bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of kinesthetic cues supported by physiotherapist during a motor training intervention with virtual reality-based games on functioning in people with Parkinson’s disease: A prospective, single-blinded, parallel-group, randomized clinical trial
Authors: Igarasi Barbosa Pamela Yuki , Falconi Amarilis , D’Alencar Matheus , Okamoto Erika , Pimentel Piemonte Maria Elisa
Published: September 2023

Background

It’s been suggested that kinesthetic cues (KC) could be a useful tool in helping individuals improve their motor learning process by facilitating muscle activation. However, while virtual reality-based games (VRG) are becoming more popular as a intervention tool, the potential effects of KC on this kind of intervention have yet to be investigated in people with Parkinson’s disease (PwPD). Therefore, this study aimed to compare the effects of motor intervention using VRG coupled or not with KC provided by a physiotherapist (PT) during training on the functioning of PwPD.

Methods

Thirty-eight PwPD in 1-3 Hoehn and Yahr (HY) stage were randomized into two groups: (1) VRG with KC Group (KCG), where KC was provided by manual assistance of a PT, and (2) VRG with NO KC group (NKCG), where no PT manual assistance was provided during the training. Both groups received 8 individual sessions with 50 minutes, twice a week, for 4 weeks: 10 minutes for warm-up and 40 minutes to play 4 games fromXBOX 360withKinect®system. Outcomes were evaluated at three time points: (1) before training (BT), (2) 1 week after training (AT), and 8 weeks after that as follow-up (FU). To reach a comprehensive evaluation of functioning, several outcomes were adopted and categorized according to International Classification of Functioning Disability and Health (ICF): Geriatric Depression Scale (GDS); Montreal Cognitive Assessment (MOCA); Unified Parkinson Disease Rating Scale (UPDRS) - section 3 (UPDRS-III); and Rapid Turns Test (RTT) used to assess motor and non-motor alteration into Function domain; Balance Evaluation Systems Test (BESTest); Falls Efficacy Scale International (FES-I); Thirty-Second Walk Test (30sWT); Six-Minute Walk Test (6mWT); 5 Times Sit to Stand Test; and UPDRS section II (UPDRS-II) used to assess motor performance into Activity domain, and finally, the Parkinson Disease Questionnaire (PDQ-39) to assess quality of life into Participation domain.

Results

ANOVA for repeated measures showed a significant effect for evaluation-time factor only (p-value<.001), (no effect for group or evaluation-time X group interaction) for all measures of three ICF domains, excluding GDS and 6mWT. The Tukey post-hoc test confirmed significant improvements in AT that remained at FU. Conclusions: A motor intervention using VRG can improve the functioning in terms of Function, Activity, and Participation of PwPD, regardless of the KC provided by PT during the training.
Insights intoPhakopsora pachyrhizieffector-effector interactions
Authors: Qi Mingsheng , Yu Haiyue , Bredow Melissa , Chicowski Aline Sartor , Fields Letícia Dias , Whitham Steven A.
Published: September 2023

The multifaceted role of pathogen-encoded effectors in plant-pathogen interactions is complex and not fully understood. Effectors operate within intricate host environments, interacting with host proteins and other effectors to modulate virulence. The complex interplay between effectors raises the concept of metaeffectors, where some effectors regulate the activity of others. While previous research has demonstrated the importance of effector repertoires in pathogen virulence, only a limited number of studies have investigated the interactions between these effectors. This study explores the interactions amongPhakopsora pachyrhizieffector candidates (PpECs).P. pachyrhizihaustorial transcriptome analysis identified a collection of predictedPpECs. Among these,PpEC23 was found to interact withPpEC48, prompting further exploration into their potential interaction with other effectors. Here, we utilized a yeast-two-hybrid screen to explore protein-protein interactions betweenPpECs. A split-luciferase complementation assay also demonstrated that these interactions could occur within soybean cells. Interestingly,PpEC48 displayed the ability to interact with several small cysteine-rich proteins (SCRP), suggesting its affinity for this specific class of effectors. We show that these interactions involve a histidine-rich domain withinPpEC48, emphasizing the significance of structural motifs in mediating effector interactions. The unique nature ofPpEC48, showing no sequence matches in other organisms, suggests its relatively recent evolution and potential orphan gene status. Our work reveals insights into the intricate network of interactions amongP. pachyrhizieffector-effector interactions.
Accurate Prediction of Antibody Function and Structure Using Bio-Inspired Antibody Language Model
Authors: Jing Hongtai , Gao Zhengtao , Xu Sheng , Shen Tao , Peng Zhangzhi , He Shwai , You Tao , Ye Shuang , Lin Wei , Sun Siqi
Published: September 2023

In recent decades, antibodies have emerged as indispensable therapeutics for combating diseases, particularly viral infections. However, their development has been hindered by limited structural information and labor-intensive engineering processes. Fortunately, significant advancements in deep learning methods have facilitated the precise prediction of protein structure and function by leveraging co-evolution information from homologous proteins. Despite these advances, predicting the conformation of antibodies remains challenging due to their unique evolution and the high flexibility of their antigen-binding regions. Here, to address this challenge, we present the Bio-inspired Anti-body Language Model (BALM). This model is trained on a vast dataset comprising 336 million 40% non-redundant unlabeled antibody sequences, capturing both unique and conserved properties specific to antibodies. Notably, BALM showcases exceptional performance across four antigen-binding prediction tasks. Moreover, we introduce BALMFold, an end-to-end method derived from BALM, capable of swiftly predicting full atomic antibody structures from individual sequences. Remarkably, BALMFold outperforms those well-established methods like AlphaFold2, IgFold, ESMFold, and OmegaFold in the antibody benchmark, demonstrating significant potential to advance innovative engineering and streamline therapeutic antibody development by reducing the need for unnecessary trials.
Sexual antagonism and sex determination in three syngnathid species alongside a male pregnancy gradient
Authors: Dubin Arseny , Parker Jamie , Böhne Astrid , Roth Olivia
Published: September 2023

