune cell infiltration and subsequent activation, but whether these types of materials can serve as an immunization platform is unknown. We developed an injectable immunization platform that uses a slurry of antigen-loaded hydrogel microparticles that anneal to form a porous scaffold with high surface area for antigen uptake by infiltrating immune cells as the biomaterial degrades to maximize humoral immunity. Antigen-loaded-microgels elicited a robust cellular humoral immune response, with increased CD4+T follicular helper (Tfh) cells and prolonged germinal center (GC) B cells comparable to the commonly used adjuvant, aluminum hydroxide (Alum). By simply increasing the weight fraction of polymer material, we enhanced material stiffness and further increased antigen-specific antibody titers superior to Alum. Vaccinating mice with inactivated influenza virus loaded into this more highly crosslinked formulation elicited a strong antibody response and provided better protection against a high dose viral challenge than Alum. Thus, we demonstrate that by tuning physical and chemical properties alone, we can enhance adjuvanticity and promote humoral immunity and protection against a pathogen, leveraging two different types of antigenic material: individual protein antigen and inactivated virus. The flexibility of the platform may enable design of new vaccines to enhance innate and adaptive immune cell programming to generate and tune high affinity antibodies, a promising approach to generate long-lasting immunity against specific pathogens.
novoRNABreak: local assembly for novel splice junction and fusion transcript detection from RNA-seq data
Authors: Tan Yukun , Mohanty Vakul , Liang Shaoheng , Ma Jun , Kim Kun Hee , Bonder Marc Jan , Shi Xinghua , Lee Charles ,   Human Genome Structural Variation Consortium, Chong Zechen , Chen Ken
Published: December 2022

We present novoRNABreak, a unified framework for cancer specific novel splice junction and fusion transcript detection in RNA-seq data obtained from human cancer samples. novoRNABreak is based on a local assembly model, which offers a tradeoff between the alignment-based and de novo whole transcriptome assembly (WTA) approaches, namely, being more sensitive in assembling novel junctions that cannot be directly aligned, and more efficient due to the strategy that focuses on junctions rather than full-length transcripts. The performance of novoRNABreak is demonstrated by a comprehensive set of experiments using synthetic data generated based on genome reference, as well as real RNA-seq data from breast cancer and prostate cancer samples. The results show that novoRNABreak can detect novel splice junctions and fusion transcripts efficiently with high sensitivity and reasonable specificity.
Primary infection with Zika virus provides one-way heterologous protection against Spondweni virus infection in rhesus macaques
Authors: Jaeger Anna S. , Crooks Chelsea M. , Weiler Andrea M. , Bliss Mason I. , Rybarczyk Sierra , Richardson Alex , Einwalter Morgan , Peterson Eric , Capuano Saverio , Barkhymer Alison , Becker Jordan T. , Greene Joseph T. , Freedman Tanya S. , Langlois Ryan A. , Friedrich Thomas C. , Aliota Matthew T.
Published: December 2022

Spondweni virus (SPONV) is the closest known relative of Zika virus (ZIKV). SPONV pathogenesis resembles that of ZIKV in pregnant mice, and both viruses are transmitted byAedes aegyptimosquitoes. We aimed to develop a translational model to further understand SPONV transmission and pathogenesis. We found that cynomolgus macaques (Macaca fascicularis) inoculated with ZIKV or SPONV were susceptible to ZIKV, but resistant to SPONV infection. In contrast, rhesus macaques (Macaca mulatta) supported productive infection with both ZIKV and SPONV and developed robust neutralizing antibody responses. Crossover serial challenge in rhesus macaques revealed that SPONV immunity did not protect against ZIKV infection, whereas ZIKV immunity was fully protective against SPONV infection. These findings establish a viable model for future investigation into SPONV pathogenesis, and suggest the risk of SPONV emergence is low in areas with high ZIKV seroprevalence due to one-way cross-protection between ZIKV and SPONV.