The allocation of energy towards gamete production, parental care, mate choice, sex roles, and sexual dimorphism generates divergence in selection pressures between the sexes, leading to opposing fitness strategies and sexual antagonism (SA). Due to the shared genetic makeup, a single genomic locus can contain a gene or allele with differing fitness impacts on each sex. This intralocus sexual conflict can be resolved via intersex bias in gene expression and/or formation of sex-linked genomic regions, that may also regulate sex determination. Sex determination (SD) encompasses environmental SD (ESD), monogenic SD, and polygenic SD. Occasionally, shifts from one SD locus to another can occur. While the precise mechanisms driving these shifts are unknown, SA is believed to be a major contributor. To investigate the link between SA and SD, we selected three syngnathid species along the gradient of male pregnancy that evolved with different sex roles and intensities of sexual dimorphism. By looking at intersex genetic divergence (Fst) and sex-biased expression patterns, we uncovered that sex role and mate competition, rather than male pregnancy, primarily drive SA. Furthermore, we identified processes related to non-coding RNAs and biased allele expression as mediators of SA. Most notably, we discovered intraspecies sex chromosome polymorphism inHippocampus erectus. Overall, we report important details on the interplay between SA and SD, and suggest that understanding SA and its resolution mechanisms is crucial for unraveling the evolution of SD in diverse species.
Explainable machine learning for the identification of proteome states via the data processing kitchen sink
Authors: Scott Aaron M. , Hartman Erik , Malmström Johan , Malmström Lars
Published: September 2023

The application of machine learning algorithms to facilitate the understanding of changes in proteome states has emerged as a promising methodology in proteomics research. Unfortunately, these methods can prove difficult to interpret, as it may not be immediately obvious how models reach their predictions. We present the data processing kitchen sink (DPKS) which provides reproducible access to classic statistical methods and advanced explainable machine learning algorithms to build highly accurate and fully interpretable predictive models. In DPKS, explainable machine learning methods are used to calculate the importance of each protein towards the prediction of a model for a particular proteome state. The calculated importance of each protein can enable the identification of proteins that drive phenotypic change in a data-driven manner while classic techniques rely on arbitrary cutoffs that may exclude important features from consideration. DPKS is a free and open source Python package available athttps://github.com/InfectionMedicineProteomics/DPKS.
Rapid translocation of NGR proteins drives polarization of PIN-activating D6 protein kinase during root gravitropism
Authors: Kulich Ivan , Schmid Julia , Teplova Anastasia , Qi Linlin , Friml Jiří
Published: September 2023

Root gravitropic bending represents a fundamental aspect of terrestrial plant physiology. Gravity is perceived by sedimentation of starch-rich plastids (statoliths) to the bottom of the central root cap cells. Following gravity perception, intercellular auxin transport is redirected downwards leading to an asymmetric auxin accumulation at the lower root side causing inhibition of cell expansion, ultimately resulting in downwards bending. How gravity-induced statoliths repositioning is translated into asymmetric auxin distribution remains unclear despite PIN auxin efflux carriers and the Negative Gravitropic Response of roots (NGR) proteins polarize along statolith sedimentation, thus providing a plausible mechanism for auxin flow redirection. In this study, using a functional NGR1-GFP construct, we visualized the NGR1 localization on the statolith surface and plasma membrane (PM) domains in close proximity to the statoliths, correlating with their movements. We determined that NGR1 binding to these PM domains is indispensable for NGR1 functionality and relies on cysteine acylation and adjacent polybasic regions as well as on lipid and sterol PM composition. Detailed timing of the early events following graviperception suggested that both NGR1 repolarization and initial auxin asymmetry precede the visible PIN3 polarization. This discrepancy motivated us to unveil a rapid, NGR-dependent translocation of PIN-activating AGCVIII kinase D6PK towards lower PMs of gravity-perceiving cells, thus providing an attractive model for rapid redirection of auxin fluxes following gravistimulation.
Development and validation of a respiratory syncytial virus multiplex immunoassay
Authors: Marsall Patrick , Fandrich Madeleine , Griesbaum Johanna , Harries Manuela , Lange Berit ,   RESPINOW study consortium, Ascough Stephanie , Dayananda Pete , Chiu Christopher , Remppis Jonathan , Ganzenmueller Tina , Renk Hanna , Strengert Monika , Schneiderhan-Marra Nicole , Dulovic Alex
Published: September 2023

Respiratory syncytial virus (RSV) is one of the leading causes of severe respiratory disease in infants and adults. RSV exists as two subtypes A and B, which co-circulate throughout the season, although one will usually become dominant. While vaccines and monoclonal therapeutic antibodies either are or will shortly become available, correlates of protection remain unclear. For this purpose, we developed an RSV multiplex immunoassay that analyses antibody titers towards the post-F, Nucleoprotein, and a diverse mix of G proteins. Technical and clinical validation showed outstanding performance, while methodological developments enabled identification of the subtype of previous infections through use of the diverse G proteins for approximately 50% of samples. As a proof of concept to show the suitability of the assay in serosurveillance studies, we then evaluated titer decay and age- dependent antibody responses within population cohorts. Overall, the developed assay shows robust performance, is scalable, provides additional information on infection subtype, and is therefore ideally suited to be used in future population cohort studies.

Importance

Although respiratory syncytial virus (RSV) is endemic and re-infections are common and harmless to the majority of the population, it is a leading cause of hospitalization in young children, the elderly, or immunocompromised individuals. A better characterization of RSV immunology and spreading dynamics is thus critical for preparedness, especially when interventions aiming to mitigate other diseases (e.g., COVID-19) disturb its endemic cycles. This requires high-throughput information-dense assays. We therefore developed a bead-based multiplex immunoassay that allows measurements of antibodies against multiple RSV antigens simultaneously. We identified antibodies which were strong indicators of previous infection, while others allowed identification of the subtype of the previous infection. The assay itself was shown to be robust and scalable, making it ideal for to keep track of the temporal variation RSV immunity profiles within the population.
The olfactory-based neurofeedback of the EEG alpha rhythm
Authors: Medvedeva Alexandra , Ninenko Ivan , Kleeva Daria F , Fedoseev Alexey , Bazhenov Artem , Cabrera Miguel Altamirano , Tsetserukou Dzmitry , Lebedev Mikhail A
Published: September 2023

Neurofeedback (NFB) is a form of biofeedback that enables subjects to monitor and control their own brain activity. To communicate with the subject, modern NFB training methodologies utilize various signaling mechanisms, typically visual and/or auditory stimuli. Olfaction has not been explored yet as a way to deliver NFB. Here we developed an olfactory-based NFB system based on electroencephalographic (EEG) recordings in human participants. The system incorporates an EEG recording apparatus, a custom olfactory display for the automated delivery of Sniffin’ Sticks, and a Python application for the conversion of EEG rhythms into NFB signals and controlling behavioral tasks. We tested occipital alpha rhythm as the source of NFB. Fifteen healthy participants were randomly assigned to three groups: olfactory neurofeedback, auditory NFB, and mock-olfactory NFB. NFB training resulted in an increase of alpha power in the true NFB groups, but not in the mock NFB group where the alpha power decreased, probably because of fatigue and drowsiness. Based on these results, we conclude that olfactory NFB is feasible, and lay out a framework for its future development.
Genome-wide CRISPRi screens reveal the essentialome and determinants for susceptibility to dalbavancin inStaphylococcus aureus
Authors: Liu Xue , de Bakker Vincent , Heggenhougen Maria Victoria , Mårli Marita Torrissen , Frøynes Anette Heidal , Salehian Zhian , Porcellato Davide , Angeles Danae Morales , Veening Jan-Willem , Kjos Morten
Published: September 2023

Antibiotic resistance and tolerance remain a major problem for treatment of staphylococcal infections. Knowing genes that influence antibiotic susceptibility could open the door to novel antimicrobial strategies, including targets for new synergistic drug combinations. Here, we developed a genome-wide CRISPR interference library forStaphylococcus aureus, demonstrated its use by quantifying the essentialome in different strains through CRISPRi-seq, and used it to identify genes that modulate susceptibility to the lipoglycopeptide dalbavancin. By exposing the library to sublethal concentrations of dalbavancin using both CRISPRi-seq and direct selection methods, we found genes previously reported to be involved in antibiotic susceptibility, but also identified genes thus far unknown to affect antibiotic tolerance. Importantly, some of these genes could not have been detected by more conventional knock-out approaches because they are essential for growth, stressing the complementary value of CRISPRi-based methods. Notably, knockdown of a gene encoding the uncharacterized protein KapB specifically sensitizes the cells to dalbavancin, but not to other antibiotics of the same class, while knockdown of the Shikimate pathway surprisingly has the opposite effect. The results presented here demonstrate the potential of CRISPRi-seq screens to identify genes and pathways involved in antibiotic susceptibility and pave the way to explore alternative antimicrobial treatments through these insights.
VCP increases or decreases tau seeding using specific cofactors
Authors: Batra Sushobhna , Vaquer-Alicea Jaime , Manon Victor A. , Kashmer Omar M. , Lemoff Andrew , Cairns Nigel J. , White Charles L. , Diamond Marc I.
Published: September 2023

Background

Neurodegenerative tauopathies may progress based on seeding by pathological tau assemblies, whereby an aggregate is released from one cell, gains entry to an adjacent or connected cell, and serves as a specific template for its own replication in the cytoplasm.In vitroseeding reactions typically take days, yet seeding into the complex cytoplasmic milieu can happen within hours. A cellular machinery might regulate this process, but potential players are unknown.

Methods

We used proximity labeling to identify factors that control seed amplification. We fused split-APEX2 to the C-terminus of tau repeat domain (RD) to reconstitute peroxidase activity upon seeded intracellular tau aggregation. We identified valosin containing protein (VCP/p97) 5h after seeding. Mutations in VCP underlie two neurodegenerative diseases, multisystem proteinopathy and vacuolar tauopathy, but its mechanistic role is unclear. We utilized tau biosensors, a cellular model for tau aggregation, to study the effects of VCP on tau seeding.