Teaser

Identification of asymmetric immune interactions between Zika and Spondweni viruses in macaque monkeys.
The microRNomes of Chinese Hamster Ovary (CHO) cells and their extracellular vesicles, and how they respond to osmotic and ammonia stress
Authors: Belliveau Jessica , Papoutsakis Eleftherios T.
Published: December 2022

A new area of focus in Chinese Hamster Ovary (CHO) biotechnology is the role of small (exosomes) and large (microvesicles or microparticles) extracellular vesicles (EVs). CHO cells in culture exchange large quantities of proteins and RNA through these EVs, yet the content and role of these EVs remain elusive. MicroRNAs (miRs) are central to adaptive responses to stress and more broadly to changes in culture conditions. Given that EVs are highly enriched in miRs, and that EVs release large quantities of miRs bothin vivoandin vitro, EVs and their miR content likely play an important role in adaptive responses. Here we report the miRNA landscape of CHO cells and their EVs under normal culture conditions and under ammonia and osmotic stress. We show that both cells and EVs are highly enriched in five miRs (among over 600 miRs) that make up about half of their total miR content, and that these highly enriched miRs differ significantly between normal and stress culture conditions. Notable is the high enrichment in mir-92a and miR-23a under normal culture conditions, in contrast to the high enrichment in let-7 family miRs (let-7c, let-7b and let-7a) under both stress conditions. The latter suggests a preserved stress-responsive function of the let-7 miR family, one of the most highly preserved miR families across species, where among other functions, let-7 miRs regulate core oncogenes, which, depending on the biological context, may tip the balance between cell cycle arrest and apoptosis. While the expected –based on their profound enrichment – important role of these highly enriched miRs remains to be dissected, our data and analysis constitute an important resource for exploring the role of miRs in cell adaptation as well as for synthetic applications.
Census of exposed aggregation-prone regions in proteomes
Authors: Falgarone Théo , Villain Etienne , Richard Francois , Osmanli Zarifa , Kajava Andrey V.
Published: December 2022

Loss of solubility usually leads to the detrimental elimination of protein function. In some cases, the protein aggregation is also required for beneficial functions. Given the duality of this phenomenon, it remains a fundamental question how natural selection controls the aggregation. The exponential growth of genomic sequence data and recent progress within silicopredictors of the aggregation allows approaching this problem by a large-scale bioinformatics analysis. Most of the aggregation-prone regions are hidden within the 3D structures and, therefore, they cannot realize their potential to aggregate. Thus, the most realistic census of the aggregation prone regions requires crossing aggregation prediction with information about the location of the natively unfolded regions. This allows us to detect so-called “Exposed Aggregation-prone Regions” (EARs). Here, we analyzed the occurrence and distribution of the EARs in 76 full reference proteomes from the three kingdoms of life. For this purpose, we used a bioinformatics pipeline, which provides a consensual result based on several predictors of aggregation. Our analysis revealed a number of new statistically significant correlations about the presence of EARs in different organisms, their dependence on protein length, cellular localizations, co-occurrence with short linear motifs, and the level of protein expression. We also obtained a list of proteins with the conserved aggregation-prone sequences for further experimental tests. Insights gained from this work led to a deeper understanding of the functional and evolutionary relations of the protein aggregation.
Drug-induced changes in connectivity to midbrain dopamine cells revealed by rabies monosynaptic tracing
Authors: Bartas Katrina , Hui May , Derdeyn Pieter , Tian Guilian , Vasquez Jose J. , Azouz Ghalia , Yamamoto Cindy M. , Beier Kevin T.
Published: December 2022