Results

VCP knockdown reduced tau seeding. However, distinct chemical inhibitors of VCP and the proteasome had opposing effects on aggregation, but only when given <8h of seed exposure. ML-240 increased seeding efficiency ∼40x, whereas NMS-873 decreased seeding efficiency by 50%, and MG132 increased seeding ∼10x. We screened VCP co-factors in HEK293 biosensor cells by genetic knockout or knockdown. Reduction of ATXN3, NSFL1C, UBE4B, NGLY1, and OTUB1 decreased tau seeding, as did NPLOC4, which also uniquely increased soluble tau levels. Reduction of FAF2 and UBXN6 increased tau seeding.

Conclusions

VCP uses distinct cofactors to determine seed replication efficiency, consistent with a dedicated cytoplasmic processing complex that directs seeds towards dissolution vs. amplification.
Genome-wide screens reveal shared and strain-specific genes that facilitate enteric colonization byKlebsiella pneumoniae
Authors: Cheung Bettina H , Alisoltani Arghavan , Kochan Travis J , Lebrun-Corbin Marine , Nozick Sophia H , Axline Christopher MR , Bachta Kelly ER , Ozer Egon A , Hauser Alan R
Published: September 2023

Gastrointestinal (GI) colonization byKlebsiella pneumoniaeis a risk factor for subsequent infection as well as transmission to other patients. Additionally, colonization is achieved by many strain types that exhibit high diversity in genetic content. Thus, we aimed to study strain-specific requirements forK. pneumoniaeGI colonization by applying transposon insertion sequencing to three classical clinical strains: a carbapenem-resistant strain, an extended-spectrum beta-lactamase producing strain, and a non-epidemic antibiotic-susceptible strain. The transposon insertion libraries were screened in a murine model of GI colonization. At three days post-inoculation, 27 genes were required by all three strains for colonization. Isogenic deletion mutants for three genes/operons (acrA,carAB,tatABCD) confirmed colonization defects in each of the three strains. Additionally, deletion ofacrAreduced bile tolerancein vitro, while complementation restored both bile tolerancein vitroand colonization abilityin vivo. Transposon insertion sequencing suggested that some genes were more important for colonization of one strain than the others. For example, deletion of the sucrose porin-encoding genescrYresulted in a colonization defect in the carbapenemase-producing strain but not in the extended-spectrum beta-lactamase producer or the antibiotic-susceptible strain. These findings demonstrate that classicalK. pneumoniaestrains use both shared and strain-specific strategies to colonize the mouse GI tract.

IMPORTANCE

Klebsiella pneumoniaeis a common cause of difficult-to-treat infections due to its propensity to express resistance to many antibiotics. For example, carbapenem-resistantK. pneumoniae(CR-Kp) has been named an urgent threat by the United States Centers for Disease Control and Prevention. Gastrointestinal colonization of patients withK. pneumoniaehas been linked to subsequent infection, making it a key process to control in prevention of multidrug-resistant infections. However, the bacterial factors which contribute toK. pneumoniaecolonization are not well understood. Additionally, individual strains exhibit large amounts of genetic diversity, begging the question of whether some colonization factors are strain-dependent. This study identifies the enteric colonization factors of 3 classical strains using transposon mutant screens to define a core colonization program forK. pneumoniaeas well as detecting strain-to-strain differences in colonization strategies.
Ras-mediated homeostatic control of front-back signaling dictates cell polarity
Authors: Lin Yiyan , Pal Dhiman Sankar , Banerjee Parijat , Banerjee Tatsat , Qin Guanghui , Deng Yu , Borleis Jane , Iglesias Pablo A. , Devreotes Peter N.
Published: September 2023

Studies in the model systems,Dictyosteliumamoebae and HL-60 neutrophils, have shown that local Ras activity directly regulates cell motility or polarity. Localized Ras activation on the membrane is spatiotemporally regulated by its activators, RasGEFs, and inhibitors, RasGAPs, which might be expected to create a stable ‘front’ and ‘back’, respectively, in migrating cells. Focusing on C2GAPB in amoebae and RASAL3 in neutrophils, we investigated how Ras activity along the cortex controls polarity. Since existing gene knockout and overexpression studies can be circumvented, we chose optogenetic approaches to assess the immediate, local effects of these Ras regulators on the cell cortex. In both cellular systems, optically targeting the respective RasGAPs to the cell front extinguished existing protrusions and changed the direction of migration, as might be expected. However, when the expression of C2GAPB was induced globally, amoebae polarized within hours. Furthermore, within minutes of globally recruiting either C2GAPB in amoebae or RASAL3 in neutrophils, each cell type polarized and moved more rapidly. Targeting the RasGAPs to the cell backs exaggerated these effects on migration and polarity. Overall, in both cell types, RasGAP-mediated polarization was brought about by increased actomyosin contractility at the back and sustained, localized F-actin polymerization at the front. These experimental results were accurately captured by computational simulations in which Ras levels control front and back feedback loops. The discovery that context-dependent Ras activity on the cell cortex has counterintuitive, unanticipated effects on cell polarity can have important implications for future drug-design strategies targeting oncogenic Ras.
Effect of cannabigerol on sleep and quality of life in Veterans: A decentralized, randomized, placebo-controlled trial
Authors: Emerson Chris R , Webster Courtney E , Daza Eric J , Klamer Brett G , Tummalacherla Meghasyam
Published: September 2023

Background/Objective

This decentralized, randomized, triple-blind, placebo-controlled study evaluated the efficacy and safety of an oral cannabigerol (CBG) formulation in Veterans with sleep issues.

Methods

After inclusion, randomization and a two-week run-in phase, participants received CBG (25 mg daily for two weeks, escalated to 50 mg daily for a further two weeks) or placebo. The primary endpoint was change in sleep quality, assessed via the Medical Outcomes Study Sleep Problems Index II (MOS-SS SPI-II). Additional endpoints included change in quality of life (WHODAS-2.0–12), post-traumatic stress disorder symptoms (PCL-5) and sleep actigraphy data. Safety was assessed based on adverse event reporting.

Results

A total of 63 participants were randomized to receive CBG (n=33) or placebo (n=30) and formed the intention-to-treat (ITT) population. Of these, 35 completed the study without major protocol deviations (CBG [n=18]; placebo [n=17]) and formed the per-protocol (PP) population. During active treatment (between day 14 and day 42) MOS-SS SPI-II scores declined numerically (indicating improved sleep) in both treatment groups (in both the ITT and the PP populations) with no statistically discernible difference between the CBG and placebo groups. Similar patterns were observed for WHODAS-2.0–12 and PCL-5 scores. Actigraphy data indicated no discernible difference in sleep patterns between the treatment groups. Five mild, nonserious, adverse events were reported with CBG.

Conclusions

Both CBG and placebo tended towards sleep and QoL improvement in Veterans. While no firm conclusion on the efficacy of CBG in improving sleep can be made, the favorable safety profile supports future studies to investigate the benefit of CBG.
Clinical evaluation of deep learning-based risk profiling in breast cancer histopathology and comparison to an established multigene assay
Authors: Wang Yinxi , Sun Wenwen , Karlsson Emelie , Lövgren Sandy Kang , Ács Balázs , Rantalainen Mattias , Robertson Stephanie , Hartman Johan
Published: September 2023

A significant proportion of oestrogen receptor (ER)-positive and human epidermal growth factor receptor 2 (HER2)-negative early breast cancer patients are categorised as intermediate risk based on classic clinicopathological variables, thus providing limited information to guide treatment decisions. The Prosigna assay is one of the established prognostic multigene assays in clinical practice for risk profiling. Stratipath Breast is a novel deep learning-based image analysis tool that utilises haematoxylin and eosin (HE)-stained histopathological images for risk profiling. In this study, we aimed to evaluate the Stratipath Breast tool for image-based risk profiling and compare it with the Prosigna assay. In a real-world breast cancer case series comprising 234 invasive tumours from patients with early ER+/HER2-breast cancer, clinically intermediate risk and eligible for chemotherapy, clinicopathological data including Prosigna results and corresponding HE-stained tissue slides were retrieved. The digitised HE slides were analysed by Stratipath Breast. Our findings showed that the Stratipath Breast analysis identified 49.6% of the clinically intermediate tumours as low risk and 50.4% as high risk. The Prosigna assay classified 32.5%, 47.0% and 20.5% tumours as low, intermediate and high risk, respectively. Among Prosigna intermediate-risk tumours, 47.3% were stratified as Stratipath low risk and 52.7% as high risk. In addition, 89.7% of Stratipath low-risk cases were classified as Prosigna low/intermediate risk. The overall agreement between the two tests for low-risk and high-risk groups was 71.0%, with a Cohen’s kappa of 0.42. For both risk profiling tests, grade and Ki67 differed significantly between risk groups. In conclusion, for the first time, we here present the results from a clinical evaluation of image-based risk stratification and show a considerable agreement to an established gene expression assay in routine breast pathology. The findings demonstrate that image-based risk profiling may aid in the identification of low-risk patients who could potentially be spared adjuvant chemotherapy.
Antiviral activity of intracellular nanobodies targeting the influenza virus RNA-polymerase core
Authors: Bessonne Mélissa , Morel Jessica , Nevers Quentin , Ballandras-Colas Allison , Grange Magali , Roussel Alain , Crépin Thibaut , Delmas Bernard
Published: September 2023

Influenza viruses transcribe and replicate their genome in the nucleus of the infected cells, two functions that are supported by the viral RNA-dependent RNA-polymerase (FluPol). FluPol displays structural flexibility related to distinct functional states, from an inactive form to conformations competent for replication and transcription. FluPol machinery is constituted by a structurally-invariant core comprising the PB1 subunit stabilized with PA and PB2 domains, whereas the PA endonuclease and PB2 C-domains can pack in different configurations around the core. To get insights into the functioning of FluPol, we selected single-domain nanobodies (VHHs) specific of the influenza A FluPol core. When expressed intracellularly, several of them exhibited inhibitory activity on type A FluPol, but not on the type B one. The most potent VHH (VHH16) targets PA, but preferentially bind the PA-PB1 dimer with an affinity below the nanomolar range. Ectopic intracellular expression of VHH16 in virus permissive cells blocks multiplication of different influenza A subtypes, even when induced at late times post-infection. VHH16 was found to impair the transport of the PA-PB1 dimer to the nucleus, without affecting its handling by the importin β RanBP5 and subsequent steps in FluPol assembly. These data suggest that the VHH16 neutralization activity is likely due to an alteration of the import of the PA-PB1 dimer into the nucleus, resulting to an inhibition of FluPol functioning. VHH16 binding site represent a potential target for antiviral development.