Drugs of abuse cause long-lasting changes in connectivity from inputs onto ventral tegmental area dopamine cells (VTADA) that contribute to drug-induced behavioral adaptations. However, it is not known which inputs are altered. Here we used a rabies virus-based mapping strategy to quantify rabies-labeled inputs to VTA cells after a single exposure to one of a variety of abused drugs – cocaine, amphetamine, methamphetamine, morphine, and nicotine – and compared the relative global input labeling across conditions. We observed that all tested drugs of abuse elicited similar input changes onto VTADAcells, in particular onto DA cells projecting to the lateral shell of the nucleus accumbens and amygdala. In addition, repeated administration of ketamine/xylazine to induce anesthesia induces a change in inputs to VTADAcells that is similar to but different from those elicited by a single exposure to drugs of abuse, suggesting that caution should be taken when using ketamine/xylazine-based anesthesia in rodents when assessing motivated behaviors. Furthermore, comparison of viral tracing data to an atlas of gene expression in the adult mouse brain showed that the basal expression patterns of several gene classes, especially calcium channels, were highly correlated with the extent of both addictive drug- or ketamine/xylazine-induced changes in rabies-labeled inputs to VTADAcells. Reducing expression levels of the voltage-gated calcium channelCacna1ein cells in the nucleus accumbens lateral shell reduced rabies-mediated input labeling of these cells into VTADAcells. These results directly link genes controlling cellular excitability and the extent of input labeling by the rabies virus.
Lung extracellular matrix modulates KRT5+basal cell activity in pulmonary fibrosis
Authors: Hewitt Richard J. , Puttur Franz , Gaboriau David C. A. , Fercoq Frédéric , Fresquet Maryline , Traves William J. , Yates Laura L. , Walker Simone A. , Molyneaux Philip L. , Kemp Samuel V. , Nicholson Andrew G. , Rice Alexandra , Lennon Rachel , Carlin Leo M. , Byrne Adam J. , Maher Toby M. , Lloyd Clare M.
Published: December 2022

Aberrant expansion of KRT5+basal cells in the distal lung accompanies progressive alveolar epithelial cell loss and tissue remodelling during fibrogenesis in idiopathic pulmonary fibrosis (IPF). The mechanisms determining activity of KRT5+cells in IPF have not been delineated. Here, we reveal a potential mechanism by which KRT5+cells migrate within the fibrotic lung, navigating regional differences in collagen topography.In vitro, KRT5+cell migratory characteristics and expression of remodelling genes are modulated by extracellular matrix (ECM) composition and organisation. Mass spectrometry-based proteomics revealed compositional differences in ECM components secreted by primary human lung fibroblasts (HLF) from IPF patients compared to controls. Over-expression of ECM glycoprotein, Secreted Protein Acidic and Cysteine Rich (SPARC) in the IPF HLF matrix restricts KRT5+cell migrationin vitro. Together, our findings demonstrate how changes to the ECM in IPF directly influence KRT5+cell behaviour and function contributing to remodelling events in the fibrotic niche.
A chromosome-scale high-contiguity genome assembly of the threatened cheetah (Acinonyx jubatus)
Authors: Winter Sven , Meißner René , Greve Carola , Hamadou Alexander Ben , Horin Petr , Prost Stefan , Burger Pamela A.
Published: December 2022

The cheetah (Acinonyx jubatus, SCHREBER 1775) is a large felid and is considered the fastest land animal. Historically, it inhabited open grassland across Africa, the Arabian Peninsula, and southwestern Asia; however, only small and fragmented populations remain today. Here, we present a de novo genome assembly of the cheetah based on PacBio continuous long reads and Hi-C proximity ligation data. The final assembly (VMU_Ajub_asm_v1.0) has a total length of 2.38 Gb, of which 99.7% are anchored into the expected 19 chromosome-scale scaffolds. The contig and scaffold N50 values of 96.8 Mb and 144.4 Mb, respectively, a BUSCO completeness of 95.4% and a k-mer completeness of 98.4%, emphasize the high quality of the assembly. Furthermore, annotation of the assembly identified 23,622 genes and a repeat content of 40.4%. This new highly contiguous and chromosome-scale assembly will greatly benefit conservation and evolutionary genomic analyses and will be a valuable resource, e.g., to gain a detailed understanding of the function and diversity of immune response genes in felids.
A computational model ofPseudomonas syringaemetabolism unveils the role of branched-chain amino acids in virulence expression at the early stages of Arabidopsis colonization
Authors: Tubergen Philip J. , Medlock Greg , Moore Anni , Zhang Xiaomu , Papin Jason A. , Danna Cristian H.
Published: December 2022

Leaf mesophyll-colonizing bacterial pathogens infect their plant hosts by adjusting their metabolism to the leaf mesophyll environment. Soon after the inoculation of naïve, susceptible plants, the model bacterial pathogenPseudomonas syringaepv.tomatoDC3000 (PstDC3000) expresses virulence factors that suppress plant immunity, a requirement to produce robust infections. However, if plant immunity was elicited with Microbe-Associated Molecular-Patterns (MAMPs) prior to bacterial inoculation,PstDC3000 slows down virulence expression and only produces symptomless mild infections. To understand how bacterial metabolism adapts to these two contrasting conditions, we created iPst19, anin silicoensemble of genome-scale metabolic reconstructions. Constraining thein silicogrowth of iPst19 within planta PstDC3000 gene expression data revealed that sugar catabolism is highly active in bacteria that have been inoculated in mock-treated plants. In contrast, branched-chain amino acids (BCAAs) catabolism is highly active in bacteria that have been inoculated in MAMP-pretreated plants. Bacterial growth and gene expression analysis showed that BCAAs suppress virulence gene expression without affecting bacterial growthin vitro. In planta, however, BCAAs suppress the expression of virulence genes at the early stages of the infection and significantly impair leaf colonization of the host plantArabidopsis thaliana. While the overexpression of the conserved bacterial leucine-responsive transcriptional regulatorLrpinduced the expression of virulence genes, its downregulation had the opposite effect, suggesting that BCAA-free Lrp induces virulence while BCAA-Lrpdoes not. Overall, our data provide mechanistic connections to understand how plant immunity impactsPstDC3000 metabolism and virulence, furthering our understanding of bacterial pathogenesis and plant disease.
A data assimilation method to track time-varying changes in the excitation-inhibition balance using scalp EEG
Authors: Yokoyama Hiroshi , Kitajo Keiichi
Published: December 2022

Recent neuroscience studies have suggested that controlling the excitation and inhibition (E/I) balance is essential for maintaining normal brain function. However, while control of time-varying E/I balance is considered essential for perceptual and motor learning, an efficient method for estimating E/I balance changes has yet to be established. To tackle this issue, we propose a new method to estimate E/I balance changes by applying neural-mass model-based tracking of the brain state using the Ensemble Kalman Filter. In this method, the parameters of synaptic E/I gains in the model are estimated from observed electroencephalography (EEG) signals. Moreover, the index of E/I balance was defined by calculating the ratio between synaptic E/I gains based on estimated parameters. The method was validated by showing that it could estimate E/I balance changes from human EEG data at the sub-second scale, indicating that it has the potential to quantify how time-varying changes in E/I balance influence changes in perceptual and motor learning. Furthermore, this method could be used to develop an E/I balance-based neurofeedback training method for clinical use.
LRRK2 Suppresses Lysosome Degradative Activity in Macrophages and Microglia Through MiT-TFE Transcription Factor Inhibition
Authors: Yadavalli Narayana , Ferguson Shawn M.
Published: December 2022

Cells maintain optimal levels of lysosome degradative activity to protect against pathogens, clear waste and generate nutrients. Here we show that LRRK2, a protein that is tightly linked to Parkinson’s disease, negatively regulates lysosome degradative activity in macrophages and microglia via a transcriptional mechanism. Depletion of LRRK2 and inhibition of LRRK2 kinase activity enhanced lysosomal proteolytic activity and increased the expression of multiple lysosomal hydrolases. Conversely, the kinase hyperactive LRRK2 G2019S Parkinson’s disease mutant suppressed lysosomal degradative activity and gene expression. We identified MiT-TFE transcription factors (TFE3, TFEB and MITF) as mediators of LRRK2-dependent control of lysosomal gene expression. LRRK2 negatively regulated the abundance and nuclear localization of these transcription factors and their depletion prevented LRRK2-dependent changes in lysosome protein levels. These discoveries define a role for LRRK2 in controlling lysosome degradative activity and support a model wherein LRRK2 hyperactivity may increase Parkinson’s disease risk by suppressing lysosome degradative activity.