Author Summary

The influenza virus RNA-polymerase (FluPol) ensures genome transcription and replication in the nucleus of the infected cells. To select ligands able to block FluPol activities, we screened a library of phages encoding nanobodies and resulting from the immunization of a llama with FluPol subunits. When expressed intracellularly, one of the nanobodies displays highly efficient FluPol blocking and virus neutralizing activities. This nanobody binds FluPol with high affinity and recognizes preferentially the PA-PB1 assembled subunits. Furthermore, it was found to interfere with the transport of the PA-PB1 dimer into the nucleus, suggesting that targeting FluPol trafficking between the cytoplasm and the nucleus may constitute a powerful strategy to develop new antivirals.
The Host miR-17-92 Cluster Negatively Regulates Mouse Mammary Tumor Virus (MMTV) Replication via miR-92a by Targeting the Viral Genome
Authors: Baby Jasmin , Gull Bushra , Ahmad Waqar , Khader Thanumol Abdul , Panicker Neena G. , Akhlaq Shaima , Rizvi Tahir A. , Mustafa Farah
Published: September 2023

The mouse mammary tumor virus (MMTV) induces mammary tumors in mice. Recently, we have shown that MMTV perturbs expression of the host miR-17-92 cluster in MMTV-infected mammary glands and MMTV-induced mammary tumors. Known as oncomiR-1, this cluster is often dysregulated in a number of cancers, especially breast cancer. Therefore, we investigated whether there was a functional interaction between MMTV and the cluster. Our results reveal that MMTV expression led to dysregulation of the cluster in both mammary epithelial HC11 and HEK293T cells with the expression of miR-92a cluster member being affected the most. Conversely, over-expression of the whole or partial cluster significantly repressed MMTV expression. Interestingly, overexpression of miR-92a alone resulted in MMTV repression to the same extent as observed with the whole or partial cluster. Inhibition of miR-92a led to nearly a complete recovery of MMTV gene expression. Deletion/substitution of the miR-92a seed sequence within the cluster rescued MMTV expression. Dual luciferase assays identified MMTV genomic RNA as the target of miR-92a. These results show that the miR-17-92 cluster acts as part of the cell’s well-known miRNA-based anti-viral response to thwart incoming MMTV infection. Interestingly, miR-92a expression was the least amongst cluster members in MMTV-induced breast tumors, suggesting that the elevated level of MMTV expression in tumors is due perhaps to MMTV subverting miR-92a overexpression, restricting its ability to control MMTV replication during tumorigenesis. Thus, our study provides the first evidence demonstrating the biological significance of host miRNAs in controlling MMTV replication and influencing the process of tumorigenesis.

Importance

MMTV is a non-acute, slow-transforming retrovirus that causes breast cancer and lymphomas in mice. Consequently, understanding how MMTV interacts with its host can reveal critical aspects of cellular anti-viral responses and the process of tumorigenesis. This study reveals that the host miR-17-92 cluster plays an anti-viral role during MMTV infection which seems to be counteracted by MMTV during tumorigenesis. This observation is significant since MMTV-induced tumors share similarity with human breast cancer and the MMTV/mouse model is the best mammalian animal model to study breast cancer initiation, progression, and therapy. Moreover, MMTV is increasingly (though controversial) being implicated in human breast cancer, leukemias/lymphomas, and biliary cirrhosis due to its potential zoonosis into humans. Thus, if proven true, understanding how miRNAs modulate MMTV replication will be critical in appreciating how hosts miRNAs affect virus replication/tumorigenesis, and towards the development of novel miRNA-based therapeutics (e.g., anti-miRs) to thwart MMTV replication in humans.
Structural basis of aggregate binding by the AAA+ disaggregase ClpG
Authors: Katikaridis Panagiotis , Simon Bernd , Jenne Timo , Moon Seongjoon , Lee Changhan , Hennig Janosch , Mogk Axel
Published: September 2023

Severe heat stress causes massive loss of essential proteins by aggregation necessitating a cellular activity that rescues aggregated proteins. This activity is executed by ATP-dependent, ring-forming, hexameric AAA+ disaggregases. Little is known about the recognition principles of stress-induced protein aggregates. How can disaggregases specifically target aggregated proteins while avoiding binding to soluble non-native proteins? Here, we determined by NMR spectroscopy the core structure of the aggregate targeting N1 domain of the bacterial AAA+ disaggregase ClpG, which confers extreme heat resistance to bacteria. N1 harbors a Zn2+-coordination site that is crucial for structural integrity and disaggregase functionality. Conserved hydrophobic N1 residues located on a β-strand were found crucial for aggregate targeting and disaggregation activity. Mixing experiments with N1-truncated AAA+ hexamers revealed that a minimal number of four N1 domains must be present in a AAA+ ring for high disaggregation activity. We suggest that multiple N1 domains increase substrate affinity through avidity effects. These findings define the recognition principle of a protein aggregate by a disaggregase, involving simultaneous contacts with multiple hydrophobic substrate patches located in close vicinity on an aggregate surface. This binding mode ensures selectivity for aggregated proteins while sparing soluble, non-native protein structures from disaggregase activity.
The crystal structure of bacteriophage λ RexA provides novel insights into the DNA binding properties of Rex-like phage exclusion proteins
Authors: Adams Myfanwy C. , Schiltz Carl J. , Sun Jing , Hosford Christopher J. , Johnson Virginia M. , Pan Hao , Borbat Peter P. , Freed Jack H. , Thomason Lynn C. , Court Carolyn , Court Donald L. , Chappie Joshua S.
Published: September 2023

RexA and RexB function as an exclusion system that prevents bacteriophage T4rIImutants from growing onE. coliλ phage lysogens. Recent data established that RexA is a non-specific DNA binding protein that can act independently of RexB to bias the λ bistable switch toward the lytic state, preventing conversion back to lysogeny. The molecular interactions underlying these activities are unknown, owing in part to a dearth of structural information. Here we present the 2.05-Å crystal structure of the λ RexA dimer, which reveals a two-domain architecture with unexpected structural homology to the recombination-associated protein RdgC. Modelling suggests that our structure adopts a closed conformation and would require significant domain rearrangements to facilitate DNA binding. Mutagenesis coupled with electromobility shift assays, limited proteolysis, and double electron-electron spin resonance spectroscopy support a DNA-dependent conformational change. In vivophenotypes of RexA mutants suggest that DNA binding is not a strict requirement for phage exclusion but may directly contribute to modulation of the bistable switch. We further demonstrate that RexA homologs from other temperate phages also dimerize and bind DNAin vitro. Collectively, these findings advance our mechanistic understanding of Rex functions and provide new evolutionary insights into different aspects of phage biology.
Strain-specific differences in the interactions of the cucumber mosaic virus 2b protein with the viral 1a and host Argonaute 1 proteins
Authors: Crawshaw Sam , Watt Lewis G. , Murphy Alex M. , Carr John P.
Published: September 2023

Abstract/Summary

The cucumber mosaic virus (CMV) 2b protein is a potent counter-defense protein and symptom determinant that inhibits antiviral silencing by titration of short double-stranded RNAs. Expression of the 2b protein from the CMV Subgroup IA strain Fny-CMV in transgenicArabidopsis thalianaplants disrupts microRNA-mediated cleavage of host mRNAs by binding ARGONAUTE 1 (AGO1), leading to symptom-like phenotypes. This also triggers AGO2-mediated resistance against CMV and strong resistance to CMV’s aphid vectors, which would be deleterious to viral fitness. However, in authentic viral infections the Fny-CMV 1a protein modulates 2b-AGO1 interactions, which inhibits induction of AGO2-mediated virus resistance and resistance to aphid vectors. Contrastingly, the 2b proteins encoded by the Subgroup II LS-CMV strain or the recently discovered Subgroup IA strain Ho-CMV induce no apparent symptoms. Confocal laser scanning microscopy, bimolecular fluorescence complementation and co-immunoprecipitation showed that the Fny-CMV and Ho-CMV 2b proteins interact with the Fny-CMV and LS-CMV 1a proteins whilst the CMV-LS 2b protein does not. However, the Fny-CMV, Ho-CMV and LS-CMV 2b proteins all interacted with AGO1, but while AGO1-Fny2b complexes occurred in the host cell nucleus and cytoplasm, the corresponding AGO1-2b complexes for LS-CMV and Ho-CMV accumulated almost exclusively in nuclei.AGO2transcript accumulation was used to assess the inhibition of AGO1-mediated miRNA-regulated mRNA cleavage. While Fny-CMV 2b induced a five-fold increase inAGO2accumulation, the LS-CMV and Ho-CMV 2b proteins induced only two-fold increases. Thus, these 2b proteins bind AGO1 but are less effective at inhibiting AGO1 activity. We conclude that the intracellular localization sites of 2b-AGO1 complexes influences the degree to which a 2b protein can inhibit microRNA-mediated host mRNA degradation and that cytoplasmic AGO1 has the strongest influence on miRNA-mediated cellular mRNA turnover.
Single-cell transcriptomics reveals functional insights into a non-model aquatic phytoflagellate and its metabolically linked bacterial community
Authors: Florenza Javier , Jeevannavar Aditya , Divne Anna-Maria , Tamminen Manu , Bertilsson Stefan
Published: September 2023

Single-cell transcriptomics is a vital tool for unraveling metabolism and tissue diversity in model organisms. Its potential for elucidating the ecological roles of microeukaryotes, especially non-model ones, remains largely unexplored. This study employed the Smart-seq2 protocol onOchromonas triangulata, a microeukaryote lacking a reference genome, showcasing how transcriptional states align with growth phases. Unexpectedly, a third transcriptional state was identified, across both growth phases. Metabolic mapping revealed a down-regulation trend in pathways associated with ribosome functioning, CO2 fixation, and carbohydrate catabolism from fast to slow growth to the third transcriptional state. Using carry-over rRNA reads, taxonomic identity ofOchromonas triangulatawas re-confirmed and distinct bacterial communities associated with transcriptional states were identified. This study underscores single-cell transcriptomics as a powerful tool for characterizing metabolic states in microeukaryotes without a reference genome, offering insights into unknown physiological states and individual-level interactions with different bacterial taxa. This approach holds broad applicability for uncovering ecological roles, surpassing alternative methods like metagenomics or metatranscriptomics.
Leaf and shoot apical meristem transcriptomes of quinoa (Chenopodium quinoaWilld.) in response to photoperiod and plant development
Authors: Maldonado-Taipe Nathaly , Rey Elodie , Tester Mark , Jung Christian , Emrani Nazgol
Published: September 2023