Significance Statement

This study defines a homeostatic mechanism that allows macrophages and microglia to match the degradative activity of their lysosomes to ongoing changes in cellular demand. It shows that the leucine rich repeat kinase 2 (LRRK2) protein suppresses lysosome degradative activity by inhibiting the expression and nuclear localization of the MiT-TFE family of transcription factors that control the expression of multiple genes that encode lysosome proteins. It further demonstrates that a Parkinson’s disease mutation that hyperactivates LRRK2 kinase activity limits the degradative activity of lysosomes more strongly. These findings support a model wherein LRRK2 protects cells from excessive lysosome degradative activity and suggest that overactivation of this pathway may increase Parkinson’s disease risk by limiting the degradative activity of lysosomes.
Evaluation ofMycobacterium Tuberculosisenrichment in metagenomic samples using ONT adaptive sequencing and amplicon sequencing for identification and variant calling
Authors: Su Junhao , Lui Wui Wang , Lee YanLam , Zheng Zhenxian , Siu Gilman Kit-Hang , Ng Timothy Ting-Leung , Zhang Tong , Lam Tommy Tsan-Yuk , Lao Hiu-Yin , Yam Wing-Cheong , Tam Kingsley King-Gee , Leung Kenneth Siu-Sing , Lam Tak-Wah , Leung Amy Wing-Sze , Luo Ruibang
Published: December 2022

Sensitive detection ofMycobacterium Tuberculosis(TB) in small percentages in metagenomic samples is essential for microbial classification and drug resistance prediction, and assisting in diagnosis and treatment planning. However, traditional methods, such as bacterial culture and microscopy, are time-consuming and sometimes have limited TB detection sensitivity. Oxford Nanopore Technologies’ MinION sequencing allows rapid and simple sample preparation for whole genome and amplicon sequencing. Its recently developed adaptive sequencing selects reads from targets, while allowing real-time base-calling during sequencing to achieve sequence enrichment or depletion. Another common enrichment method is PCR amplification of the target TB genes. In this study, we compared both methods with MinION sequencing for TB detection and variant calling in metagenomic samples using both simulation runs and those with synthetic and patient samples. We found that both methods effectively enrich TB reads from a high percentage of human and other microbial DNA. We provide a simple automatic analysis framework, including quality filtering, taxonomic classification, variant calling, and antimicrobial resistance prediction, to support these detection workflows for clinical use, openly available athttps://github.com/HKU-BAL/ONT-TB-NF. Depending on the patient’s medical condition and sample type (commonly including bronchoalveolar lavage fluid, blood samples, sputum, and tissues), we discuss the findings and recommend that users optimize their workflow to improve the detection limit.
A subclass of the IS1202family of bacterial insertion sequences targets XerCD recombination sites
Authors: Siguier Patricia , Rousseau Philippe , Cornet François , Chandler Michael
Published: December 2022

IS1202, originally isolated fromStreptococcus pneumoniain the mid-1990s had been previously tagged as an emerging IS family in ISfinder. While searching for plasmid-associated Xer recombinase recombination sites (xrs) inAcinetobacter baumannii, we observed that some insertion sequences related to IS1202were repeatedly found abutting these sites in a number of plasmids. The plasmids often carried repeatedxrsthought to form a new type of mobile genetic element (MGE) which uses the chromosomally-encoded XerCD recombinase for mobility. The MGE (xrscassette) consist ofxrsflanking one or a small number of genes often including different clinically important carbapenemase-encodingbla-OXA. The IS1202-related IS are inserted with their left, transposase proximal extremity, IRL, five base pairs fromxrsand include a characteristic 5bp flanking target duplication. Further searches revealed that many different plasmid- and chromosome-bornexrscan be targeted and that IS1202-xrscombinations are not limited toAcinetobacter baumanniibut occur in other bacteria.

In addition to 28 IS1202group ISs in ISfinder and a number which had been subsequently submitted, we undertook a survey of the NCBI (February 2020) and identified 138 additional IS1202-related IS. These could be divided into 3 principal subgroups based on their transposase sequences and on the length of the DR generated on insertion: subgroup IS120227-28bp DR); ISTde1(15-17bp); and ISAba32(5-6bp). Members of each group which lacked DR were also found. But since other examples of most of these were subsequently identified having DR, those lacking DR may have been generated by intra-replicon recombination. Only members of the group which generate 5bp DR were found to targetxrs. These were not only identified in plasmids but also occurred at some individualxrssites,dif, located at the chromosome replication terminus and involved in post-replication chromosome segregation. Further analysis showed the presence of subgroup-specific indels in their transposases which may be responsible for the differences in their behavior.