Our study aimed to identify candidate genes for flowering time regulation and photoperiod response in quinoa. We investigated the timing of photoperiod-driven floral transition and analyzed the transcriptomes of leaf and shoot apical meristems in photoperiod-sensitive and -insensitive quinoa accessions. Histology analysis of the apical meristem showed that floral transition in quinoa initiates two to three weeks after sowing. We found four groups of differentially expressed genes responding to plant development and floral transition, which were annotated in the QQ74-V2 reference genome, including (i) 222 genes differentially responding to photoperiod in leaves, (ii) 1,812 genes differentially expressed between accessions under long-day conditions in leaves, (iii) 57 genes responding to developmental changes between weeks under short-day conditions in leaves, and (iv) 911 genes responding to floral transition within the shoot apical meristem. Interestingly, out of the thousands of candidates, two putativeFTorthologues and several others have been reported as key regulators of flowering time in other species (e.g.,SOC1,COL,AP1). Additionally, we used co-expression networks to associate novel transcripts to a putative biological process based on the annotated genes within the same co-expression cluster. The candidate genes in this study would benefit quinoa breeding by identifying and integrating their beneficial haplotypes in crossing programs to develop adapted cultivars to diverse environmental conditions.
An assessment of the prevalence and risk factors of hypertensive crisis in patients who visit the Emergency Outpatient Department (EOPD) at Adama Hospital Medical College, Adama, Oromia, Ethiopia: A 6-month prospective study
Authors: Abebe Abel Tezera , Kebede Yabets Tesfaye , Mohammed Bekri Delil
Published: September 2023

Background

Over 1 billion people worldwide suffer from the common chronic medical condition of hypertension. A hypertensive crisis occurs when blood pressure exceeds 180/110 mmHg. Depending on whether the target organ is harmed, the situation may be presented as a hypertensive emergency or urgency.

Objective

To assess the prevalence and risk factors of patients with hypertensive crises who visited the Emergency Outpatient Department (EOPD) at Adama Hospital Medical College in Adama, Oromia, Ethiopia, between January 01 and August 31, 2021 G.C.

Methodology

A cross sectional, prospective study on hypertensive crisis was conducted at Adama Hospital Medical College from January 01 to August 31, 2021 G.C. The data was collected using a standardized questionnaire, validated for completeness, and analyzed using SPSS. The findings were tabulated, and conclusions and recommendations were conveyed.

Result

A total of 444 individuals with hypertension in crisis were identified. Of these, 56.8% were men, resulting in a M:F ratio of 1.31:1. Those between the ages of 66 and 75 were the most affected. At presentation, 91.0% of the study participants were known hypertensive patients. Of the known hypertensive patients, the majority (34.9%) were known to have been hypertensive for 5 to 10 years. Of the known hypertensive patients, 48.6% were found to be adherent. Hypertensive urgency was discovered to be far more common than hypertensive emergencies (63.5% vs. 36.5%). Headache was the most common presenting symptom, and most of the patients (36.5%) presented to the health setup in less than 24 hours. The main risk variables identified were drug discontinuation, family history of hypertension, salt consumption, and alcohol usage. The main excuse for lack of adherence was the cost of the medications. More than half of the patients do not have any additional comorbidities, and of those who do, diabetes mellitus is the most prevalent. A stroke was identified as a major complication.

Conclusions and recommendations

Hypertensive crises are one of the most prevalent reasons for EOPD admission and are linked to significant consequences. At presentation, most of the study subjects were known hypertension patients. Diabetes mellitus was discovered to be a comorbid condition in one quarter of them. Although more than half of the patients have improved, the death rate remains high. Infrastructure and capacity building to provide hospitals with the requisite baseline investigations are among the government’s recommendations. Health practitioners are expected to make improvements, such as educating the public about the need for lifestyle changes and evaluating and managing any hypertension problems.
Learning the sequence code of protein expression in human immune cells
Authors: Nicolet Benoît P. , Jurgens Anouk P. , Bresser Kaspar , Guislain Aurélie , Bradariç Antonia , Wolkers Monika C.
Published: September 2023

Accurate protein expression in human immune cells is essential for appropriate cellular function. The mechanisms that define protein abundance are complex and executed on transcriptional, post-transcriptional and post-translational level. Here, we present SONAR, a machine learning pipeline that learns the endogenous sequence code and that defines protein abundance in human cells. SONAR uses thousands of sequence features (SFs) to predict up to 63% of the protein abundance independently of promoter or enhancer information. SONAR uncovered the cell type-specific and activation-dependent usage of SFs. The deep knowledge of SONAR provides a map of biologically active SFs, which can be leveraged to manipulate the amplitude, timing, and cell type-specificity of protein expression. SONAR informed on the design of enhancer sequences to boost T cell receptor expression and to potentiate T cell function. Beyond providing fundamental insights in the regulation of protein expression, our study thus offers novel means to improve therapeutic and biotechnology applications.

One Sentence Summary

SONAR informs the design of cell type-specific protein expression in human cells