We propose that this collection of IS be classed as a new insertion sequence family: the IS1202family composed of at three subfamilies, only one of which specifically targets plasmid-bornexrs. We discuss the implications ofxrstargeting for gene mobility.
The Neurophysiological Modality Effect in Native and Second Language Processing: An ERP Study
Authors: Gallagher Daniel , Yano Masataka , Ohta Shinri
Published: December 2022

Experimental psychology has long discussed the modality effect, whereby the stimulus modality significantly affects retention of the information presented. In neurolinguistics, however, the effect of stimulus modality on language processing has gained little attention. We conducted a multi-modal event-related potential (ERP) experiment on both native and non-native Spanish speakers to investigate the possibility of a neurophysiological modality effect in language processing. Using morphosyntactically violated and orthographically/phonologically violated stimuli, we elicited a robust N400 and P600 in native speakers. We showed that the N400 has consistent features across modalities, while the P600 has modality-specific features. Specifically, the auditory evoked P600 was characterized by a more gradual slope and a later peak than the visual evoked P600. We discuss this in detail along with other modality effects observed post sensory perception. Among second language (L2) learner groups, those with higher proficiency exhibited more nativelike neurophysiological responses in both modalities when compared to those with lower proficiency. We additionally observed fewer modality-specific differences in low proficiency learners than in higher proficiency learners suggesting that modality-specific specialization in language processing comes with increased proficiency. We further discuss the question of modality-specific differences in the process of neurophysiological nativization, whereby L2 learners’ ERPs become increasingly on nativelike.
Population size shapes the evolution of lifespan
Authors: Bagic Martin , Valenzano Dario Riccardo
Published: December 2022

Biological aging results from the age-dependent change in the force of natural selection, which increases the probability of germline variants that limit survival to accumulate in genes acting predominantly in late life1. The evolutionary mechanisms underlying the accumulation of neutral mutations and antagonistically pleiotropic gene variants that cause biological aging have been analyzed to date under the assumption of infinitely large population size. However, even though population size importantly shapes genetic and phenotype variation via drift and selection2,3, we still have a limited understanding of how finite population size impacts the evolution of mortality at the population level. Here, we study the impact of population size on lifespan evolution under mutation accumulation and antagonistic pleiotropy. We found that larger population size leads to lower age-independent, as well as age-dependent mortality under mutation accumulation, due to more effective purifying selection against deleterious germline variants. Strikingly, large population size can lead to extended lifespan under antagonistic pleiotropy, due to more effective positive selection on gene variants increasing survival in early-life, while leading to increased post-maturation mortality. Our findings provide a comprehensive numerical framework for the two major evolutionary genetic theories of aging and reveal a fundamental and yet non-appreciated role for population size in the evolution of mortality trajectories.
Transcriptional and translational dynamics underlying heat shock response in the thermophilic CrenarchaeonSulfolobus acidocaldarius
Authors: Baes Rani , Grünberger Felix , dit Ruys Sébastien Pyr , Couturier Mohea , De Keulenaer Sarah , Skevin Sonja , Van Nieuwerburgh Filip , Vertommen Didier , Grohmann Dina , Ferreira-Cerca Sébastien , Peeters Eveline
Published: December 2022

High-temperature stress is critical for all organisms and induces a profound cellular response. For Crenarchaeota, little information is available on how heat shock affects cellular processes and on how this response is regulated. In this work, we set out to study heat shock response in the thermoacidophilic model crenarchaeonSulfolobus acidocaldarius, which thrives in volcanic hot springs and has an optimal growth temperature of 75°C. Pulse-labeling experiments demonstrated that a temperature shift to 86°C induces a drastic reduction of the transcriptional and translational activity, but that RNA and protein neosynthesis still occurs. By combining RNA sequencing and TMT-labeled mass spectrometry, an integrated mapping of the transcriptome and proteome was performed. This revealed that heat shock causes an immediate change in the gene expression profile, with RNA levels of half of the genes being affected, followed by the more subtle reprogramming of the protein landscape. A limited correlation was observed in differential expression on the RNA and protein level, suggesting that there is a prevalence of post-transcriptional and post-translational regulation upon heat shock. Furthermore, based on the finding that promoter regions of heat shock regulon genes lack a conserved DNA-binding motif, we propose that heat-shock responsive transcription regulation is likely not to be accomplished by a classical transcription factor. Instead, in contrast to histone-harboring Euryarchaeota that have heat-shock transcription factors, it is hypothesized that Sulfolobales and other histone-lacking thermophilic archaea employ an evolutionary ancient mechanism relying on temperature-responsive changes in DNA organization and compaction, induced by the action of nucleoid-associated proteins.
The ins and outs of membrane bending by intrinsically disordered proteins
Authors: Yuan Feng , Lee Christopher T. , Sangani Arjun , Houser Justin R. , Wang Liping , Lafer Eileen M. , Rangamani Padmini , Stachowiak Jeanne C.
Published: December 2022

Membrane curvature is essential to diverse cellular functions. While classically attributed to structured domains, recent work illustrates that intrinsically disordered proteins are also potent drivers of membrane bending. Specifically, repulsive interactions among disordered domains drive convex bending, while attractive interactions, which lead to liquid-like condensates, drive concave bending. How might disordered domains that contain both repulsive and attractive domains impact curvature? Here we examine chimeras that combine attractive and repulsive interactions. When the attractive domain was closer to the membrane, its condensation amplified steric pressure among repulsive domains, leading to convex curvature. In contrast, when the repulsive domain was closer to the membrane, attractive interactions dominated, resulting in concave curvature. Further, a transition from convex to concave curvature occurred with increasing ionic strength, which reduced repulsion while enhancing condensation. In agreement with a simple mechanical model, these results illustrate a set of design rules for membrane bending by disordered proteins.
Excellent repigmentation was observed in the treatment of refractory vitiligo with autologous cultured epithelium grafting: a real-world retrospective cohort study
Authors: Li Jian , Zeng Xuanhao , Chen Shujun , Tang Luyan , Zhang Qi , Lv Minzi , Uyama Taro , Wu Fuyue , Lian Weiling , Wang Jinqi , Lv Haozhen , Liu Yating , Wu Jinfeng , Xu Jinhua
Published: December 2022

Surgical intervention is considered as the mainstream therapy for refractory vitiligo. In this study, we developed a modified autologous cultured epithelial grafting (ACEG) technique for the surgical treatment of vitiligo. A total of 726 patients with vitiligo treated with ACEG were enrolled from January 2015 to June 2019 in China. Patient characteristics, such as sex, age, clinical type, lesion sites, course of the disease, and disease stable period, were recorded. In 2118 skin lesions from 726 patients who received ACEG, total efficacy rate was 82.81% (1754/2118).

However, the repigmentation rate of the ACEG was 64.87%, which was higher than that of conventional surgical interventions (52.69%). Patients with segmental vitiligo, skin lesions in the lower limbs, aged 18 years or below, and a stable period of over 3 years might have a good response to ACEG. Single-cell RNA sequencing was performed to observe different cell compositions in the skin before and after ACEG. The number of melanocytes increased by 50% after transplantation. In addition, there was a significant increase in hair follicle outer root sheath-derived keratinocytes in ACEG, and the numbers of these cells in the repigmentation sites 1 year after ACEG were still higher than those in the skin lesions. Therefore, ACEG is a promising therapeutic agent for refractory vitiligo. Age, clinical type, lesion site, and lesion stable period before surgery have significant impacts on repigmentation in ACEG. ACEG can increase the number of melanocytes and KRT6C+ keratinocytes in skin lesions, thereby restoring a skin microenvironment suitable for melanocyte survival.

One sentence summary

Autologous cultured epithelial grafting (ACEG) technique is a promising therapy for refractory vitiligo.